K values of NPX-CyAA-Pyr dyad diastereomers (column 2) determined for the different ratios of diastereomer concentrations (column 1).
\r\n\t
",isbn:"978-1-83969-150-8",printIsbn:"978-1-83969-149-2",pdfIsbn:"978-1-83969-151-5",doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,hash:"7409b2acd5150a93004300800918b736",bookSignature:"Prof. Karmen Pažek",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/10548.jpg",keywords:"Lean Manufacturing, Agriculture, Production and Process, Costs Reduction, Lean Principles, Industry, Tools, Implementation, Sustainability, Modeling, Environment, Planning",numberOfDownloads:11,numberOfWosCitations:0,numberOfCrossrefCitations:0,numberOfDimensionsCitations:0,numberOfTotalCitations:0,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"October 20th 2020",dateEndSecondStepPublish:"November 17th 2020",dateEndThirdStepPublish:"January 16th 2021",dateEndFourthStepPublish:"April 6th 2021",dateEndFifthStepPublish:"June 5th 2021",remainingDaysToSecondStep:"2 months",secondStepPassed:!0,currentStepOfPublishingProcess:4,editedByType:null,kuFlag:!1,biosketch:"Dr. Pažek is Head of the undergraduate study program Agricultural economics and rural development and Vice-dean for education. She is the author or co-author of 61 scientific papers, 6 scientific books, and 24 book chapters.",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"179642",title:"Prof.",name:"Karmen",middleName:null,surname:"Pažek",slug:"karmen-pazek",fullName:"Karmen Pažek",profilePictureURL:"https://mts.intechopen.com/storage/users/179642/images/system/179642.jpg",biography:"Karmen Pažek achieved her Ph.D. at University of Maribor, Faculty of Agriculture in 2006. She is active as Full Professor for Farm management in the Department for Agriculture Economics and Rural Development on Faculty of Agriculture and Life Sciences, University of Maribor. Her research includes development of decision support tools and systems for farm management (simulation modeling, multi-criteria decision analysis, option models, investment analysis) and economics of agricultural production. She is involved in teaching activities as thesis supervisor at postgraduate study programs and involved in national and international research projects. She is author or coauthor of 61 scientific papers (including 34 papers in journals with impact factor), 6 scientific books and 24 book chapters. Currently she is Head of the undergraduate study program Agricultural economics and rural development and Vice dean for education. \r\n\r\nAcademic activities\r\nResearch:\r\n-\tFarm management\r\n-\tDecision support, simulation, forecasting, multi criteria decision making in the area of agriculture with emphasis on field crops, farm tourism and fruit producon\r\n\r\nCurrent Research work:\r\n- Financial parameters assessment based on perfect and in-perfect information in agrifood \r\n systems \r\n- Option modeling of agrifood projects\r\n-\tEfficiency assessment in farm tourism \r\n-\tEfficiency of sugar beet production systems \r\n\r\nTeaching:\r\nUndergraduate Programmes and Courses\r\n-\tFarm management I and II\r\n-\tIntroduction to decision theory\r\n-\tOrganic fam management\r\n-\tManagement od supplementary activities\r\n-\tEconomics and management of rural tourism\r\n-\tSelected issues in agricultural entrepreneurship\r\n\r\nMaster Programmes and Courses\r\n\r\n-\tResearch methods in farm management\r\n-\tDecision theory\r\n-\tProject planning and quality management\r\n-\tOrganic fam management\r\n\r\n \r\nPhD Programme and Course\r\n\r\n-\tProject management (transferable skills)\r\n-\tSelected issues in farm management",institutionString:"University of Maribor",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"3",totalChapterViews:"0",totalEditedBooks:"0",institution:{name:"University of Maribor",institutionURL:null,country:{name:"Slovenia"}}}],coeditorOne:null,coeditorTwo:null,coeditorThree:null,coeditorFour:null,coeditorFive:null,topics:[{id:"11",title:"Engineering",slug:"engineering"}],chapters:[{id:"74769",title:"Development of Integrated Lean Six Sigma-Baldrige Framework for Manufacturing Waste Minimization: A Case of NAS Foods Plc",slug:"development-of-integrated-lean-six-sigma-baldrige-framework-for-manufacturing-waste-minimization-a-c",totalDownloads:16,totalCrossrefCites:0,authors:[null]}],productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"},personalPublishingAssistant:{id:"247865",firstName:"Jasna",lastName:"Bozic",middleName:null,title:"Ms.",imageUrl:"https://mts.intechopen.com/storage/users/247865/images/7225_n.jpg",email:"jasna.b@intechopen.com",biography:"As an Author Service Manager, my responsibilities include monitoring and facilitating all publishing activities for authors and editors. From chapter submission and review to approval and revision, copyediting and design, until final publication, I work closely with authors and editors to ensure a simple and easy publishing process. I maintain constant and effective communication with authors, editors and reviewers, which allows for a level of personal support that enables contributors to fully commit and concentrate on the chapters they are writing, editing, or reviewing. I assist authors in the preparation of their full chapter submissions and track important deadlines and ensure they are met. I help to coordinate internal processes such as linguistic review, and monitor the technical aspects of the process. As an ASM I am also involved in the acquisition of editors. Whether that be identifying an exceptional author and proposing an editorship collaboration, or contacting researchers who would like the opportunity to work with IntechOpen, I establish and help manage author and editor acquisition and contact."}},relatedBooks:[{type:"book",id:"1591",title:"Infrared Spectroscopy",subtitle:"Materials Science, Engineering and Technology",isOpenForSubmission:!1,hash:"99b4b7b71a8caeb693ed762b40b017f4",slug:"infrared-spectroscopy-materials-science-engineering-and-technology",bookSignature:"Theophile Theophanides",coverURL:"https://cdn.intechopen.com/books/images_new/1591.jpg",editedByType:"Edited by",editors:[{id:"37194",title:"Dr.",name:"Theophanides",surname:"Theophile",slug:"theophanides-theophile",fullName:"Theophanides Theophile"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"3092",title:"Anopheles mosquitoes",subtitle:"New insights into malaria vectors",isOpenForSubmission:!1,hash:"c9e622485316d5e296288bf24d2b0d64",slug:"anopheles-mosquitoes-new-insights-into-malaria-vectors",bookSignature:"Sylvie Manguin",coverURL:"https://cdn.intechopen.com/books/images_new/3092.jpg",editedByType:"Edited by",editors:[{id:"50017",title:"Prof.",name:"Sylvie",surname:"Manguin",slug:"sylvie-manguin",fullName:"Sylvie Manguin"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"3161",title:"Frontiers in Guided Wave Optics and Optoelectronics",subtitle:null,isOpenForSubmission:!1,hash:"deb44e9c99f82bbce1083abea743146c",slug:"frontiers-in-guided-wave-optics-and-optoelectronics",bookSignature:"Bishnu Pal",coverURL:"https://cdn.intechopen.com/books/images_new/3161.jpg",editedByType:"Edited by",editors:[{id:"4782",title:"Prof.",name:"Bishnu",surname:"Pal",slug:"bishnu-pal",fullName:"Bishnu Pal"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"72",title:"Ionic Liquids",subtitle:"Theory, Properties, New Approaches",isOpenForSubmission:!1,hash:"d94ffa3cfa10505e3b1d676d46fcd3f5",slug:"ionic-liquids-theory-properties-new-approaches",bookSignature:"Alexander Kokorin",coverURL:"https://cdn.intechopen.com/books/images_new/72.jpg",editedByType:"Edited by",editors:[{id:"19816",title:"Prof.",name:"Alexander",surname:"Kokorin",slug:"alexander-kokorin",fullName:"Alexander Kokorin"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"1373",title:"Ionic Liquids",subtitle:"Applications and Perspectives",isOpenForSubmission:!1,hash:"5e9ae5ae9167cde4b344e499a792c41c",slug:"ionic-liquids-applications-and-perspectives",bookSignature:"Alexander Kokorin",coverURL:"https://cdn.intechopen.com/books/images_new/1373.jpg",editedByType:"Edited by",editors:[{id:"19816",title:"Prof.",name:"Alexander",surname:"Kokorin",slug:"alexander-kokorin",fullName:"Alexander Kokorin"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"57",title:"Physics and Applications of Graphene",subtitle:"Experiments",isOpenForSubmission:!1,hash:"0e6622a71cf4f02f45bfdd5691e1189a",slug:"physics-and-applications-of-graphene-experiments",bookSignature:"Sergey Mikhailov",coverURL:"https://cdn.intechopen.com/books/images_new/57.jpg",editedByType:"Edited by",editors:[{id:"16042",title:"Dr.",name:"Sergey",surname:"Mikhailov",slug:"sergey-mikhailov",fullName:"Sergey Mikhailov"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"371",title:"Abiotic Stress in Plants",subtitle:"Mechanisms and Adaptations",isOpenForSubmission:!1,hash:"588466f487e307619849d72389178a74",slug:"abiotic-stress-in-plants-mechanisms-and-adaptations",bookSignature:"Arun Shanker and B. Venkateswarlu",coverURL:"https://cdn.intechopen.com/books/images_new/371.jpg",editedByType:"Edited by",editors:[{id:"58592",title:"Dr.",name:"Arun",surname:"Shanker",slug:"arun-shanker",fullName:"Arun Shanker"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"878",title:"Phytochemicals",subtitle:"A Global Perspective of Their Role in Nutrition and Health",isOpenForSubmission:!1,hash:"ec77671f63975ef2d16192897deb6835",slug:"phytochemicals-a-global-perspective-of-their-role-in-nutrition-and-health",bookSignature:"Venketeshwer Rao",coverURL:"https://cdn.intechopen.com/books/images_new/878.jpg",editedByType:"Edited by",editors:[{id:"82663",title:"Dr.",name:"Venketeshwer",surname:"Rao",slug:"venketeshwer-rao",fullName:"Venketeshwer Rao"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"4816",title:"Face Recognition",subtitle:null,isOpenForSubmission:!1,hash:"146063b5359146b7718ea86bad47c8eb",slug:"face_recognition",bookSignature:"Kresimir Delac and Mislav Grgic",coverURL:"https://cdn.intechopen.com/books/images_new/4816.jpg",editedByType:"Edited by",editors:[{id:"528",title:"Dr.",name:"Kresimir",surname:"Delac",slug:"kresimir-delac",fullName:"Kresimir Delac"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"3621",title:"Silver Nanoparticles",subtitle:null,isOpenForSubmission:!1,hash:null,slug:"silver-nanoparticles",bookSignature:"David Pozo Perez",coverURL:"https://cdn.intechopen.com/books/images_new/3621.jpg",editedByType:"Edited by",editors:[{id:"6667",title:"Dr.",name:"David",surname:"Pozo",slug:"david-pozo",fullName:"David Pozo"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},chapter:{item:{type:"chapter",id:"64898",title:"Peculiarities of Electron Transfer in Chiral Linked Systems",doi:"10.5772/intechopen.82684",slug:"peculiarities-of-electron-transfer-in-chiral-linked-systems",body:'\nChiral systems are in the top of interest for a long time to this day. From the chemical point of view, the most interesting are the following issues: sources of the chiral biomolecules appearance in prebiological period, and physicochemical reasons of the difference in biological and medical activity of enantiomers. It is a matter of debate up to now [1]. On the both directions, significant progress has been made in the last decade of the twentieth century and the beginning of the twenty-first century. This review will focus on elementary ET process in chiral systems. These systems are donor-acceptor dyads with two chiral centers studied by spin chemistry and photochemistry methods. Similar dyads are frequently used to simulate the binding of chiral drugs with receptors or enzymes [2, 3]. They are diastereomers, where one partner is a drug enantiomer. Such systems attract attention since drug enantiomers demonstrate different, often just opposite, medical activity. It is a practical problem of pharmacology and medicine [1]. Despite this, numerous biochemical studies did not give a definite answer about the reasons for the difference in enantiomer medical activities.
\nBecause enantiomers have identical physicochemical properties, there are no physical reasons of the difference in reactivity. But in active sites of enzymes and receptors, chiral enantiomers interact with chiral amino acid residues, and it lets us expect that drug-enzyme or drug-receptor complexes will be similar to diastereomers. If binding process includes stages with charge transfer, the ET in dyads would be a convenient model to simulate drug-receptor or drug-enzyme interaction. Suggested approach is based on two major assumptions. First, it is suggested that the reactivity of active intermediates would not depend on their generation pathway. Second, using photoirradiation instead of enzymatic one for the studied processes, activation is possible since an activation barrier is decreased as a result of increasing of drug’s oxidation potential in its excited state [3].
\nET in chiral systems is studied by the example of nonsteroidal anti-inflammatory drugs (NSAIDs), medicines that demonstrate remarkable difference in enantiomers medical activity [1, 4]. PET was studied by the UV irradiation of dyads including NSAIDs linked by bridges with various electron donors [5, 6, 7, 8, 9, 10]. The exploration by using fluorescence and laser flash photolysis has shown that ET in dyads is stereoselective.
\nThe application of photochemistry and CIDNP methods to study dyads with known representative NSAID naproxen (NPX) and donors N-methylpyrrolidine (Pyr) and tryptophan (Trp) has shown that, under the UV irradiation, partial and full charge transfer occur. Both processes demonstrate stereoselectivity [3, 11, 12, 13, 14].
\nThis difference in reactivity of (S)- and (R)-NPX isomers, which are part of diastereomers, is in qualitative agreement with some results of biochemical research [3]. So, (R)-isomer is more active in cytochrome P-450-induced metabolism that involves stage of ET. At that, indeed, the rate constant of the local excited state transition to a state with partial charge transfer systematically is greater for (R,S)-diastereomers. If we imagine that the process of partial charge transfer leading to exciplex formation can simulate the binding process, then large quantum yields of exciplex fluorescence for (S,S)-diastereomers compared with (R,S)- will mean the better binding of (S)-NPX with a donor.
\nThe following peculiarity of ET process in chiral systems is spin selectivity [15]. Spin selectivity means the difference in CIDNP effects of dyad diastereomers, shown by the example of two dyads: NPX linked by a rigid bridge with the donor Pyr, and NPX-Trp dyad. Spin selectivity appearance lets us assume the difference in electron density distribution of (R,S)- and (S,S)-dyad diastereomers.
\nThus, the above results have shown that full ET and partial charge transfer processes play a significant role in understanding of chiral systems reactivity.
\nFurther, this chapter will be devoted to the description of two features of ET in chiral systems: stereo- and spin selectivity and the influence of environment on these peculiarities.
\nIn this part, we will discuss the appearance of stereoselectivity of processes with PET in the linked systems with different NSAIDs.
\nPhotoinduced interactions of (S)-/(R)-flurbiprofen (FBP) and thymidine (dThd) were studied in covalent-linked systems in work [5] (for structures, see Figure 1). Under the illumination of (R)-FBP-dThd and (S)-FBP-dThd solutions in aerated acetonitrile, chromophore 1FBP* was excited and its fluorescence quantum yield (φfl) and lifetime (τfl) values were smaller than those of free FBP in solution. Authors suggested this dynamical quenching carries out through ET (where FBP is a donor) or exciplex formation. This theory is supported by Gibbs energies (ΔG) estimation in solutions of different polarity (acetonitrile and dioxane) by using classical Rehm-Weller-Zacharias criterion [16]. This criterion application has demonstrated that both partial and full charge transfer are possible in acetonitrile but exciplex formation is prevailing in dioxane. Fluorescence quantum yields and lifetimes have shown stereoselectivity of these processes—for (R)-isomer, φfl and τfl (0.015 and 0.27 ns) were smaller than for (S)- (0.018 and 0.31 ns).
\nChemical structures of FBP-dThd dyads.
For systems with amino acid Trp (structures in Figure 2), stereoselectivity of quenching process was also demonstrated. In this case, the excitation wavelength 266 nm was chosen where about 60% of light is absorbed by FBP and 40% by Trp, respectively. Significant fluorescence quenching was detected, and residual spectrum was referred to Trp chromophore. Also wide exciplex band was registered (380–500 nm), and its lifetimes for (R,S)- and (S,S)-isomers revealed the biggest stereoselectivity—5.1 and 7.6 ns (in N2 bubbled acetonitrile solutions), respectively. As for fluorescence quantum yields and lifetimes of the main spectrum band attributed to the dyad excited state, the values were φfl (R,S) = 0.02, φfl (S,S) = 0.04; τfl (R,S and S,S) = 0.9 ns. Possibility of electron transfer and exciplex formation was checked by ΔG values estimation (using Rehm-Weller-Zacharias equation [16]), in both cases processes turned out to be exergonic [6].
\nChemical structures of (R)-/(S)-FBP-Trp dyads.
For systems with amino acid tyrosine, some stereoselectivity of photoinduced charge transfer was also found and, in this case, main attention was focused on the influence of spacer between partners [7]. One system FBP-Tyr had flurbiprofen and tyrosine connected directly, and another one had them separated by a cyclic spacer (see Figure 3).
\nChemical structures of FBP-Tyr (top) and FBP-CyAA-Tyr (bottom).
As in previous flurbiprofen systems, these systems were studied by means of time-resolved fluorescence and flash-photolysis spectroscopy in acetonitrile solutions. Significant fluorescence quenching was found for both systems with short and cyclic bridge. It was attributed to electron transfer. Expectedly, this process was more efficient for the system with short bridge. Fluorescence quantum yields were 0.04 and 0.09 (for (R,S)- and (S,S)-FBP-Tyr, respectively), 0.15 and 0.11 (for (R,S)- and (S,S)-FBP-CyAA-Tyr). Average fluorescence lifetimes were 0.36 and 0.9 ns (for (R,S)- and (S,S)-FBP-Tyr, respectively), 1.2 and 0.93 ns (for (R,S)- and (S,S)-FBP-CyAA-Tyr). Interestingly, that for systems with a cyclic spacer exciplex formation was registered in fluorescence spectra and its lifetimes were 3.05 ± 0.01 and 3.27 ± 0.01 ns for (R,S)- and (S,S)-FBP-CyAA-Tyr, respectively. Possibility of electron transfer and exciplex formation was confirmed by authors with ΔG values estimation (by using Rehm-Weller-Zacharias equation [16]). Even more stereodifferentiation was observed for these systems triplet state: 0.14 and 0.27 (for (R,S) and (S,S)-FBP-Tyr), 0.39 and 0.22 (for (R,S) and (S,S)-FBP-CyAA-Tyr). No stereoselectivity was detected at early times, only after about 10 ps. The kinetics of (R,S)-FBP-Tyr was found to decay faster than its (S,S)-analogue after 10 ps; on the contrary, for the dyads with rigid bridge, the opposite behavior was observed [8]. These differences were analyzed in view of conformational peculiarities of studied systems. Computations have shown that geometrical arrangement of FBP and Tyr was favored for donor-acceptor interaction in (R,S) with short bridge due to its folded conformation, which was in agreement with fluorescence data. And geometry of this dyad (S,S)-configuration was more distorted, and fluorescence quenching was less efficient. Concerning dyads with rigid bridge, authors claimed more unfavorable geometry for interaction, especially for the (R,S)-diastereomer, where the two chromophores are practically in orthogonal arrangement. That is also in line with (R,S)- lower fluorescence quenching compared to (S,S)-FBP-CyAA-Tyr. Exciplex formation was not analyzed in these computations.
\nChiral systems photoinduced interactions with amino acids were also studied in work [9]. In this case, an acceptor role is played by another NSAID—carprofen (CPF-Trp, see systems structure in Figure 4) and a donor amino acid was tryptophan (in dyad and in human serum albumin).
\nChemical structures of dyads (R)-/(S)-CPF-Trp.
In the presence of albumin in the solution of CPF, similar transient absorption spectra were observed for both CPF stereoisomers. However, time-resolved measurements have shown significant difference—for each isomer, triplet state quenching profile had two components with different lifetimes. Authors corresponded these times to CPF complexation with two possible albumin bonding sites (site I and site II). This mapping was confirmed in experiments with another NSAID—ibuprofen as a substitute in site II. Therefore, shorter components, which had shown more significant stereodifferentiation (τR/τS~4), were associated with CPF triplet state in site I; and lifetime shortening authors linked with excitation quenching by charge transfer from only one possible partner in protein—Trp amino acid residue. Laser flash photolysis study of model linked system—CPF-Trp dyad (Figure 4) confirmed this idea. Trp cation-radical was registered (with absorption maximum at ~580 nm); this is the evidence of electron transfer mechanism. And triplet lifetimes of model systems demonstrated stereodifferentiation—τRS = 2.4 μs and τSS = 3.0 μs. Interestingly, that in this case, in opposite to all flurbiprofen-contained dyads, no reactivity was observed for singlet excited state—all fluorescence quantum yields and lifetimes were equal to carprofen itself φfl = 0.068 and τfl = 1.55 ns.
\nAnother study involving drug in complex with α1-acid glycoprotein (BAAG) was performed by photophysical methods [10]. Laser flash photolysis in solutions of BAAG/(S)-CPFMe (methyl ester of carprofen) and BAAG/(R)-CPFMe in proportion 2:1 was performed and decay traces have shown stereodifferentiation. Thus, the triplet lifetime value for the (S)-enantiomer was much shorter (13 μs) than that of (R)-analogue (18 μs). Authors proposed that this could be related to the more efficient quenching of triplet excited state 3CPFMe* by electron transfer from Trp in the case of (S)-CPFMe, which is in agreement with a closer distance from (S)-CPFMe to a Trp residue than in case of (R)-CPFMe, within the binding site pocket [9].
\nNSAID NPX as a part of linked systems with electron donors was studied in many works. Interestingly, system with NPX was dyad NPX-oxetane (OXT) (Figure 5) [17]. Illumination of systems in acetonitrile and chloroform solutions at NPX absorption wavelength (300 nm) leaded to ring opening.
\nLinked systems structure: left—(S)-NPX-(S,R,S)-OXT, right—(S)-NPX-(R,S,R)-OXT.
Authors explained this reaction mechanism also by involving photoinduced electron transfer (reaction scheme is presented in Figure 6).
\nScheme of photoinduced ring opening in systems (S)-NPX-(S,R,S)-OXT and (S)-NPX-(R,S,R)-OXT.
Interestingly, that high reactivity was observed in acetonitrile but high stereoselectivity was in chloroform (τS,R,S/τR,S,R ~ 1.5). Allegedly, this is because of twisted conformations that prevail in dyad (S)-diastereomer. In this conformation, naphthalene fragment oriented to oxetane that favors intramolecular electron transfer [17].
\nMore detailed investigation of processes with partial and full charge transfer in linked systems was performed by examples of several dyads from Figure 7 [3, 11, 12, 13, 14].
\nLinked system structures: NPX-Pyr—“short,” NPX-AA-Pyr—“flexible,” NPX-CyAA-Pyr—“rigid,” and NPX-Trp—“tryptophan dyads.”.
In this case, besides using time-resolved fluorescence measurement, CIDNP method has been applied to detect the state with full charge transfer between donor and acceptor—biradical-zwitterion (BZ) formed as a result of intramolecular ET.
\nCIDNP is phenomenon that manifested as unusual NMR signals, with the population of nuclear spins different from Boltzmann population. These signals are observed in the products of radical reaction, carried out directly in the probe of NMR spectrometer. There are singlets and multiplets with enhanced absorption or emission existing for a short time comparable with time of nuclear relaxation. These effects are the result of weak magnetic interactions in the radical pair (RP), which is a precursor of products. In essence, CIDNP, formed in high magnetic field of NMR spectrometer, reflects the difference in the recombination probability of RPs with αN and βN nuclear spin projections on the magnetic field direction. It depends on hyperfine coupling constants, the difference in g-factors of two radicals in RP, multiplicity of the RP precursors, the type of reaction, and some other parameters [18]. Today, CIDNP is the most direct method for detecting paramagnetic precursors of radical reaction products.
\nThe formation of BZ under the UV irradiation of dyads pictured in Figure 7 was detected using CIDNP [3]. In accordance with spin density distribution in BZ, maximal CIDNP effect demonstrates methyl protons of N-methylpyrrolidine group (Figure 8).
\nNMR and CIDNP spectra detected after laser irradiation of (S,S)-NPX-Pyr “short” dyad in CD3CN; 1—N-CH3, 2—CH, 3—CH2 signals of α protons in N-centered radical-cation of N-methylpyrrolidine fragment.
The same CIDNP effects have demonstrated all other dyads with donor N-methylpyrrolidine. Unlike the mentioned dyads, diastereomers of dyad NPX-Trp have shown CIDNP of both aromatic and aliphatic protons (Figure 9).
\nNMR and pseudo steady state (PSS) CIDNP spectra of dyad’s NPX-Trp solutions (5 mM in 40% C6D6, 0.17% H2O, the rest is CD3CN): (R,S)-top, (S,S)-bottom. Reproduced with permission from Ref. [11] © John Wiley & Sons, Ltd., 2018.
The reason of the distinction of CIDNP pattern in dyads with N-methylpyrrolidine and Trp relates with the difference in the ratios of HFI constants in paramagnetic centers of donor and acceptor. In the case of NPX-Trp dyad, HFI constants of both fragments are the same order of magnitude, so CIDNP effects are appeared on all groups of protons. As it was mentioned above, in BZs of dyads with Pyr HFI constants of α protons in pyrrolidine fragment prevail; therefore, they determine the singlet-triplet evolution in RP.
\nThus, CIDNP spectra in all studied dyads point at NPX excited state quenching via ET, followed by the formation of BZ (Scheme 1). CIDNP signs analysis carried out according to the known rules of R. Kaptein modified by G. Closs showed that the singlet excited state of dyads is quenched by an electron donor in intramolecular process [18]. An intramolecular ET is favored since a difference in order of magnitude between the rates of monomolecular and bimolecular quenching: Wintra = 2–8 × 108 s−1 and Winter = 2 × 109 × 5 × 10−3 = 107 M−1 s−1 [11, 12, 14].
\nMechanism of NPX-Pyr singlet excited state quenching.
Well-known criterion of Rehm-Weller-Zacharias points at the possibility of the back ET from both singlet and triplet spin states of the BZ [12, 16]. According to this criterion, ET for studied systems is possible if the value of free energy (ΔG) is negative ΔG = [Eox(D/D+) – Ered(A−/A)]ε0 – e/εa + ΔGsolv – E(1,3D*). Here ΔGsolv = e2/2(1/RD+ + 1/RA−) × (1/ε – 1/ε0), Eox (Pyr) = 1.0 eV, Ered (2-methoxynaphthalene) = −2.6 eV, and E0–0(NPX) = 3.69 eV.
\nSince for NPX ET = 3.25 eV and ΔH in BZ is equal to Eox(D/D+) – Ered(A−/A)]ε0 = 3.6 eV, then NPX in excited triplet state can be formed via the back ET in BZ of triplet collective spin state.
\nThus, analysis of the CIDNP, detected upon the UV irradiation of studied dyads, allows one to confirm ET between (S)- and (R)-NPX and donors. Note, in the case of dyads with Pyr, CIDNP analysis using Kaptein rule [18] has shown that back ET occurs predominantly from the singlet collective spin state of BZ, whereas for the NPX-Trp dyad, it occurs mainly from the triplet spin state of the BZ [14].
\nAccording to the calculated curves in Figure 10, the experimental CIDNP dependences on dielectric constant in both cases of partial and full charge transfer should be smooth curves without extremes.
\nDependence of free energies of the radical-ion pair of N-methylpyrrolidine radical-cation and methoxynaphthalene radical-anion on solvent permittivity, calculated using Rehm-Weller-Zacharias equation and dependences of CIDNP on solvent permittivity for short, flexible, and rigid dyads (bottom). Reproduced from Ref. [3] with permission from the PCCP Owner Societies.
However, the experimental dependences of CIDNP for dyads with N-methylpyrrolidine on the dielectric constant are curves with maxima position, depending on the bridge length. Such shapes mean that the CIDNP effects are formed in at least two processes, and their contributions depend on the permittivity and dyad structures. Therefore, the resulting polarization has to be formed from the back ET in both singlet and triplet collective spin states of the BZ. According to CIDNP signs analysis, the rate constant of back ET in BZ singlet spin state (kS) has to be higher than that for triplet state (kT). Meanwhile, there are reference data informing that in the pair of amine radical-cation and naphthalene radical-anion, kT value is higher than kS [11, 12]. It is reasonable to assume that the predominance of kS is related to the formation of a singlet exciplex, being in equilibrium with the BZ. The detailed mechanism of NPX singlet excited state quenching by the N-methylpyrrolidine, provided on the basis of CIDNP analysis, is shown in Scheme 1.
\nSo, CIDNP analysis points that NPX excited state quenching in above dyads goes through stages of full and partial charge transfer. Details of the processes of partial charge transfer: rate constants of local excited state (LES) and exciplex accumulation and decay and exciplex fluorescence quantum yields were obtained from fluorescence measurements, including time-resolved experiments.
\nThe fluorescence spectra of dyad diastereomers with short, flexible, and rigid bridges compared with NPX methyl ether in acetonitrile are presented in Figure 11.
\nFluorescence spectra of (S)-NPX-OMe (1), (R,S)-NPX-Pyr (2a), and (S,S)-NPX-Pyr (2b), (R,S)-NPX-AA-Pyr (3a), (S,S)-NPX-AA-Pyr (3b), (R,S)-NPX-CyAA-Pyr (4a), and (S,S)-NPX-AA-Pyr (4b) in acetonitrile, excitation wavelength 320 nm. Magnified long-wavelength bands of exciplex are shown in insert.
There are two emission bands; they refer to local excited state of NPX moiety (350 nm) and exciplex (520 nm). The decreasing of dyad fluorescence intensity was accounted for new quenching channel—transition from LES to exciplex [13]. Note that LES fluorescence quantum yield is higher for dyad with flexible bridge, whereas exciplex fluorescence quantum yield is higher for dyad with rigid bridge. At that, fluorescence decay traces of both LES and exciplex were approximated by two exponential models with short and long lifetimes for LES and rise and decay times for exciplex. Fluorescence spectra and kinetics of its quenching were investigated in solvents with different dielectric constants (Figure 12).
\nDependences of fluorescence lifetimes for NPX-AA-Pyr dyad (left) and NPX-CyAA-Pyr (right). LES—squares (τshort, τlong), exciplex—circles (τrise, τfall), solid—(R,S)-diastereomers, open—(S,S)-diastereomers. Reproduced from Ref. [3] with permission from the PCCP Owner Societies.
Careful analysis of fluorescence data allows authors [3] to suggest the existence of two fast equilibria: “exciplex—LES” and “exciplex—BZ”. In this work, the summary scheme of NPX chromophore quenching by electron donor was proposed (Scheme 2).
\nMechanism of NPX chromophore quenching in dyads with flexible and rigid bridges. Dashed lines denote the dependence of LES and exciplex energy levels on solvent polarity. Reproduced from Ref. [3] with permission from the PCCP Owner Societies.
In this figure, two different dyad conformations in ground state, namely, expanded and folded, transfer to LES and exciplex through pathways W1 and W2, respectively. The development of this mechanism is based on the kinetic curves fitting, considering that LES and the exciplex are formed simultaneously. According to Scheme 2, the exciplex exists in dynamic equilibrium with LES (k4, k5) and BZ (k7, k8). BZ, in turn, can be in singlet or triplet spin states that have equal energy. Spin conversion (kS-T) occurs under the magnetic interactions in BZ. Back ET from both BZ spin states leads to the formation of parent dyad in singlet ground state and in triplet excited state (with corresponding rate constants kS and kT). Emission of LES and exciplex is determined by k3 and k6 rate constants. Exciplex also undergoes internal conversion with rate constant kisc(exc).
\nThe kinetics of LES and exciplex fluorescence were calculated by solution of differential equation system using Runge-Kutta method [3], to obtain rate constants of separate stages. Thus, the values of quenching rate constants (k3–k7) were obtained in [3]. It is remarkable that there is a difference between the values of diastereomers rate constants, in particular k4, k7, related with charge transfer processes (see Figure 13).
\nDependences of k4, k7 rate constants on solvent polarity for three dyads linked with N-methylpyrrolidine. Reproduced from Ref. [3] with permission from the PCCP Owner Societies.
The greatest difference between the values of diastereomers k7 (exciplex transformation into BZ) is observed only in polar media. At that, the constant k4 (transition from LES to exciplex) is higher for (R,S)-diastereomers of all three dyads.
\nNote that NPX-Pyr diastereomers were previously investigated by using fluorescence technique in work [19], where stereoselectivity was also revealed (τfl (RS) = 2.35 ns, τfl (SS) = 3.02 ns, rate constants of ET kRS = 2.8 × 108 s−1, and kSS = 1.8 × 108 s−1).
\nThus, the appearance of stereoselectivity in ET was shown for a number of dyads. In particular, the stereoselectivity of the rate constants of separate stages of NPX excited state quenching was demonstrated by the example of dyads with donor N-methylpyrrolidine. The biggest difference is shown by rate constants of LES transfer to exciplex for (R,S)- and (S,S)-diastereomers of dyads with short bridge. The similar results were obtained for exciplex fluorescence quantum yields measured in solvents with different dielectric constants. So, the stereoselectivity in the systems under study significantly depends on the distance between the chiral centers and the structure of the bridge.
\nThe following peculiarity of ET in chiral systems is spin selectivity. Spin selectivity implies the difference between CIDNP effects of dyad diastereomers, shown by the example of two NPX-based dyads: linked by rigid bridge with donor N-methylpyrrolidine, and linked with amino acid Trp (Figure 7).
\nThis part of the chapter focuses on the application of CIDNP to study the difference of spin density distribution in diastereomeric BZs of abovementioned two dyads. For this purpose, in works [14, 15], the CIDNP effects in diastereomers of these dyads were compared.
\nFigure 14 represents the difference in the intensity of polarized signals of N-methyl protons in diastereomers of NPX-Pyr dyad.
\nNMR and pseudo-steady-state (PSS) CIDNP spectra of NPX-CyAA-Pyr dyad diastereomers (5 mM) solution in CD3CN (H2O 0.05%): (S,S)—top, (R,S)—bottom. Negative polarized line situated at 2.3 ppm (red arrows) refers to N-methyl protons of dyad diastereomers. Reproduced with permission from Ref. [15] © John Wiley & Sons, Ltd., 2018.
Diastereomers of NPX-Trp dyad have demonstrated even greater distinction in CIDNP effects (see Figure 8, Section 2.3).
\nTo study in detail the nature of differences in the CIDNP effects formed in BZs of these dyads, in work [15], so-called CIDNP enhancement coefficients per one paramagnetic particle were used.
\nFor determination of the CIDNP enhancement coefficients per one paramagnetic particle, ratios of the intensities of polarized and equilibrium signals in the NMR spectra were divided by BZ concentrations, estimated from fluorescence data [3]. It should be noted that the extinction coefficients for two diastereomers are the same. Further, we will operate only with the ratio of enhancement coefficients determined as follows:
As can be seen from figures, the CIDNP enhancement coefficients are appreciably higher for the (R,S)-diastereomers and the maximum ratio of enhancement coefficients, K, varies from 2.1 to 2.3 for both dyads. Since this is a polarization per one paramagnetic particle, the difference in the coefficients is certain to be the result of the distinctions in the magnetoresonance parameters of the (R,S)- and (S,S)-BZs. CIDNP effects for the diastereomers of NPX-CyAA-Pyr dyad were compared in acetonitrile, because in this case the contribution from exciplex to CIDNP effects is negligible and K values would be mainly determined by the magnetoresonance parameters of BZ (see Scheme 1). Further, to understand the origin of the differences in CIDNP enhancement coefficients in dyad diastereomers, these coefficients were calculated by varying the magnetoresonance parameters and lifetimes of BZ for NPX-CyAA-Pyr diastereomers. The calculation was carried out in the frame of radical pair theory by using two-position model and Green function method [15, 18]. According to this model, two states of the system are available. One is the state of a direct contact; the radicals are located in the reaction zone with characteristic time τr. The other is the state where the radicals are out of the reaction zone. The overall system lifetime is τc. Time that the system spends in the reaction zone (τr) is much less than the total lifetime of the system (τc). Calculations of CIDNP dependence on the HFI constant under various values of Δg and characteristic times (τc) were carried out. Next, authors tried to trace what variations of the abovementioned parameters could give the required adjustment—a twofold change in the CIDNP effects of diastereomers. The dependence of CIDNP on the hyperfine coupling constant is known to be bell-shaped for long-lived radical ion pairs [18]. The location of the dependence extremum is defined by the ratio of Δg and the hyperfine interaction constant. In the studied systems, we used the following BZ magnetoresonance parameters: τc = 10 ns, whereas the experimental values are 7 and 9 ns for (R,S)- and (S,S)-diastereomers of NPX-CyAA-Pyr dyad, τr = 1 ns, and ∆g = 10−3 [3, 20]. The latter is also the experimental value. From a comparison of the listed parameters, and by taking into account the bell-shaped CIDNP dependence, one can draw the following conclusion: because 2ΔgH0 values are larger than hyperfine coupling constant values, greater polarization will correspond to a larger HFI constant. However, quantitative calculation can be carried out only for NPX-CyAA-Pyr dyad. In this case, the HFI constants of the N-methyl protons predominate in BZ, and therefore, one nuclear approximation can be applied for the calculations.
\nIf we set the lifetime of τc = 10 ns, which is close to the experimental values of 7 and 9 ns accordingly, τr = 1 ns, and experimental value Δg = 10−3, then it allows one to describe the difference observed in the values of CIDNP enhancement coefficients of NPX-Trp diastereomers. In this case, the values of hyperfine interaction constants for the (S,S)- and (R,S)-diastereomers differ by a factor of 2 (Figure 15). The same result will be achieved by proportional varying of Δg values. The changes required in the values of Δg factors are unlikely to be accepted as plausible. In addition to the variation of HFI constants and Δg factors, it is worthwhile to consider the influence of changes in characteristic lifetimes of the studied system. However, the experimental data points at much less than the required difference in BZ lifetimes.
\nStarting points and full curves (in insert) of the CIDNP intensity dependences on HFI constant values under B = 4.7 T, τr = 1 ns, Δg = 0.001 and various τc. Reproduced with permission from Ref. [15] © John Wiley & Sons, Ltd., 2018.
Therefore, it seems reasonable to associate changes in the CIDNP enhancement coefficients of NPX-CyAA-Pyr diastereomers with the variation in HFI constants. Moreover, this conclusion is in accordance with the reference data [21, 22, 23]. EPR investigations of chiral stable radicals and biradicals revealed changes in the HFI constants of 1.5 and 2.
\nAnother peculiarity of CIDNP in chiral systems is the dependence of enhancement coefficient ratios for (R,S)- and (S,S)-configurations on the ratios of diastereomer concentrations [15]. It was detected upon UV irradiation of diastereomers mixture of NPX-CyAA-Pyr dyad (Figure 16).
\nNMR and time-resolved (TR) CIDNP spectra of N-methyl protons area (full are presented in insert) detected in the mixture of NPX-CyAA-Pyr dyad diastereomers in acetonitrile (the ratio of (R,S)-/(S,S)- = 0.8). Reproduced with permission from Ref. [15] © John Wiley & Sons, Ltd., 2018.
The K values obtained for different ratios of diastereomers concentrations are presented in Table 1. The noticeable changes in the coefficient ratios, K, are visible.
\nRatio of (R,S)/(S,S) concentrations | \nK | \n
---|---|
0.4 | \n1.70 ± 0.09 | \n
0.7 | \n1.80 ± 0.09 | \n
0.8 | \n1.80 ± 0.09 | \n
1.0 | \n1.9 ± 0.1 | \n
1.3 | \n2.0 ± 0.1 | \n
1.8 | \n2.3 ± 0.1 | \n
2.1 | \n2.3 ± 0.1 | \n
2.3 | \n2.3 ± 0.1 | \n
K values of NPX-CyAA-Pyr dyad diastereomers (column 2) determined for the different ratios of diastereomer concentrations (column 1).
Reproduced with permission from Ref. [15] © John Wiley & Sons, Ltd., 2018.
These changes in the K values obtained for different ratios of diastereomers concentrations point to the contribution of some intermolecular processes to CIDNP formation. The reference data on the tendency of substituted naphthalenes, including naproxen, to association [24, 25, 26] forces authors to suggest that the abovementioned intermolecular process might be the association of dyad diastereomers in solution—most likely, dimer formation. Since the studied dyads contain “naproxen” part and “tails”—the donor groups and bridges, including amide fragment -C(O)NH-, they can form associates due to the hydrogen bonds between amide groups of two molecules, specifically H-bond between amide group of one molecule and carbonyl or carboxyl oxygen of another one (Figure 17). XRD analysis in the solid state of NPX-Trp dyad diastereomers confirmed such H-bond formation [15].
\nSelf-associated (I) and collision complexes (II and III) of dyads with aromatic solvents. Reproduced with permission from Ref. [15] © John Wiley & Sons, Ltd., 2018.
As for the manifestation of the dyads association in solutions, dependences of both chemical shifts and the lines widths of diastereomers on the solvent composition, demonstrated in work [15], also indicate at the possibility of association. According to the reference data, it might be not only dimers (I), but the collision complexes of amide group and indole ring with aromatic solvents (II,III) (Figure 17).
\nBesides, the line widths of amide groups NH protons for both dyads showed the selective broadening (Figure 18).
\nNMR spectra (the region of aromatic protons) of (R,S)-/(S,S)-NPX-Trp and (R,S)-/(S,S)-NPX-CyAA-Pyr dyads in the solvent mixture: CD3CN + 0.17% H2O. Reproduced with permission from Ref. [15] © John Wiley & Sons, Ltd., 2018.
These broadenings, along with chemical shifts solvent dependence, were referred to the slow exchange between monomers and dimers of dyads (Scheme 3, the first equilibrium) and to the solvation of dyads via H-bond formation (Scheme 3, the second equilibrium) [15].
\nPossible ways of the participation of amide fragment of studied dyads in H-bond formation.
On the other side, the fraction of the dyad associates and weak collisional complexes in acetonitrile and benzene mixture may vary depending on the ratios of components in the mixture. Then, we can assume that processes in solution would be different for (R,S)- and (S,S)-optical configurations of the dyads and might result in the K value variation in different solvents, accordingly.
\nThe impact of solvent on CIDNP coefficients of diastereomers was studied in work [15] on the example of NPX-Trp dyad, known not to form exciplex upon the UV irradiation. For this dyad, the minimal ratio of CIDNP enhancement coefficients of diastereomers is observed in acetonitrile 1.4 ± 0.1. The presence of benzene in the mixture provides a large difference between the CIDNP of diastereomers, and this difference grows with the increasing of water concentration in solvent mixture (volume fraction % CD3CN/% C6D6/% H2O): 1.5 ± 0.1 (19.9/80/0.1) < 1.7 ± 0.1 (59.8/40/0.2) < 2.0 ± 0.1(79.7/20/0.3). To explain the distinctions in K values in the different solvents mixtures, in work [11], several circumstances were taken into account: first, the capacity of the amide fragments for self-associating in polar solvents (Figure 17, I), such as acetonitrile, and second, the formation of collision complexes with aromatic compounds (Figure 17, II and III), including benzene [27].
\nSo, the difference in the solvent effects, that is, the association of optical isomers or formation of collisional complexes, can lead to distinctions in the BZ conformations. The latter also assumes the difference in the efficiency of singlet-triplet conversion that is responsible for CIDNP formation and magnitudes of diastereomers enhancement coefficients.
\nFurther study of the effect of association on charge transfer processes in chiral dyads was undertaken in work [15]. Authors, in order to confirm the relation of K values dependence on (R,S)- and (S,S)-concentration ratios with dimers formation, calculated the dependence of K values on diastereomer concentration ratios in the frame of Frank theory of chiral catalysis [28].
\nFrank theory is one of the most accepted theories among chemists, in which it was mathematically shown that a small amount of one chiral compound could increase its own reproduction and suppress the formation of another optical isomer [28]. The main requirement of Frank conception is the following: to obtain one isomer’s prevalence, the presence of a chiral associate, for example, (S,S), (R,R), and (R,S) dimers, in the reaction mixture, is needed. In this case, the driving force of chiral enrichment is the competition between the catalyzing action of one enantiomer on the formation of the same isomer, and the inhibition of the formation of another optical isomer. So, the formation of homodimers (S,S), (R,R) will lead to chiral enrichment with one isomer and heterodimer (R,S) does not.
\nSo, in work [15], to prove that polarized dyad diastereomers are products of back ET in homo- and heterodimers (R,S)-(R,S), (S,S)-(S,S), and (R,S)-(S,S), it was shown how a different ratio of diastereomers changes the value of K. Below is the scheme, where CIDNP values, alpha and beta, are proportional to back ET efficiency (Scheme 4).
\nPhotoinduced processes, occurring in homo-(DRS,RS, DSS,SS) and hetero-DRS,SS chiral dyad dimers. Reproduced with permission from Ref. [15] © John Wiley & Sons, Ltd., 2018.
By Frank concept, αRS and αSS are appreciably higher than β, and, according to the experimental data, αRS is twice as high as αSS. Concentration of dimers was believed to be considerably higher than concentration of monomers. The calculation of K values was carried out by using a quasi-steady-state approximation. On account of the above assumption, the dimer’s stability constants have to be about 105 M−1 at the initial monomers concentration 10−3 M. For detailed description of calculations, readers are referred to work [15].
\nFigure 19 displays the calculated (curve) and experimental (red circles) dependences of CIDNP coefficient ratios on diastereomer concentration ratios.
\nExperimental dependence of CIDNP enhancement coefficients K (data from Table 1, red balls) and calculated Kc (black line) dependence on (R,S)-diastereomer concentration. [SS]0 = 10−3 M, αSS = 10, αRS = 20, βRS = 1, βSS = 1; KRS = 2 × 105 M−1, KSS = 2 × 105 M−1, KRS,SS = 1 × 105 M−1. Reproduced with permission from Ref. [15] © John Wiley & Sons, Ltd., 2018.
The agreement of experimental and calculated K values confirms the impact of association on ET process in dyad’s diastereomers. So, back ET occurs in BZs, being part of dimers. It might be the additional reason of HFI constants variation in BZ of (R,S)- and (S,S)-configurations, resulting from different conformations of BZs. Since the dimerization also changes ET efficiency, the impact of dyad’s dimerization on back ET efficiency might be considered as an example of ET chiral catalysis.
\nSo, this chapter shows that the set of dyads, including (R)/(S)-NPX and chiral partner, have demonstrated stereoselectivity—difference in ET rates 0f diastereomers. The joint studies by spin chemistry and photochemistry techniques, including the time-resolved measurements of dyads with donor N-methylpyrrolidine, have shown the stereoselectivity of separate stages of NPX excited state quenching, LES, and exciplex quantum yields.
\nThe scale of these differences was found to be influenced by the donor-acceptor properties of the partners and the length and structure of the bridge. Another peculiarity of ET—spin selectivity was observed on the example of two NPX-based dyads. It is the different CIDNP effects formed via back ET in BZ of diastereomers. Spin selectivity is explained by the difference between HFI constants, which, in turn, determine back ET in BZs of (R,S)- and (S,S)-configuration. The impact of dyad’s dimerization on ET efficiency might be considered as the first example of chiral catalysis in elementary process—ET.
\nThis work was supported by the Russian Science Foundation (grant no. 18-13-00047). The authors are truly grateful to Prof. Plyusnin who contributed a lot to the work of Russian scientists.
\nColorectal cancer is the third most common cancer worldwide and remains an important cause of death. CRC diagnosis and treatment require a multidisciplinary approach, and in stage IV disease combination chemotherapy (CT) and regional multimodality treatments − like metastasectomy and other local treatments − are increasingly used. Systemic therapy has evolved over the past few decades, with the emergence of combination CT and targeted agents (Figure 1).
Targeted therapies that have been approved or are currently under investigation for advanced colorectal cancer.
In the present review, genomic and tumor microenvironment alterations driving treatment selection are discussed.
Metastatic CRC (mCRC) presents with synchronous metastatic disease at initial diagnosis in 20% of cases, with 50–60% of patients developing metachronous metastases. Approximately 56% of patients with CRC will ultimately die from their cancer [1]. The cornerstone of CRC treatment for 20 years has been fluoropyrimidine-based CT doublets, with either irinotecan (FOLFIRI or CAPIRI) or oxaliplatin (FOLFOX or CAPOX) in the first- and second-line settings [2].
In the past two decades, remarkable progress has been achieved in mCRC treatment with the introduction of molecular targeted agents (Figure 2). Today, the median overall survival (OS) for these patients in phase III trials is approximately 30 months, more than doubling that of 20 years ago [3]. Simultaneously, mortality has declined, what is attributed to earlier diagnosis (due to screening tests) and improved treatment options, including new systemic CT agents and biologic agents targeting specific pathways [1].
Timeline of development of targeted therapies in colon cancer.
More recently, consensus molecular subtypes (CMS) defined by gene expression profiling have identified biologically different CRC subtypes, which seem to have a prognostic and predictive value. However, CMS subtyping is not a standard test with therapeutic application at present, being more relevant in the research field [2].
New targeted therapies against the epidermal growth factor receptor (EGFR) had an impressive impact on mCRC prognosis, with an actual median OS over 30 months (varying according to therapeutics options) [4, 5, 6].
As part of the ErbB tyrosine kinase family, EGFR is a transmembrane receptor and its activation by extracellular ligands stimulates downstream pathways, such as RAS–RAF–MEK-MAPK, PIK3CA-AKT, the SRC family kinases, PLCγ-PKC, and JAK/STATs, inducing proliferation, migration, invasion, survival, and angiogenesis [6, 7]. Thus, EGFR is an important factor in tumor development and progression, being expressed in various cancers and in 60–80% of CRCs [8].
Target therapy against EGFR is now a standard of care in RAS wild-type mCRC. Two monoclonal antibodies (mAbs) are approved: cetuximab (human-mouse chimeric mAb) and panitumumab (fully human mAb). By recognizing and binding to the extracellular domain of the EGFR receptor, these mAbs prevent binding of other extracellular ligands and subsequent receptor internalization and degradation, thus inhibiting and blocking downstream pathways and signaling [9]. Tumor RAS mutational status predicts efficacy of anti-EGFR agents in mCRC patients, with RAS mutations being a well-established negative predictive biomarker for patient selection [10].
Several phase II and III clinical trials have established the efficacy of cetuximab and panitumumab, either in monotherapy or in association with CT, in terms of progression-free survival (PFS), OS, and overall response rate (RR), while maintaining quality of life (Table 1) [6, 11, 12, 13].
Setting | Study | Treatment | RR□, % | PFS□, months | OS□, months |
---|---|---|---|---|---|
1st line | PRIME | PAN+FOLFOX4 FOLFOX4 | 59* 46* | 10.1* 7.9* | 26.0* 20.2* |
1st line | PEAK | PAN-mFOLFOX6 mFOLFOX6 | 64 61 | 13.0* 9.5* | 41.3 28.9 |
1st line | PLANET-TTD | PAN-FOLFOX4 PAN-FOLFIRI | 74 67 | 12.8 14.8 | 39.0 45.8 |
1st line | 314 | PAN-FOLFIRI | RASwt: 56* RASmt: 38* | RASwt: 8.9* RASmt: 7.2* | NR |
1st line | COIN | CET-OXAL OXAL | 64* 57* | 8.6 8.6 | 17.9 17.0 |
1st line | OPUS | CET-FOLFOX4 FOLFOX4 | 61* 37* | 8.3* 7.2* | 22.8 18.5 |
1st line | CRYSTAL | CET-FOLFIRI FOLFIRI | 46.9* 38.7* | 9.9* 8.7* | 24.9 21.0 |
1st line | FIRE-3 | CET-FOLFIRI BEVA-FOLFIRI | 62.0 58.0 | 10.0 10.3 | 28.7* 25.0* |
1st line | CALGB 80405 | CET-FOLFOX/FOLFIRI BEVA-FOLFOX/FOLFIRI | 59.6 55.2 | 10.5 10.6 | 30.0 29.0 |
2nd or greater | 181 | PAN-FOLFIRI FOLFIRI | 36* 10* | 5.9* 3.9* | 14.5 12.5 |
2nd or greater | PICOLLO | PAN- CPT-11 CPT-11 | 34* 12* | HR 0.78* | 10.4 10.9 |
2nd or greater | Saltz, 2004 | CET | 8.8 | 1.4 | 6.4 |
2nd or greater | Cunningham, 2014 | CET + CPT-11 CET | 22.9* 10.8* | 4.1* 1.5* | 8.6 6.9 |
2nd or greater | ASPECCT | PAN CET | 22.5 20 | 4.1 4.4 | 10.4 10.0 |
Targeted therapies against EGFR in colorectal cancer.
Results for the KRAS wild-type subgroup, except if clearly stated.
Difference between groups is statistically significant (p < 0.05).
BEVA, bevacizumab; CET, cetuximab; CPT-11, irinotecan; mt, mutated; NR, not reported; ORR, overall response rate; OS, overall survival; OXAL, oxaliplatin-containing chemotherapy regimen; PAN, panitumumab; PFS, progression-free survival; wt, wild-type.
The PRIME trial, a randomized phase III trial investigating the addition of panitumumab to FOLFOX4 as first-line therapy in RAS wild-type mCRC, showed a 2- and 6-month PFS and OS benefit, respectively, with the combination. Regarding safety, known EGFR inhibition adverse events (AE) were more frequently observed with panitumumab, including skin toxicity and diarrhea (36% vs. 2% and 18% vs. 9% in panitumumab and placebo arms, respectively) [11].
The randomized phase II PEAK trial compared the efficacy and safety of mFOLFOX6 plus panitumumab with mFOLFOX plus bevacizumab (an anti- vascular endothelial growth factor [VEGF] mAb) as first-line therapy in RAS wild-type mCRC. The study primary endpoint was met, with panitumumab showing a 3.5-month PFS increase compared with bevacizumab. An OS improvement was also observed, although not statistically significant [14]. Rivera et al. and Stintzing S et al. also demonstrated that early tumor shrinkage is an important and early predictor of treatment sensitivity and deep tumor response correlates with OS [15, 16].
The open-label phase II PLANET-TTD trial compared panitumumab with two different CT regimens (FOLFOX 4 and FOLFIRI) as first-line treatment of RAS wild-type mCRC, but no significant efficacy differences were observed between the two regimens [17].
The 314 trial, a single-arm phase II study evaluating first-line panitumumab plus FOLFIRI in mCRC patients, confirmed the impact of KRAS exon 2 status in being a negative predictor of efficacy in mutant patients. In a total of 154 patients, 59% had KRAS wild-type tumors. RR and median duration of response (DoR) were higher in the KRAS wild-type group. Additionally, more patients in the wild-type group underwent R0 resection (8% vs. 5%), and a PFS benefit was also observed in this group (8.9 vs. 7.2 months) [18].
In the COIN trial, cetuximab was added to oxaliplatin-containing CT (FOLFOX or CAPOX) in first-line setting of mCRC. In patients with KRAS wild-type tumors, no OS or PFS difference was reported between the two groups, while overall response rate (ORR) was higher with the addition of cetuximab to CT compared to CT alone [19].
Similar ORR results were seen in the OPUS trial. In KRAS wild-type tumors, the addition of cetuximab to FOLFOX-4 was associated with a clinically significant increased chance of response and a lower risk of disease progression. The same results were not seen in the overall population, confirming the relevance of KRAS mutational status [12].
Although the addition of cetuximab to oxaliplatin-containing CT had little survival impact, the CRYSTAL trial showed different results when combining cetuximab to FOLFIRI. A borderline significant PFS increase was seen in the combination arm, although with no OS differences. However, when KRAS mutational status was considered, a significant PFS increase was observed favoring cetuximab [20].
Additionally, in the phase III open-label FIRE-3 trial, cetuximab was compared with bevacizumab, both in combination with FOLFIRI. No differences were observed in the primary endpoint of ORR or in PFS, but the median OS was improved in cetuximab arm [21].
Cetuximab was further compared with bevacizumab, both combined with CT (FOLFOX or FOLFIRI), in the CALGB 80405, with no significant differences in ORR, PFS, or OS [22].
In the 181 trial, the efficacy and safety of adding panitumumab to FOLFIRI was compared with FOLFIRI alone in RAS wild-type mCRC patients who had failed the initial treatment. Addition of panitumumab to the regimen resulted in a significant PFS improvement, of approximately 2 months. Although not significant, a trend towards an OS benefit was seen with the addition of panitumumab [23].
Conversely, the randomized open-label PICOLLO trial reported no benefit with the addition of panitumumab to irinotecan after progression on fluoropyrimidine, with or without oxaliplatin. However, better PFS and more responses were reported in the panitumumab group [24].
In 2004, Saltz et al. and Cunningham et al. evidenced the role of cetuximab in heavily pretreated patients. Saltz et al. reported a median OS of 6.4 months and a median PFS of 1.4 months in 57 patients receiving cetuximab monotherapy after progression on irinotecan, and a tumor RR of 8.8% [25]. Cunningham et al. included over 300 patients and investigated the role of cetuximab (with or without irinotecan) after progression on irinotecan. A PFS and ORR benefit was observed, with a numeric but not statistically significant difference also observed in OS (8.6 vs. 6.9 months) [26].
Later, the randomized phase II ASPECCT trial compared panitumumab alone with cetuximab alone as third-line treatment for mCRC patients with RAS wild-type (exon 2) tumors. With OS as primary endpoint, panitumumab was given at a dose of 6 mg/Kg every two weeks and cetuximab at a loading dose of 400 mg/m2, followed by a weekly dose of 250 mg/m2. No efficacy differences were observed, with a median OS of 10.4 months for panitumumab and 10.0 months for cetuximab [27].
Regarding maintenance and treatment intensification, three clinical trials are worth mentioning: VOLFI, VALENTINO, and SAPPHIRE.
VOLFI was a randomized open-label phase II trial comparing the addition of panitumumab to FOLFOXIRI CT regimen. An ORR of 87,3% was seen in the FOLFOXIRI plus panitumumab arm, which was higher compared with FOLFOXIRI alone. PFS was similar in both arms, whereas OS showed a trend in favor of panitumumab [28]. This was the highest ORR reported in mCRC, suggesting that these protocols can be considered to obtain maximum cytoreduction in selected patients.
The VALENTINO trial, an open-label phase II trial, investigated maintenance therapy with panitumumab (induction therapy with FOLFOX-4 + panitumumab followed by maintenance with panitumumab ±5FU/LV). The study hypothesis that panitumumab alone was not inferior to the combination as maintenance therapy could not be proven. ORR and OS results did not differ between the two arms [29].
In the SAPPHIRE trial, patients received six cycles of mFOLFOX6 plus panitumumab as induction therapy. Patients who completed induction therapy without progression were then randomized to mFOLFOX6 plus panitumumab (group A) or 5-FU/LV plus panitumumab (group B). PFS, RR, OS, and time to treatment failure were similar between groups, adding to the concept that planned discontinuation of oxaliplatin after six cycles of mFOLFOX6 is a potential treatment option for mCRC patients, achieving similar efficacy while reducing oxaliplatin-associated peripheral neuropathy compared with mFOLFOX6 plus panitumumab [30].
Although anti-EGFR therapy has shown benefit in a particular subgroup of CRC patients, primary or innate resistance is high among unselected patients. Furthermore, even patients that initially respond to cetuximab and panitumumab, eventually develop resistance and relapse under these therapies (secondary resistance). Knowledge of the resistance mechanisms associated with the EGFR pathway is crucial to improve therapy efficacy.
RAS–RAF-MAPK is an EGFR direct downstream signaling pathway, highly deregulated in CRC. Mutations frequently found in these family members generally lead to protein constitutive activation independently of the upstream signaling cascade. Over the last decade, analysis of retrospective clinical trial data (in particular of the OPUS, CRISTAL, and PRIME trials) led to the discovery that patients harboring RAS (KRAS and NRAS) and BRAF (specially V600E) activating mutations do not benefit from cetuximab and panitumumab treatment, and that it could even be detrimental for them [31]. These results have led the European Medicines Agency (EMA) and Food and Drug Administration (FDA) to recommend against the use of EGFR-targeted therapies in patients harboring RAS and BRAF mutations. These mutations are currently the only clinically validated predictive marker of resistance to anti-EGFR therapies in CRC.
Although RAS and RAF mutations are effective in predicting resistance, not all wild-type patients respond to cetuximab and panitumumab. The EGFR receptor also signals through the PI3K-AKT pathway, resulting in tumor cell proliferation and survival [32]. Retrospective studies of cetuximab treatment in chemorefractory metastatic CRC patients revealed that KRAS wild-type patients with PIK3CA mutations in exon 20 (but not in exon 9) have lower response rates compared to unmutated patients (0.0% vs. 36.8%; 95% confidence interval [CI] 0.00–0.89; p = 0.029) [33]. PTEN is another potential marker of response to anti-EGFR therapy, given its inhibitory role on PI3K-AKT signaling pathway. Although PTEN studies are scarce and inconclusive, some works suggest that loss of PTEN expression (measured by immunohistochemistry [IHC]) is associated with decreased RR, PFS, and OS in metastatic CRC patients treated with anti-EGFR therapy [34, 35].
Evidence from cellular studies has suggested that constitutive activation of other EGFR downstream pathways, such as those including the JAK–STAT family, are implicated in resistance to the anti-EGFR gefitinib [36, 37].
Additionally, amplification of other receptor tyrosine kinases (RTKs) has been proposed as a resistance mechanism to anti-EGFR therapies. Expression of VEGF-1 or its receptor (VEGFR) has been associated with cetuximab resistance in both preclinical models and metastatic CRC patients [38]. Bertotti et al. reported that human epidermal growth factor receptor 2 (HER2) gene amplification correlated with cetuximab resistance in a patient-derived xenograft mouse model [39]. Besides HER2, also HER3 has been described to have a role in resistance mechanism to EGFR-targeted therapies. In a cohort of metastatic CRC patients treated with irinotecan and cetuximab, HER3 overexpression was associated with lower PFS and OS [40].
Finally, growing evidence implicates the MET pathway in both primary and secondary resistance mechanisms to mAbs in KRAS wild-type patients, through MET amplification or hepatocyte growth factor (HGF) increased expression [41]. In a randomized phase II clinical trial of chemorefractory KRAS wild-type anti-EGFR-naïve patients, the combination of anti-HGF mAbs and panitumumab led to higher RR and a trend towards better outcomes in the population with MET overexpression [42].
Although RAS mutations are negative predictors of efficacy in cetuximab and panitumumab treatment, it is acknowledged that not all RAS wild-type patients respond to these agents. To investigate this, research efforts were driven downwards in the MAPK pathway, putting the spotlight on BRAF. This is the main effector in EGFR pathway and is usually mutated in 5–10% of mCRC patients. BRAF and KRAS are usually mutually exclusive, with BRAF V600E mutation (class I) accounting for most alterations found and conferring worse prognosis to these patients.
Regardless of EGFR blockade, BRAF mutations can keep the downstream signaling persistently activated, suggesting that they can confer EGFR blockade resistance. In fact, in a retrospective trial, De Roock et al. showed that chemorefractory mCRC patients with BRAF V600E mutations have significantly lower RR to cetuximab than patients with wild-type tumors (8.3% vs. 38.0%; odds ratio 0.15; p = 0.0012) [43]. Several multicentre trials and meta-analyses have subsequently confirmed that BRAF V600E mutation results in shorter PFS and OS compared to the wild-type phenotype, emphasizing its role in resistance to anti-EGFRs in patients with chemorefractory mCRC.
Multiple combinations with drugs targeting the MAPK pathway have been tested in BRAF-mutant CRC. Monotherapy results were disappointing when compared to the clinical activity seen in melanoma. In contrast to melanoma, CRC expresses high levels of activated EGFR, which reactivate the MAPK pathway after single BRAF inhibition [44, 45]. In view of the possibility of therapy resistance via EGFR signaling feedback activation, the trial was amended to include safety and efficacy assessment of vemurafenib combined with cetuximab in a heavily pretreated population, with positive results (median PFS of 3.7 months and median OS of 7.1 months). Similar results were observed when combining dabrafenib with panitumumab (median PFS of 3.5 months) and encorafenib with cetuximab (RR of 23.1%, median PFS of 3.7 months), with phase II results of the latter showing a median PFS of 4.2 months and an ORR of 22% [46].
CT was also combined with BRAF and EGFR inhibition in a phase II trial of irinotecan, cetuximab, and vemurafenib. A total of 106 patients were enrolled, with the study reporting a PFS benefit of 4.3 months with the addition of vemurafenib compared to 2.0 months in the control arm [47].
BRAF inhibition can also induce EGFR overactivation or PI3K modulation, and triplet combos targeting EGFR, MAPK, and PI3K have shown positive results. The MEK116833 trial included 24 patients receiving full-dose combination of panitumumab, trametinib, and dabrafenib and reported an ORR of 21%, a median PFS of 4.1 months, and an OS of 9.1 months. Additionally, a randomized phase II trial combining encorafenib, cetuximab, and the PI3K inhibitor alpelisib reported a median PFS of 5.4 months and an ORR of 27% in interim analysis [48, 49, 50, 51].
More recently, the phase 3 BEACON trial investigated the doublet of encorafenib plus cetuximab and the triplet of encorafenib plus cetuximab plus binimetinib in patients with BRAF-mutant CRC after one or two prior regimens. The updated analysis confirmed an ORR of 27% with the triplet versus 20% with the doublet versus 2% in the control arm. Median OS was 9.3 months with the duplet and 5.9 months in the control group (hazard ratio [HR] 0.61). The benefit was seen across all subgroups. Numerically identical median OS was observed when comparing the triplet and doublet, with higher toxicity for the triplet (mainly gastrointestinal toxicity and anemia). Subgroup analysis suggested survival benefits in some subgroups, such as those with ECOG 1, three or more organs affected, and higher levels of C-reactive protein and with unresected primary tumors, suggesting that patients with higher disease burden and inflammatory drive could benefit from triple therapy. PFS was also comparable between doublet and triplet and clearly superior to the control arm [52, 53].
HER2 is a growth factor receptor involved in CRC development and progression. HER2 amplification is relatively uncommon, reported in only 3–5% of metastatic CRC patients with wild-type KRAS and wild-type BRAF [54].
Trastuzumab is a monoclonal antibody targeting HER2. The phase II HERACLES trial included mCRC patients with KRAS wild-type, HER2-positive (defined as 2+/ 3+ HER2 score in >50% of cells by IHC or HER2:CEP17 ratio > 2 in >50% of cells by fluorescent in situ hybridization [FISH]) tumors who were refractory to standard therapy with EGFR inhibitors and were treated with trastuzumab and lapatinib. ORR was 30%, with one complete response, and median OS was 46 weeks [55]. The most common AEs were diarrhea, rash, and fatigue (78%, 48%, and 48%, respectively). These findings suggested that HER2 positivity was an important driver in CRC. In the phase IIa multi-basket MYPATHWAY trial, patients with HER2-amplified tumors (including CRC) received dual blockade therapy with pertuzumab and trastuzumab. Preliminary results showed promising response, with an ORR of 37.5%, and suggested durable responses with HER2-targeting agents, with a median DoR of 11 months [56].
Both the TRIUMPH (trastuzumab and pertuzumab) and MOUNTAINEER (trastuzumab and tucatinib) trials reported high response rates (35% and 52%, respectively) and encouraging median PFS (4.0 and 8.1 months, respectively), supporting dual HER2 blockade in patients with HER2-amplified metastatic CRC [57, 58]. Conversely, the combination of pertuzumab and TDM-1 did not show an enhanced objective response in the HERACLES-B trial, although achieving a similar disease control to the HERACLES-A trial (ORR of 10% and median PFS of 4.8 months at cut-off) [59].
Regarding new antibody-drug conjugates, the phase 2 DESTINY-CRC01 trial, of trastuzumab deruxtecan (T-DXd; DS-8201) and also in patients with metastatic HER2-amplified CRC, reported significant responses (ORR of 45.3%, disease control rate [DCR] of 83%), including in patients previously submitted to HER2 blockade [60].
Tumor angiogenesis is one of the hallmarks of cancer and a key process in tumor development [61, 62]. One of the most relevant pathways involved in angiogenesis is the vascular endothelial growth factor/vascular endothelial growth factor receptor (VEGF/VEGFR) signaling pathway. VEGF-A is a heparin-binding glycoprotein with potent angiogenic activity. VEGF is produced by different cell types, such as immune cells, fibroblasts, and cancer cells, in response to tumor hypoxia via hypoxia-inducible factor (HIF)-1a pathway, inducing an angiogenic switch [63]. Overproduction of pro-angiogenic growth factors leads to formation of chaotic blood vessels in the tumor, with a leaky endothelial wall [64].
In CRC, primary tumor growth and distant metastases development are highly dependent on new vessel formation, making VEGF signaling pathway an attractive therapeutic target. Inhibition of VEGF signaling pathway can be achieved through neutralizing antibodies binding VEGF ligands or blocking VEGFR, or tyrosine kinase inhibitors (TKIs) blocking intracellular VEGFR-dependent signaling [65].
Bevacizumab. The first angiogenesis inhibitor approved for mCRC was bevacizumab, an immunoglobulin G (IgG)1 monoclonal antibody with affinity to VEGF-A. Several trials have evaluated the benefit of adding bevacizumab to cytotoxic regimens as first-line treatment of patients with mCRC, with inconsistent PFS and OS results (Table 2).
Study | Treatment | PFS, months | OS, months | HR (p-value) |
---|---|---|---|---|
Hurwitz et al. (III) | BEVA-ILF PLACEBO-IFL | 10.6* 6.2* | 20.3 15.6 | PFS - 0.54 (<0.001) OS - 0.66 (>0.001) |
Saltz et al. (III) | XELOX BEVA-FOLFOX PLACEBO | 9.4* 8.0* - | 23.3 19.9 - | PFS - 0.83 (0.002) OS - 0.89 (0.077) |
AVEX (III) | BEVA-CAP CAP | 9.1* 5.1* | — | PFS - 0.53 (<0.001) |
ITACa (III) | BEVA-FOLFIRI/FOLFOX PLACEBO-FOLFIRI/FOLFOX | 9.6 8.4 | — | PFS - 0.86 (0.182) |
SOLSTICE (III) | BEVA-Trifluridine/tipiracil BEVA-CAP | — | — | Ongoing |
VELOUR (III) | Aflibercept-FOLFIRI PLACEBO-FOLFIRI | 6.90* 4.67* | 13.50* 12.06* | PFS - 0.758 \t(<0.001) OS - 0.817 (0.003) |
AFFIRM (II) | Aflibercept-FOLFOX PLACEBO-FOLFOX | 8.48 8.77 | — | PFS - 1.00 |
RAISE (III) | Ramucirumab-FOLFIRI PLACEBO-FOLFIRI | — | 13.3* 11.7* | OS - 0.844 (0.022) |
CORRECT (III) | Regorafenib PLACEBO | — | 6.4* 5.0* | OS - 0.77 (0.005) |
CONCOUR (III) | Regorafenib PLACEBO | — | 8.8* 6.3* | OS - 0.55 (<0.001) |
CONSIGN (III) | Regorafenib | AEs: hypertension (15%), hand-foot skin reaction (14%), fatigue (13%), diarrhea (5%), and elevated aminotransferase (6%), aspartate aminotransferase (7%), and bilirubin (13%). |
Targeted therapies against VEGF in colorectal cancer.
Difference between groups is statistically significant (p < 0.05).
AEs, adverse events; BEVA, bevacizumab; CAP, capecitabine; HR, hazard ratio; OS, overall survival; PFS, progression-free survival.
A phase III trial conducted by Hurwitz et al. compared the efficacy of irinotecan, bolus fluorouracil, and leucovorin (IFL) plus bevacizumab versus IFL plus placebo in untreated mCRC patients. Bevacizumab was intravenously administered at a dose of 5 mg/kg every two weeks along with CT. Bevacizumab arm showed a meaningful improvement in OS (20.3 versus 15.6 months in placebo arm) and PFS (10.6 versus 6.2 months in placebo arm) [66]. Saltz et al. assigned mCRC patients in a 2x2 factorial design to receive CAPOX or FOLFOX4 followed by bevacizumab or placebo as first-line treatment. Median PFS was higher in the bevacizumab group compared with placebo (9.4 versus 8.0 months). OS differences did not reach statistical significance, but only 29% of bevacizumab recipients were treated until disease progression or toxicity [67]. For elderly patients with untreated and unresectable mCRC not candidates for oxaliplatin- or irinotecan-based therapies, the phase III AVEX trial compared the efficacy and safety of capecitabine combined with bevacizumab versus capecitabine alone. Capecitabine was given at a dose of 1000 mg/m2 orally twice a day on days 1–14 and bevacizumab was administered intravenously at a dose of 7.5 mg/kg on day 1, every 21 days. Longer PFS was documented in the bevacizumab arm (9.1 versus 5.1 months for capecitabine alone), with acceptable tolerance. Grade ≥ 3 adverse events reported in the combination arm included hand-foot syndrome (16%), diarrhea (7%), and venous thromboembolic events (8%) [68].
Despite these results, the 2015 phase III ITACa trial reported no statistically significant PFS and OS differences when bevacizumab was added to standard first-line CT (FOLFIRI or FOLFOX4) [69]. Other previous trials reported the same negative results. Considering these discrepancies, a 2017 meta-analysis based on 9 studies examined the survival impact of bevacizumab plus CT in first-line treatment of mCRC patients, showing that the combination significantly prolonged PFS (HR 0.66; p < 0.0001) and OS (HR 0.84; p = 0.0001) compared with CT alone. Subgroup analyses suggested that irinotecan-based regimens might be a better partner for bevacizumab than oxaliplatin-based regimens, with superior PFS and OS benefit [70].
Sidedness of the primary tumor is known to be an important prognostic factor in metastatic setting of CRC, with worst survival outcomes for right-sided tumors. Several clinical trials investigated the prognostic role of bevacizumab in the treatment of patients with right-sided and left-sided CRC. A post-hoc analysis of 16 randomized trials including PEAK, FIRE-3, and CALGB/SWOG trials showed that right-sided tumors have impaired CT sensitivity, while addition of bevacizumab to cytotoxic regimens can be an optimal first-line treatment for RAS-wild-type right-sided mCRC [71].
Although continuing bevacizumab with second-line chemotherapy showed benefit after disease progression, other anti-VEGF drugs should be considered for fast progressors (PFS <3–4 months) [72].
In patients with unresectable mCRC who are not candidates for intensive therapy, the ongoing phase III SOLSTICE trial is currently comparing trifluridine/tipiracil (TAS-102) plus bevacizumab versus capecitabine plus bevacizumab as first-line treatment [73].
Aflibercept. Aflibercept is a recombinant fusion protein composed by VEGF-binding portions from VEGFR-1 and -2 extracellular domains fused to the Fc portion of human IgG1. It acts by blocking the activity of VEGF-A and -B, preventing their binding to VEGFR on endothelial and tumor cells [74].
The role of aflibercept was evaluated in the phase III VELOUR trial, of mCRC patients previously treated with oxaliplatin-based regimens in first line, including with bevacizumab. Second-line FOLFIRI was intravenously administered with placebo or aflibercept at the dose of 4 mg/kg every two weeks. Aflibercept improved the median OS (13.50 vs. 12.06 months) and median PFS (6.90 versus 4.67 months) compared to placebo [74]. These results lead to approval of the drug in combination with FOLFIRI as second-line treatment for patients pretreated with oxaliplatin-based doublet with bevacizumab. The most common grade ≥ 3 AEs reported in the VELOUR trial included neutropenia, diarrhea, stomatitis, hypertension, and fatigue. Additionally, there was no evidence of greater toxicity in patients previously treated with bevacizumab [74].
More recently, the phase II AFFIRM trial investigated the addition of aflibercept to first-line oxaliplatin-based regimens in mCRC patients. Patients received mFOLFOX6 plus aflibercept or mFOLFOX6 alone. Despite VELOUR results, this study did not reach the primary endpoint of PFS. Adding aflibercept to first-line mFOLFOX6 did not increase efficacy and was associated with higher toxicity [75].
Ramucirumab. Ramucirumab is a human IgG1 monoclonal antibody against VEGFR-2. Efficacy and safety of ramucirumab in combination with second-line FOLFIRI was evaluated in the phase III RAISE trial. Patients with progressive mCRC during or after first-line treatment with bevacizumab, oxaliplatin, and fluoropyrimidine were randomized to receive intravenous ramucirumab 8 mg/kg plus FOLFIRI or placebo plus FOLFIRI every 2 weeks. Ramucirumab significantly improved survival in this subpopulation, reaching a median OS of 13.3 months, against 11.7 months in the placebo arm. Grade ≥ 3 AEs included neutropenia (38%), hypertension (11%), diarrhea (11%), and fatigue (12%). Febrile neutropenia was only reported in 3% of patients and most toxicities reported were manageable [76]. This trial lead to the approval of ramucirumab in combination with FOLFIRI in the second-line setting of mCRC previously treated with bevacizumab, oxaliplatin, and fluoropyrimidine in first line.
Regorafenib. The only TKI approved for mCRC treatment is regorafenib, a multi-kinase inhibitor of angiogenic pathway members, including VEGFR-1 and -2, platelet-derived growth factor receptor (PDGFR)-β, and tyrosine kinase with immunoglobulin-like and EGF-like domains 2 (TIE2) [77].
Several phase III trials evaluated the role and efficacy of regorafenib as single-agent in mCRC patients progressing after several standard lines of treatment (Table 2). The CORRECT trial was the first to compare treatment with regorafenib 160 mg daily for 21 days, every 28-day cycle, versus placebo. Final study results reported a quality of life (QoL) and OS (6.4 vs. 5.0 months in placebo arm) improvement in favor of regorafenib [78]. The phase III CONCOUR trial was similar to the CORRECT trial but exclusively recruited Asian patients, holding similar OS results [79]. The CONSIGN trial was designed to specifically evaluate regorafenib safety. In a total of 2864 patients (median age of 62 years), the most common grade ≥ 3 AEs were hypertension (15%), hand-foot syndrome (14%), fatigue (13%) and diarrhea (5%). Grade ≥ 3 laboratory toxicities included elevated alanine aminotransferase (6%), aspartate aminotransferase (7%), and bilirubin (13%) [80].
Despite the outcome benefits seen with anti-VEGF agents in CRC, these are usually transient and followed by relapse and tumor growth [81]. Several resistance mechanisms to anti-VEGF therapies have been described, including VEGF axis-dependent alterations, non-VEGF axis-dependent upregulation, and stromal cell interactions [82].
Upregulation of alternative VEGFR-2 angiogenic ligands, such as VEGF-C, −D, and placental growth factor (PIGF), can bypass VEGF-A inhibition and elicit bevacizumab resistance [82]. In a phase II trial, Kopetz et al. showed that PlGF, VEGF-C, and VEGF-D plasma levels in mCRC patients receiving FOLFIRI plus bevacizumab were elevated prior to and at the time of disease progression [83].
Complementary angiogenic pathways other than VEGF/VEGFR signaling exert control on tumor angiogenesis and may explain acquired resistance to anti-VEGF therapies. These pathways involve members of the platelet-derived growth factor (PDGF) family, HIF, members of the fibroblast growth factor (FGF) family, angiopoietin (Ang), and Notch [84, 85].
The PDGF family consists of five ligands that bind to tyrosine kinases PDGFR-α and -β, activating downstream signal transduction pathways, as PI3K/Akt and PLCγ. PDGF-C was shown to be upregulated in cancer-associated fibroblasts (CAFs) of anti-VEGF-resistant tumors in vivo [86], making it a possible resistance mediator.
HIF-1 is a transcription factor with a key role in cellular response to reduced oxygen levels. Among its multiple downstream effects is induction of VEGF-A, VEGFR, PIGF, and PDGF expression [85].
Growth factors of the FGF family are potent mediators of tumor angiogenesis. Binding of FGF to fibroblast growth factor receptor (FGFR) tyrosine kinase activates downstream pathways such as MAPK/ERK, PI3K/Akt, and STAT [86], acting synergistically with VEGFA to induce angiogenesis via endothelial cell proliferation, survival, and migration [87]. FGF-2 upregulation is observed in anti-VEGF-resistant tumors, especially in tumors exposed to a hypoxic environment, [86] while FGF-2 blockade results in decreased tumor growth in in vivo models [88].
Ang-Tie signaling is a vascular-specific pathway essential for blood vessel development and vascular permeability regulation. Ang-2 acts as an antagonist of the Tie2 receptor, leading to development of vascular sprouts in the context of VEGF exposure [86]. mCRC patients with poor bevacizumab response showed high serum Ang2 levels, suggesting its relevance in resistance to anti-angiogenic therapy [89].
Delta-like ligand 4 (DII4) is a Notch ligand overexpressed in several solid malignancies, including CRC. DII4 upregulation is thought to contribute to bevacizumab resistance, which can be overcome by Notch inhibition with a γ-secretase inhibitor [90].
TGF-β is a ligand for type II TGF-β receptors and endoglin (CD105). It has important regulatory functions in angiogenesis, either directly, or indirectly by activating fibroblasts to produce extracellular matrix and stimulating the tube formation in endothelial cells [91]. Anti-VEGF therapy-resistant tumors can exhibit high levels of TGF-β1 expression. Additionally, in preclinical models VEGF pathway blockade led to increased CD105 levels, suggesting a role for CD105 in anti-VEGF therapy resistance [92].
It has been recently suggested that tumor stromal cells and bone marrow-derived cells (BMDCs) recruited to the tumor microenvironment by secreted cytokines play an important role in acquired resistance to anti-VEGF therapies [81].
CAFs entail a large portion of stromal cells present in the tumor environment. These cells secrete a number of pro-angiogenic mediators, including IGF, FGF, EGF, cytokines, and chemokines, and are capable of recruiting endothelial progenitor cells (EPCs) to the tumor site [93, 94]. Interestingly, Kinugasa et al. showed that CAFs from anti-VEGF-resistant tumors express high levels of CD44, a marker for cancer stem cells and cytotoxic resistance. CAFs can hence be considered a promising target for overcoming resistance to anti-angiogenic agents [95].
BMDCs are comprised of endothelial and pericyte progenitors, macrophages, and myeloid-derived suppressor cells (MDSCs) [96]. Preclinical models suggest that EPCs in the tumor microenvironment are able to secrete different proangiogenic factors and accelerate angiogenesis [97]. More importantly, endothelial precursor cells can differentiate into endothelial cells and participate in new vessel formation [98, 99].
Tumor-associated macrophages (TAMs) are also involved in angiogenesis. VEGF blockade by bevacizumab seems to promote TAM proliferation and reprogramming to pro-angiogenic macrophages [81]. This type of macrophages can secrete VEGF-A, TNFα, and IL-8, all of which affect different stages of angiogenesis by modifying the local extracellular matrix, promoting proliferation and migration of endothelial cells, and inhibiting development of differentiated capillaries [81].
A study by Shojaei et al. demonstrated that MDSCs were present in higher levels in anti-VEGF-resistant tumors and were functionally different from those in anti-VEGF-sensitive tumors. This population was able to sustain tumor growth even in presence of anti-VEGF inhibitors, although the exact mechanism behind this is not been fully established [100].
CD4+ T-helper cells mediate anti-VEGF resistance through IL-17 production in the tumor microenvironment and BMDC recruitment. These cells have been shown to regulate secretion of several proangiogenic factors from CAFs and other stromal cells. Additionally, Numasaki et al. reported that tumor microvessel density correlates with levels of infiltrating IL-17-producing CD4 T-cells [25, 42, 81, 101].
The main side effects of the anti-EGFR therapies cetuximab and panitumumab are dermatological toxicities, reported in 85–96% of patients (Table 3) [102]. The most common AE is papulopustular skin rash, generally developing over a period of 6 weeks after starting treatment and potentially impacting quality of life and therapy adherence. General prevention and management principles include the use of skin moisturizer, sunscreen, hydrocortisone cream, and oral tetracycline. The STEPP trial compared pre-emptive with reactive skin treatment and showed an over 50% reduction in grade ≥ 2 skin toxicities and less QoL impairment with the pre-emptive compared with reactive treatment [103]. In cases of grade 3 rash, treatment should be delayed until toxicity has resolved to grade 2 or less and dose should be reduced in a second occurrence. In grade 1 or 2 rash, dose reduction is not indicated. Other dermatological symptoms, including hair growth, periungual and nail plate abnormalities, xerosis, telangiectasias, and pruritus can occur at lower rates [102].
Target | Effect | Drug-incidence | Prevention/treatment | Dose reduction/delay treatment |
---|---|---|---|---|
EGFR | Rash | C 52–89% P 20–50% | Skin moisturizer, sunscreen, hydrocortisone cream, and oral tetracycline | Reduction in 2nd G3 occurrence, delay until ≤ G2 |
Infusion reactions | C 14–21% P-3% | Antihistamines and corticosteroids Low rate, gradual titration | Grade dependent | |
Hypomagnesemia | C 4–38% P 27% | Magnesium replacement | Some G3/4 toxicity delay until recovery | |
Diarrhea | 2% G3/4 | Loperamide, hydration, electrolyte replacement, hospitalization | Reduction in 1st G3 or 2nd G2 occurrence | |
VEGF | Hypertension | B 25% A 42.4% Reg 15% Ram 11% | Blood pressure monitoring, antihypertensive drugs | Cease if G4 or persisting G3 toxicity |
Proteinuria | 18.7% | Screening for proteinuria angiotensin receptor blockers | Discontinue if nephrotic syndrome | |
Hand-foot syndrome | B 16% Reg 14% | Emollient, analgesia | Reduction in 1st G3 or 2nd G2 occurrence, delay until ≤ G1 | |
Thromboembolic events | B 8% | Anticoagulation therapy | Cease bevacizumab |
Adverse effects of any severity with anti-EGFR and -VEGF therapies.
A, aflibercept; B bevacizumab; C cetuximab; EGFR, epidermal growth factor receptor; G grade; P, panitumumab; Ram, ramucirumab; Reg, regorafenib, VEGF, vascular endothelial growth factor.
Infusion reactions commonly occur with cetuximab and should be prevented with premedication, antihistamines, and corticosteroids. Other adverse effects, like hypomagnesemia, ocular toxicities as conjunctivitis and blepharitis, and less commonly diarrhea, can also occur [104]. Toxicity management is grade-depend and, in some cases, should be addressed by a multidisciplinary team.
The main anti-VEGF side effects are cardiovascular and kidney problems (Table 3). Hypertension has been observed at high rates in all phase III studies of anti-VEGF drugs and is normally manageable with standard antihypertensive medications, but this treatment should not be initiated in patients with uncontrolled hypertension. Proteinuria is another side effect, defined as protein content in the urine >300 mg/dL. No standard treatment is established, but anti-angiogenic drugs should be disused if protein content in the urine is >2 g/24 h, and evaluation by a nephrologist should be considered. Hand-foot syndrome is also common with this class of drugs [105].
Bevacizumab has also been associated with other side effects, like thromboembolic events (8%), delayed wound healing, bleeding, fistulae, and gastrointestinal perforation (1.7%). Bevacizumab treatment should be ceased in cases of hemorrhagic events ≥grade 3, pulmonary embolism, cerebrovascular events or arterial insufficiency, arterial thromboembolic events, grade 4 or persistent grade 3 hypertension, nephrotic syndrome, or gastrointestinal perforation [106]. Potentially life-threatening events have occurred only in a small number of patients, with bevacizumab being well tolerated by the majority.
The constitutive activation of RTKs promoted by genomic translocations play an important role in tumorigenesis across different malignancies, including CRC. Examples include ALK, ROS1, and NTRK1–2-3 (NTRK), which altogether occur in 0.2–2.4% of CRCs and may represent new therapeutic targets (Table 4) [107].
Target | Frequency | Clinicopathological features | Testing methods | Agent | Mechanism of action | Current status |
---|---|---|---|---|---|---|
NTRK genes (NTRK 1, NTRK 2, NTRK 3) fusions | 0.5–2.0% in mCRC (4% in MSI-H) | Associated with MSI-H/dMMR; wt BRAF/RAS, elderly females and right sided tumors; associated with poor prognosis; resistance to anti-EGFR monoclonal antibodies | IHC and NGS | Larotrectinib Entrectinib | Small molecule inhibitor targeting TRK proteins | Approved by the FDA and EMA |
ALK/ROS1 | 0.8–2.5% | Associated with MSI-H/dMMR; wt BRAF/RAS, elderly females and right-sided tumors; associated with poor prognosis; resistance to anti-EGFR monoclonal antibodies | FISH, RT-PCR, NGS | Clinical trials; Ceritinib | Small-molecule inhibitor targeting ALK/ROS1 | Under investigation |
FGFR | 3–5% | FGFR3 related with worse prognosis | NGS plus FISH | Regorafenib and newly developing FGFR-specific TKIs | Small-molecule inhibitor targeting FGFR signaling | Under investigation |
c-Met overexpression | Overexpressed in 50–60%, amplified in 10% and mutated in 5% of CRCs | Shorter OS, shorter PFS with bevacizumab treatment; poor prognosis; resistance to anti-EGFR monoclonal antibodies | IHC | Clinical trials | Under investigation | Under investigation |
RET fusions | 0.2% | Worse prognosis, poor treatment response, and reduced OS | IHC and FISH | Vandetanib, cabozantinib | Under investigation | Under investigation |
Summary of new targeted therapies in mCRC.
dMMR, deficient mismatch repair; FGFR, fibroblast growth factor receptor; FISH, fluorescent in situ hybridization; IHC, immunohistochemistry; mCRC, metastatic colorectal cancer; MSI-H, microsatellite instability-high; NGS, next-generation sequencing; OS, overall survival; PFS, progression-free survival; RT-PCR, reverse transcription polymerase chain reaction; TKI, tyrosine kinase inhibitor; wt, wild-type.
The NTRK (neurotrophic tropomyosin receptor kinase) 1, 2, and 3 genes encode three tropomyosin receptor kinase (TRK) receptors —TrkA, TrkB, and TrkC— which are transmembrane proteins [2, 108, 109]. Gene fusions involving those genes lead to constitutively activated NTRK proteins and, consequently, tumorigenesis [107]. The prevalence of NTRK fusions in mCRC is estimated to be 0.5–2.0% [110], but increases to 4% in microsatellite instability-high (MSI-H) mCRC [2].
NTRK gene rearrangements are more commonly detected in non-Lynch syndrome MSI-H/ deficient mismatch repair (dMMR) tumors with MLH1 promoter hypermethylation and wild-type BRAF/KRAS/NRAS, and define a molecular subgroup associated with poor prognosis [111]. They are also more frequent in elderly females with right-sided tumors [107, 109, 112].
Fusion-detection options include targeted DNA and RNA panels, RNA sequencing, FISH, and IHQ [2]. Recent ESMO recommendations for NTRK fusion detection state that, in tumors with low NTRK fusion frequency, as mCRC, detection can be done via one-step next-generation sequencing (NGS) or via IHQ followed by NGS (if IHQ positive) [113].
Larotrectinib and entrectinib are TRK inhibitors approved by the FDA and EMA in more than 10 tumor types. Larotrectinib, a small-molecule inhibitor targeting all three TRK proteins, has been tested in the multicenter single-arm LOXO-TRK-14001, SCOUT, and NAVIGATE clinical trials [111]. Larotrectinib at the dose of 100 mg twice daily showed a good safety profile and good responses (75% of ORR, 1-year PFS of 55%) [114]. In November 2018, the FDA granted accelerated tissue-agnostic approval to larotrectinib for solid tumors with NTRK gene fusions [2, 111, 112] Entrectinib is an oral pan-TRK, -ROS1, and -ALK inhibitor that is clinically active in patients with NTRK-rearranged tumors and is able to penetrate the blood–brain barrier [107]. Three clinical trials (ALKA-372-001, STARTRK-1, and STARTRK-2) have investigated this agent [107]. Pooled analyses of the three trials presented at the ESMO 2018 Congress and ASCO 2019 Meeting showed that entrectinib induced clinically meaningful durable responses in patients with solid tumors with or without metastatic central nervous systemic disease harboring NTRK fusions [111].
The second-generation TRK inhibitor BAY2731954 (formerly known as Loxo-195) and the next-generation ROS1, pan-TRK, and ALK inhibitor repotrectinib are being tested, with promising results [111].
As already shown with BRAF V600E mutations, patients with ALK-, ROS-, and NTRK-rearranged tumors seem to derive no benefit from treatment with anti-EGFR monoclonal antibodies [107]. Additionally, the high prevalence of MSI-H status in rearranged tumors opens the way for evaluation of new combination approaches including targeted (ALK, ROS1, TrkA-B-C) and immunotherapy agents [107].
Regarding resistance mechanisms, a dose-dependent effect seems to affect mutation emergence. Two mutations have been associated with entrectinib resistance: NTRK1 p. G667C and NTRK1 p.G595R [108]. For larotrectinib, three different mutational categories have been described: solvent front mutations (NTRK1 p.G595R, NTRK3 p.G623R); gatekeeper mutations (NTRK1 p.F589L); and xDFG mutations (NTRK1 p.G667S, NTRK3 p.G696A). Novel agents under development intend to overcome NTRK1 p.G595R-mediated resistance to TRK inhibitors [115].
The mesenchymal-epithelial transition (MET) protooncogene (also known as N-methyl-N′-nitroso-guanidine human osteosarcoma transforming gene) encodes for c-MET, a receptor with tyrosine kinase activity targeting HGF. Activation of this pathway has been implicated in CRC metastatic progression [2].
MET receptor tyrosine kinase can be overexpressed in 50–60%, amplified in 10%, and mutated in 5% of CRCs [2]. In a study by Lee et al., c-MET overexpression showed no correlation with primary tumor site, histological type, or molecular aberrations, but correlated with shorter OS and was a predictive biomarker of shorter PFS in bevacizumab-treated patients [3].
EGFR and MET are co-expressed in CRC and MET activation has been implicated in resistance to the anti-EGFR therapy [2, 116]. Inhibition of the HGF/c-Met pathway may improve response to EGFR inhibitors in CRC and combination therapy should be further investigated [116]. This supports the hypothesis that anti-EGFR therapy selects MET-amplified (cetuximab- and panitumumab-resistant) preexisting clones, eventually limiting the efficacy of further anti-EGFR therapies [117].
Multiple clinical trials have evaluated MET inhibition, but several of those conducted in mCRC have been unsuccessful [2]. Treatment strategies targeting HGF and c-Met include HGF antagonists, c-Met and HGF-blocking antibodies, and small-molecule c-Met inhibitors [118].
Although MET genomic aberrations are commonly observed in mCRC, these remain in the research setting [2].
The EML4-ALK fusion gene is produced by inversion in the short arm of chromosome 2, where anaplastic large-cell lymphoma kinase (ALK) joins echinoderm microtubule-associated protein-like 4 (EML4), resulting in a chimeric protein with constitutive ALK activity. ROS1 is an orphan receptor tyrosine kinase phylogenetically related to ALK [110].
ALK and ROS1 gene rearrangements have not been extensively studied in CRC. Around 0.8–2.5% of patients with mCRC have been reported to have either ALK or ROS1 rearrangements [110]. ALK, ROS1, and NTRK fusions occur more frequently in elderly patients with right-sided, RAS wild-type, MSI-H mCRC, and are associated with shorter OS and poor prognosis [107, 110]. The small patient numbers make it challenging to develop a clinical trial of targeted therapies for this patient population [110]. As no FDA-approved agents targeting these genomic alterations exist for CRC patients, basket trials (as the TAPUR trial) may give valuable insights in this setting [112].
RET is a proto-oncogene encoding a transmembrane tyrosine kinase receptor for the glial-derived neurotrophic factor family [110].
RET fusions occur in 0.2% of solid tumors, being very typical in specific tumor types, such as thyroid carcinomas [119]. The effect of RET activation is less clear in CRC, but several studies suggest that it might be associated with worse prognosis, poor treatment response, and reduced OS. Due to rarity of this aberration, clinical trials in CRC are not easy to conduct, with data derived mainly from early trials or case reports [110]. Clinicopathological factors associated with RET fusions include right colon location, older age, RAS and BRAF wild-type status, and MSI-H status [119].
Fibroblast growth factor receptors (FGFRs) are a subfamily of RTKs occurring in approximately 3–5% of CRC patients [112]. Initial evidence shows poor outcomes associated with FGFR3 alterations [120]. There is no evidence of clinicopathological characteristics related to these alterations [120].
Regorafenib, a multi-kinase inhibitor also targeting FGFR, is currently approved by the FDA for metastatic CRC patients who progressed on frontline therapies. This agent can be considered in CRC patients with FGFR alterations while novel FGFR inhibitors are not available [121]. Newly developed, more potent FGFR inhibitors are currently being investigated in multiple solid tumors [112].
Microsatellite instability (MSI) is currently a key biomarker in CRC, with diagnostic, prognostic, and therapeutic implications. For these reasons, MSI analysis is becoming increasingly important and testing for deficient mismatch repair (d-MMR)/MSI is recommended, both for hereditary syndrome screening and due to prognostic and treatment implications [122].
Inactivation of a DNA mismatch repair (MMR) gene (MLH1, MSH2, MSH6, or PMS2) by mutation or transcriptional silencing results in deficient function of the MMR system, responsible for excising DNA mismatches introduced by DNA polymerase during cell division. This activity loss translates in an accumulation of DNA replication errors and mismatches in repeated sequences, leading to hypermutated tumors [123]. In most cases, d-MMR and MSI arise due to sporadic somatic hypermethylation of MLH1 and other genes, but they can also result from germline mutations in MMR genes and from Lynch syndrome in approximately 3% of all CRCs [124].
The MMR system can be assessed through different approaches, as IHC, polymerase chain reaction (PCR)-based assays, and more recently NGS. IHC looks at MLH1, MSH2, MSH6, and PMS2 staining in tumor samples to identify the protein expression loss that characterizes d-MMR [125]. PCR amplification requires both tumor and matched normal samples. Five microsatellite loci have been PCR-amplified and analyzed by capillary electrophoresis. Instability at more than one locus was defined as MSI-high (MSI-H), at a single locus as MSI-low (MSI-L), and absence of instability at any locus as microsatellite stable (MSS), proficient MMR (p-MMR) [126]. NGS detection directly targets certain genes, which are genome sequenced to retrieve information on MSI and MMR and tumor mutational burden (TMB), integrating all information in the same test. NGS requires a smaller sample and is more accurate than PCR. Ethical issues may arise with the use of this technique regarding counseling and consent for additional genetic testing [127]. In CRC, MSI varies according to tumor stage, with higher incidence reported in early stages (20% in stages I-II, 12% in stage III) and lower incidence reported in the metastatic setting (4–5%) [128].
The success of immune checkpoint inhibitors (ICI) in d-MMR over the last years has disclosed a new therapeutic scenario. Endogenous peptides are processed and presented on major histocompatibility complex (MHC) class I molecules on the surface of all cells, being recognized by T cell receptors (TCRs). TCR–MHC signaling pathways are modulated by co-stimulatory or co-inhibitory signals. ICI target co-inhibitory receptors, like cytotoxic T lymphocyte antigen 4 (CTLA-4) and programmed cell death 1 (PD-1) on T cells, or their ligands, as programmed cell death ligand 1 (PDL-1), on tumor and various immune cells [129]. ICI are approved in several malignancies. In mCRC, phase I trials reported response to immune checkpoint therapy in a subgroup of patients with MSI-H, d-MMR, or high TMB [130].
Pembrolizumab is a humanized IgG4 antibody and was the first anti-PD-1 to show efficacy in d-MMR mCRC (Table 5). In the phase II KEYNOTE-016 trial, patients with d-MMR tumors responded better to pembrolizumab (RR of 40%, 20-week PFS of 78%) than MSS tumors (RR of 0%, 20-week PFS of 11%) [131]. In the updated analysis, an ORR of 52%, 2-year PFS of 59%, and OS of 72% was reported for MSI-H CRC [132]. The phase II KEYNOTE-164 trial confirmed the efficacy of pembrolizumab in second-line setting of MSI-H CRC, with an ORR of 33%, median PFS of 2.3 months, and median OS of 31.4 months [133]. Based on these results, pembrolizumab was approved by the FDA for MSI-H/d-MMR unresectable or metastatic CRC after progression on CT. In the phase III KEYNOTE-177 trial, first-line treatment with pembrolizumab in monotherapy significantly reduced the risk of disease progression or death by 40% (HR 0.60; 95% CI 0.45–0.80; p = 0.0004), with a median PFS of 16.5 months versus 8.2 months with CT in MSI-H CRC. The study is ongoing, and OS data will be presented later this year [134]. This led to FDA approval of pembrolizumab in first-line treatment of unresectable or metastatic MSI-H/dMMR CRC.
Setting | Study | Treatment | RR | PFS | OS | Approval |
---|---|---|---|---|---|---|
CT-refractory MSI-H/d-MMR mCRC | Phase II Keynote 164 | Pembrolizumab | 33% | 2.1 m | 31.4 m | FDA (1st line, CT-refractory) |
1st line MSI-H/d-MMR mCRC | Phase III Keynote 177 | Pembrolizumab | 43.8% | 16.5 m | NR | |
CT-refractory MSI-H/d-MMR mCRC | Phase II CheckMate-142 | Nivolumab | 31% | 50% | 73% | FDA (CT-refractory) |
Phase II CheckMate-142 | Nivolumab + ipilimumab | 55% | 71% | 85% | ||
1st line MSI-H/d-MMR mCRC | Phase II CheckMate-142 | Nivolumab + ipilimumab | 60% | 77% | 83% | Not approved |
Immune checkpoint inhibitors in mCRC.
CT, chemotherapy; dMMR, deficient mismatch repair, FDA, Food and Drug Administration; mCRC, metastatic colorectal cancer; MSI-H, microsatellite instability-high; NR, not reached; OS, overall survival; PFS, progression-free survival; RR, response rate.
Nivolumab, a humanized monoclonal IgG4-based PD-1 antibody, showed activity in MSI-H/d-MMR refractory CRC in the phase II CheckMate-142 trial, with an ORR of 31.1% regardless of tumor PD-L1 expression, 1-year PFS of 50%, and OS of 73% [135]. This trial included a cohort of nivolumab in combination with the CTLA-4 inhibitor ipilimumab, which showed a 55% ORR, 71% PFS, and 85% OS. Both nivolumab and the combination of nivolumab plus ipilimumab were approved by the FDA for CT-refractory MSI-H/dMMR mCRC. The immunotherapy doublet was also evaluated in first line in the CheckMate-142 trial, with 1-year PFS and OS of 77% and 83%, respectively, ORR of 60%, and DCR of 84% [136].
Following these studies, MSI status has become a crucial biomarker to define therapeutic options for patients in the metastatic setting.
Other PD-1/PD-L1 inhibitors are under investigation, like atezolizumab, avelumab, and durvalumab, and new immune checkpoint targets are in phase I trials, such as tumor-overexpressed T cell Ig and mucin domain-containing protein 3 (TIM-3), T cell Ig, and T cell-derived lymphocyte activation gene 3 (LAG-3). [137].
Most mCRC patients are MSS/p-MMR and results with ICI have been unsatisfactory, with immune resistance mechanisms not clearly elucidated yet. Several trials have been developed exploring ways to overcome this resistance, including by modulating tumor microenvironment, reducing tumor-specific antigen expression, altering immunosuppressive pathways, and activating other immune checkpoint pathways, immune regulatory cells, and cytokines [138]. Combining immunotherapy with CT, radiotherapy, bispecific antibody therapy, other immune checkpoint modulators, and other targeted agents are among strategies explored. The rationale behind this multimodal approach is the potential synergistic effect of targeting different immune escape pathways, resulting in improved response to ICI and patient outcomes [139].
CT has anti-tumor activity due to the direct cytotoxic effect on cancer cells and to stimulating host immune response, and several clinical trials are ongoing investigating the combination of immunotherapy with CT and targeted agents [140]. Radiotherapy can activate the host immune response by upregulating expression of tumor-specific neoantigens through cell damage and increasing membrane MHC class I expression, and several studies are ongoing in CRC combining radiotherapy with ICI. Another combined strategy is ICI and MEK blockers, considering that MEK blockade seems to increase T cell response via upregulation of PD-L1 expression [141]. Following a phase Ib trial of atezolizumab and the MEK inhibitor cobimetinib in MSS CRC, other trials were conducted, with no significant survival improvement [142]. The CEA CD3 TCB (RG7802, RO6958688) is a novel T-cell bispecific antibody targeting the carcinoembryonic antigen (CEA) on tumor cells and CD3 on T cells, which displays anti-tumor activity, leading to increased intra-tumoral T cell infiltration and activation and PD-1/PD-L1 upregulation. CEA-TCB antibody was tested in phase I trials of MSS CRC plus atezolizumab, showing antitumor activity with acceptable toxicity [143].
Considering immune side effects associated with ICI and their variable efficacy, it is important to identify biomarkers that help predict response to ICI and select potentially sensitive patients that can be candidates for these agents.
PD-L1 expression level is an established biomarker in some malignancies, but the relationship between PD-L1 positivity and response has not been proven in CRC [144]. TMB has emerged as a marker of response to immunotherapy in some tumors, suggesting that tumor cells with high mutational burden generate and present more peptide neoantigens on their MHC class I molecules, increasing T cell infiltration [145]. In CRC, dMMR/MSI-H tumors have a high mutational burden, as well as some pMMR/MSS, which may present an ultramutated phenotype as DNA polymerase epsilon (POLE) mutations, found in ∼1–2% of pMMR CRC. POLE mutations cause an increased immunogenicity and upregulation of immune checkpoint genes, such as PD-1/PD-L1 and CTLA-4, which result in similar clinical responses to dMMR tumors and may predict response to anti-PD-1 therapy [146]. Some case reports link POLE mutations with efficacy to PD-1 blockade, and phase II studies are ongoing in this setting.
The interaction between tumor and microenvironment led to the development of an immunoscore based on calculation of two lymphocytic populations (CD3/CD45-CD8 or CD8/CD45) in the centre and invasive margins of the tumor, which may predict ICI response [147]. Other lines of investigation are being explored, including the study of factors that indicate cytotoxic T cell activity, such as granzymes, perforins, and IFN-γ levels.
CRC is one of the tumor types for which immunotherapy has been less effective. Better knowledge of the molecular immune mechanisms is required to develop predictive biomarkers and effective therapeutic combination strategies, converting “cold” tumors, immune-desert and immunotherapy-resistant, in “hot” tumors, inflamed, infiltrated by the immune system, and immunotherapy responsive.
CRC treatment has changed over the last decades, not only by including different chemotherapy agents and combinations, but mainly because new targeted agents have emerged.
In metastatic setting, anti-EGFR and anti-VEGF drugs are widely used and have shown gains in survival and response rate, an important marker in CRC potentially resectable liver metastases. In contrast, several trials with targeted agents have been conducted in the adjuvant setting, without survival benefit. Immunotherapy emerged as a new treatment option with survival benefit, but at the moment it is only effective in a small portion of patients. Several other agents targeting other pathways are emerging, such as NTRK, c-MET, ALK, ROS1, and FGFR inhibitors, with promising results.
In conclusion, patients with CRC are living longer with targeted treatments, but more information about resistance mechanisms and biomarkers is necessary to extend even more their survival gains.
The authors gracefully acknowledge Joana Cavaco-Silva (jo.cvsilva@gmail.com) for manuscript English language revision and Inês Gomes (ines.gomes@medicina.ulisboa.pt) for Figure 1 graphic drawing.
L. Costa performed consulting activities for Amgen, Novartis and Servier outside the scope of this manuscript. The remaining authors declare no conflicts of interest.
If you are associated with any of the institutions in our list below, you can apply to receive OA publication funds by following the instructions provided in the links.
",metaTitle:"List of Institutions by Country",metaDescription:"If you are associated with any of the institutions in our list below, you can apply to receive OA publication funds by following the instructions provided in the links. However, if your research is financed through any of the below-mentioned funders, please consult their Open Access policies or grant ‘terms and conditions’ to explore ways to cover your publication costs (also accessible by clicking on the link in their title).",metaKeywords:null,canonicalURL:"open-access-funding-institutions-list",contentRaw:'[{"type":"htmlEditorComponent","content":"Book Chapters and Monographs
\\n\\nBook Chapters
\\n\\nMonographs Only
\\n\\n\\n\\nBook Chapters and Monographs
\\n\\nMonographs Only
\\n\\nBook Chapters and Monographs
\\n\\n\\n\\nBook Chapters and Monographs
\\n\\n\\n\\nCorresponding authors will receive a 25% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters. A 20% discount for publishing a long-form monographs, 25% for compacts and 23% for short-form monographs.
\\n\\nCSIC affiliated authors can also take advantage of a central Open Access fund (amounting to 10,000 EUR) to cover up to 50% of the rest of the OAPF until it expires. Effective for chapters accepted from January 1, 2020.
\\n\\nCorresponding authors will receive a 25% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters. A 20% discount for publishing a long-form monographs, 25% for compacts and 23% for short-form monographs.
\\n\\nCorresponding authors will receive a 25% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters. A 20% discount for publishing a long-form monographs, 25% for compacts and 23% for short-form monographs.
\\n\\n\\n\\nCorresponding authors will receive a 25% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters. A 20% discount for publishing a long-form monographs, 25% for compacts and 23% for short-form monographs.
\\n\\nBook Chapters and Monographs
\\n\\nBook Chapters and Monographs
\\n\\nBook Chapters and Monographs
\\n\\nBook Chapters and Monographs
\\n\\nThe Claremont Colleges are pledging funds via the Knowledge Unlatched program to ensure academics can publish Open Access content more easily.
\\n\\nCorresponding authors will receive a 15% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters or monograph publications. To use the discount you will need to verify your institutional email address. These discounts are valid from 2020 to 2022.
\\n\\nThe University of Massachusetts, Amherst is pledging funds via the Knowledge Unlatched program to ensure academics can publish Open Access content more easily.
\\n\\nCorresponding authors will receive a 10% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters or monograph publications. To use the discount you will need to verify your institutional email address. These discounts are valid from 2020 to 2022.
\\n\\nThe University of Surrey is pledging funds via the Knowledge Unlatched program to ensure academics can publish Open Access content more easily.
\\n\\nCorresponding authors will receive a 10% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters or monograph publications. To use the discount you will need to verify your institutional email address. These discounts are valid from 2020 to 2022.
\\n\\nMonographs Only
\\n\\n\\n\\nImportant: You must be a member or grantee of the above listed institutions in order to apply for their Open Access publication funds.
\\n"}]'},components:[{type:"htmlEditorComponent",content:'Book Chapters and Monographs
\n\n\n\nBook Chapters
\n\nMonographs Only
\n\n\n\nBook Chapters and Monographs
\n\nMonographs Only
\n\nBook Chapters and Monographs
\n\n\n\nBook Chapters and Monographs
\n\n\n\nCorresponding authors will receive a 25% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters. A 20% discount for publishing a long-form monographs, 25% for compacts and 23% for short-form monographs.
\n\nCSIC affiliated authors can also take advantage of a central Open Access fund (amounting to 10,000 EUR) to cover up to 50% of the rest of the OAPF until it expires. Effective for chapters accepted from January 1, 2020.
\n\nCorresponding authors will receive a 25% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters. A 20% discount for publishing a long-form monographs, 25% for compacts and 23% for short-form monographs.
\n\nCorresponding authors will receive a 25% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters. A 20% discount for publishing a long-form monographs, 25% for compacts and 23% for short-form monographs.
\n\n\n\nCorresponding authors will receive a 25% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters. A 20% discount for publishing a long-form monographs, 25% for compacts and 23% for short-form monographs.
\n\nBook Chapters and Monographs
\n\nBook Chapters and Monographs
\n\nBook Chapters and Monographs
\n\n\n\nBook Chapters and Monographs
\n\nThe Claremont Colleges are pledging funds via the Knowledge Unlatched program to ensure academics can publish Open Access content more easily.
\n\nCorresponding authors will receive a 15% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters or monograph publications. To use the discount you will need to verify your institutional email address. These discounts are valid from 2020 to 2022.
\n\nThe University of Massachusetts, Amherst is pledging funds via the Knowledge Unlatched program to ensure academics can publish Open Access content more easily.
\n\nCorresponding authors will receive a 10% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters or monograph publications. To use the discount you will need to verify your institutional email address. These discounts are valid from 2020 to 2022.
\n\nThe University of Surrey is pledging funds via the Knowledge Unlatched program to ensure academics can publish Open Access content more easily.
\n\nCorresponding authors will receive a 10% discount on their Open Access Publication Fees (OAPF) for Open Access book chapters or monograph publications. To use the discount you will need to verify your institutional email address. These discounts are valid from 2020 to 2022.
\n\nMonographs Only
\n\n\n\nImportant: You must be a member or grantee of the above listed institutions in order to apply for their Open Access publication funds.
\n'}]},successStories:{items:[]},authorsAndEditors:{filterParams:{sort:"featured,name"},profiles:[{id:"6700",title:"Dr.",name:"Abbass A.",middleName:null,surname:"Hashim",slug:"abbass-a.-hashim",fullName:"Abbass A. Hashim",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/6700/images/1864_n.jpg",biography:"Currently I am carrying out research in several areas of interest, mainly covering work on chemical and bio-sensors, semiconductor thin film device fabrication and characterisation.\nAt the moment I have very strong interest in radiation environmental pollution and bacteriology treatment. The teams of researchers are working very hard to bring novel results in this field. I am also a member of the team in charge for the supervision of Ph.D. students in the fields of development of silicon based planar waveguide sensor devices, study of inelastic electron tunnelling in planar tunnelling nanostructures for sensing applications and development of organotellurium(IV) compounds for semiconductor applications. I am a specialist in data analysis techniques and nanosurface structure. I have served as the editor for many books, been a member of the editorial board in science journals, have published many papers and hold many patents.",institutionString:null,institution:{name:"Sheffield Hallam University",country:{name:"United Kingdom"}}},{id:"54525",title:"Prof.",name:"Abdul Latif",middleName:null,surname:"Ahmad",slug:"abdul-latif-ahmad",fullName:"Abdul Latif Ahmad",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:null},{id:"20567",title:"Prof.",name:"Ado",middleName:null,surname:"Jorio",slug:"ado-jorio",fullName:"Ado Jorio",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Universidade Federal de Minas Gerais",country:{name:"Brazil"}}},{id:"47940",title:"Dr.",name:"Alberto",middleName:null,surname:"Mantovani",slug:"alberto-mantovani",fullName:"Alberto Mantovani",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:null},{id:"12392",title:"Mr.",name:"Alex",middleName:null,surname:"Lazinica",slug:"alex-lazinica",fullName:"Alex Lazinica",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/12392/images/7282_n.png",biography:"Alex Lazinica is the founder and CEO of IntechOpen. After obtaining a Master's degree in Mechanical Engineering, he continued his PhD studies in Robotics at the Vienna University of Technology. Here he worked as a robotic researcher with the university's Intelligent Manufacturing Systems Group as well as a guest researcher at various European universities, including the Swiss Federal Institute of Technology Lausanne (EPFL). During this time he published more than 20 scientific papers, gave presentations, served as a reviewer for major robotic journals and conferences and most importantly he co-founded and built the International Journal of Advanced Robotic Systems- world's first Open Access journal in the field of robotics. Starting this journal was a pivotal point in his career, since it was a pathway to founding IntechOpen - Open Access publisher focused on addressing academic researchers needs. Alex is a personification of IntechOpen key values being trusted, open and entrepreneurial. Today his focus is on defining the growth and development strategy for the company.",institutionString:null,institution:{name:"TU Wien",country:{name:"Austria"}}},{id:"19816",title:"Prof.",name:"Alexander",middleName:null,surname:"Kokorin",slug:"alexander-kokorin",fullName:"Alexander Kokorin",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/19816/images/1607_n.jpg",biography:"Alexander I. Kokorin: born: 1947, Moscow; DSc., PhD; Principal Research Fellow (Research Professor) of Department of Kinetics and Catalysis, N. Semenov Institute of Chemical Physics, Russian Academy of Sciences, Moscow.\r\nArea of research interests: physical chemistry of complex-organized molecular and nanosized systems, including polymer-metal complexes; the surface of doped oxide semiconductors. He is an expert in structural, absorptive, catalytic and photocatalytic properties, in structural organization and dynamic features of ionic liquids, in magnetic interactions between paramagnetic centers. The author or co-author of 3 books, over 200 articles and reviews in scientific journals and books. He is an actual member of the International EPR/ESR Society, European Society on Quantum Solar Energy Conversion, Moscow House of Scientists, of the Board of Moscow Physical Society.",institutionString:null,institution:{name:"Semenov Institute of Chemical Physics",country:{name:"Russia"}}},{id:"62389",title:"PhD.",name:"Ali Demir",middleName:null,surname:"Sezer",slug:"ali-demir-sezer",fullName:"Ali Demir Sezer",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/62389/images/3413_n.jpg",biography:"Dr. Ali Demir Sezer has a Ph.D. from Pharmaceutical Biotechnology at the Faculty of Pharmacy, University of Marmara (Turkey). He is the member of many Pharmaceutical Associations and acts as a reviewer of scientific journals and European projects under different research areas such as: drug delivery systems, nanotechnology and pharmaceutical biotechnology. Dr. Sezer is the author of many scientific publications in peer-reviewed journals and poster communications. Focus of his research activity is drug delivery, physico-chemical characterization and biological evaluation of biopolymers micro and nanoparticles as modified drug delivery system, and colloidal drug carriers (liposomes, nanoparticles etc.).",institutionString:null,institution:{name:"Marmara University",country:{name:"Turkey"}}},{id:"61051",title:"Prof.",name:"Andrea",middleName:null,surname:"Natale",slug:"andrea-natale",fullName:"Andrea Natale",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:null},{id:"100762",title:"Prof.",name:"Andrea",middleName:null,surname:"Natale",slug:"andrea-natale",fullName:"Andrea Natale",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"St David's Medical Center",country:{name:"United States of America"}}},{id:"107416",title:"Dr.",name:"Andrea",middleName:null,surname:"Natale",slug:"andrea-natale",fullName:"Andrea Natale",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:{name:"Texas Cardiac Arrhythmia",country:{name:"United States of America"}}},{id:"64434",title:"Dr.",name:"Angkoon",middleName:null,surname:"Phinyomark",slug:"angkoon-phinyomark",fullName:"Angkoon Phinyomark",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/64434/images/2619_n.jpg",biography:"My name is Angkoon Phinyomark. I received a B.Eng. degree in Computer Engineering with First Class Honors in 2008 from Prince of Songkla University, Songkhla, Thailand, where I received a Ph.D. degree in Electrical Engineering. My research interests are primarily in the area of biomedical signal processing and classification notably EMG (electromyography signal), EOG (electrooculography signal), and EEG (electroencephalography signal), image analysis notably breast cancer analysis and optical coherence tomography, and rehabilitation engineering. I became a student member of IEEE in 2008. During October 2011-March 2012, I had worked at School of Computer Science and Electronic Engineering, University of Essex, Colchester, Essex, United Kingdom. In addition, during a B.Eng. I had been a visiting research student at Faculty of Computer Science, University of Murcia, Murcia, Spain for three months.\n\nI have published over 40 papers during 5 years in refereed journals, books, and conference proceedings in the areas of electro-physiological signals processing and classification, notably EMG and EOG signals, fractal analysis, wavelet analysis, texture analysis, feature extraction and machine learning algorithms, and assistive and rehabilitative devices. I have several computer programming language certificates, i.e. Sun Certified Programmer for the Java 2 Platform 1.4 (SCJP), Microsoft Certified Professional Developer, Web Developer (MCPD), Microsoft Certified Technology Specialist, .NET Framework 2.0 Web (MCTS). I am a Reviewer for several refereed journals and international conferences, such as IEEE Transactions on Biomedical Engineering, IEEE Transactions on Industrial Electronics, Optic Letters, Measurement Science Review, and also a member of the International Advisory Committee for 2012 IEEE Business Engineering and Industrial Applications and 2012 IEEE Symposium on Business, Engineering and Industrial Applications.",institutionString:null,institution:{name:"Joseph Fourier University",country:{name:"France"}}},{id:"55578",title:"Dr.",name:"Antonio",middleName:null,surname:"Jurado-Navas",slug:"antonio-jurado-navas",fullName:"Antonio Jurado-Navas",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/55578/images/4574_n.png",biography:"Antonio Jurado-Navas received the M.S. degree (2002) and the Ph.D. degree (2009) in Telecommunication Engineering, both from the University of Málaga (Spain). He first worked as a consultant at Vodafone-Spain. From 2004 to 2011, he was a Research Assistant with the Communications Engineering Department at the University of Málaga. In 2011, he became an Assistant Professor in the same department. From 2012 to 2015, he was with Ericsson Spain, where he was working on geo-location\ntools for third generation mobile networks. Since 2015, he is a Marie-Curie fellow at the Denmark Technical University. His current research interests include the areas of mobile communication systems and channel modeling in addition to atmospheric optical communications, adaptive optics and statistics",institutionString:null,institution:{name:"University of Malaga",country:{name:"Spain"}}}],filtersByRegion:[{group:"region",caption:"North America",value:1,count:5684},{group:"region",caption:"Middle and South America",value:2,count:5166},{group:"region",caption:"Africa",value:3,count:1682},{group:"region",caption:"Asia",value:4,count:10211},{group:"region",caption:"Australia and Oceania",value:5,count:887},{group:"region",caption:"Europe",value:6,count:15616}],offset:12,limit:12,total:117315},chapterEmbeded:{data:{}},editorApplication:{success:null,errors:{}},ofsBooks:{filterParams:{},books:[{type:"book",id:"7724",title:"Climate Issues in Asia and Africa - Examining Climate, Its Flux, the Consequences, and Society's Responses",subtitle:null,isOpenForSubmission:!0,hash:"c1bd1a5a4dba07b95a5ae5ef0ecf9f74",slug:null,bookSignature:" John P. Tiefenbacher",coverURL:"https://cdn.intechopen.com/books/images_new/7724.jpg",editedByType:null,editors:[{id:"73876",title:"Dr.",name:"John P.",surname:"Tiefenbacher",slug:"john-p.-tiefenbacher",fullName:"John P. Tiefenbacher"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7829",title:"Psychosis - Phenomenology, Psychopathology and Pathophysiology",subtitle:null,isOpenForSubmission:!0,hash:"a211068a33e47af974e3823f33feaa43",slug:null,bookSignature:"Dr. Kenjiro Fukao",coverURL:"https://cdn.intechopen.com/books/images_new/7829.jpg",editedByType:null,editors:[{id:"32519",title:"Dr.",name:"Kenjiro",surname:"Fukao",slug:"kenjiro-fukao",fullName:"Kenjiro Fukao"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7901",title:"Advances in Germ Cell Biology – New Technologies, Applications and Perspectives",subtitle:null,isOpenForSubmission:!0,hash:"4adab31469b82dd5a99eec04dbbe09f2",slug:null,bookSignature:"Ph.D. Sonia Oliveira and Prof. Maria De Lourdes Pereira",coverURL:"https://cdn.intechopen.com/books/images_new/7901.jpg",editedByType:null,editors:[{id:"323848",title:"Ph.D.",name:"Sonia",surname:"Oliveira",slug:"sonia-oliveira",fullName:"Sonia Oliveira"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7921",title:"Optogenetics",subtitle:null,isOpenForSubmission:!0,hash:"3ae7e24d8f03ff3932bceee4b8d3e727",slug:null,bookSignature:"Dr. Thomas Heinbockel",coverURL:"https://cdn.intechopen.com/books/images_new/7921.jpg",editedByType:null,editors:[{id:"70569",title:"Dr.",name:"Thomas",surname:"Heinbockel",slug:"thomas-heinbockel",fullName:"Thomas Heinbockel"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8485",title:"Weather Forecasting",subtitle:null,isOpenForSubmission:!0,hash:"eadbd6f9c26be844062ce5cd3b3eb573",slug:null,bookSignature:"Associate Prof. Muhammad Saifullah",coverURL:"https://cdn.intechopen.com/books/images_new/8485.jpg",editedByType:null,editors:[{id:"320968",title:"Associate Prof.",name:"Muhammad",surname:"Saifullah",slug:"muhammad-saifullah",fullName:"Muhammad Saifullah"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8575",title:"Animal Regeneration",subtitle:null,isOpenForSubmission:!0,hash:"689b9f46c48cd54a2874b8da7386549d",slug:null,bookSignature:"Dr. Hussein Abdelhay Essayed Kaoud",coverURL:"https://cdn.intechopen.com/books/images_new/8575.jpg",editedByType:null,editors:[{id:"265070",title:"Dr.",name:"Hussein Abdelhay",surname:"Essayed Kaoud",slug:"hussein-abdelhay-essayed-kaoud",fullName:"Hussein Abdelhay Essayed Kaoud"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8737",title:"Rabies Virus",subtitle:null,isOpenForSubmission:!0,hash:"49cce3f548da548c718c865feb343509",slug:null,bookSignature:"Dr. Sergey Tkachev",coverURL:"https://cdn.intechopen.com/books/images_new/8737.jpg",editedByType:null,editors:[{id:"61139",title:"Dr.",name:"Sergey",surname:"Tkachev",slug:"sergey-tkachev",fullName:"Sergey Tkachev"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8950",title:"Birds - Challenges and Opportunities for Business, Conservation and Research",subtitle:null,isOpenForSubmission:!0,hash:"404a05af45e47e43871f4a0b1bedc6fd",slug:null,bookSignature:"Dr. Heimo Juhani Mikkola",coverURL:"https://cdn.intechopen.com/books/images_new/8950.jpg",editedByType:null,editors:[{id:"144330",title:"Dr.",name:"Heimo Juhani",surname:"Mikkola",slug:"heimo-juhani-mikkola",fullName:"Heimo Juhani Mikkola"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8977",title:"Protein Kinase - New Opportunities, Challenges and Future Perspectives",subtitle:null,isOpenForSubmission:!0,hash:"6d200cc031706a565b554fdb1c478901",slug:null,bookSignature:"Dr. Rajesh Kumar Singh",coverURL:"https://cdn.intechopen.com/books/images_new/8977.jpg",editedByType:null,editors:[{id:"329385",title:"Dr.",name:"Rajesh",surname:"Singh",slug:"rajesh-singh",fullName:"Rajesh Singh"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9008",title:"Vitamin K - Recent Advances, New Perspectives and Applications for Human Health",subtitle:null,isOpenForSubmission:!0,hash:"8b43add5389ba85743e0a9491e4b9943",slug:null,bookSignature:"Prof. Hiroyuki Kagechika and Dr. Hitoshi Shirakawa",coverURL:"https://cdn.intechopen.com/books/images_new/9008.jpg",editedByType:null,editors:[{id:"180528",title:"Prof.",name:"Hiroyuki",surname:"Kagechika",slug:"hiroyuki-kagechika",fullName:"Hiroyuki Kagechika"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9016",title:"Psychoneuroendocrinology",subtitle:null,isOpenForSubmission:!0,hash:"cb4ce09b8e853bef06c572df42933500",slug:null,bookSignature:"Dr. Ifigenia Kostoglou-Athanassiou",coverURL:"https://cdn.intechopen.com/books/images_new/9016.jpg",editedByType:null,editors:[{id:"307495",title:"Dr.",name:"Ifigenia",surname:"Kostoglou-Athanassiou",slug:"ifigenia-kostoglou-athanassiou",fullName:"Ifigenia Kostoglou-Athanassiou"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9046",title:"Amyloidosis History and Perspectives",subtitle:null,isOpenForSubmission:!0,hash:"371a4ad514bb6d6703406741702a19d0",slug:null,bookSignature:"Dr. Jonathan Harrison",coverURL:"https://cdn.intechopen.com/books/images_new/9046.jpg",editedByType:null,editors:[{id:"340843",title:"Dr.",name:"Jonathan",surname:"Harrison",slug:"jonathan-harrison",fullName:"Jonathan Harrison"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}],filtersByTopic:[{group:"topic",caption:"Agricultural and Biological Sciences",value:5,count:9},{group:"topic",caption:"Biochemistry, Genetics and Molecular Biology",value:6,count:18},{group:"topic",caption:"Business, Management and Economics",value:7,count:2},{group:"topic",caption:"Chemistry",value:8,count:7},{group:"topic",caption:"Computer and Information Science",value:9,count:10},{group:"topic",caption:"Earth and Planetary Sciences",value:10,count:5},{group:"topic",caption:"Engineering",value:11,count:15},{group:"topic",caption:"Environmental Sciences",value:12,count:2},{group:"topic",caption:"Immunology and Microbiology",value:13,count:5},{group:"topic",caption:"Materials Science",value:14,count:4},{group:"topic",caption:"Mathematics",value:15,count:1},{group:"topic",caption:"Medicine",value:16,count:60},{group:"topic",caption:"Nanotechnology and Nanomaterials",value:17,count:1},{group:"topic",caption:"Neuroscience",value:18,count:1},{group:"topic",caption:"Pharmacology, Toxicology and Pharmaceutical Science",value:19,count:6},{group:"topic",caption:"Physics",value:20,count:2},{group:"topic",caption:"Psychology",value:21,count:3},{group:"topic",caption:"Robotics",value:22,count:1},{group:"topic",caption:"Social Sciences",value:23,count:3},{group:"topic",caption:"Technology",value:24,count:1},{group:"topic",caption:"Veterinary Medicine and Science",value:25,count:2}],offset:12,limit:12,total:307},popularBooks:{featuredBooks:[{type:"book",id:"9208",title:"Welding",subtitle:"Modern Topics",isOpenForSubmission:!1,hash:"7d6be076ccf3a3f8bd2ca52d86d4506b",slug:"welding-modern-topics",bookSignature:"Sadek Crisóstomo Absi Alfaro, Wojciech Borek and Błażej Tomiczek",coverURL:"https://cdn.intechopen.com/books/images_new/9208.jpg",editors:[{id:"65292",title:"Prof.",name:"Sadek Crisostomo Absi",middleName:"C. Absi",surname:"Alfaro",slug:"sadek-crisostomo-absi-alfaro",fullName:"Sadek Crisostomo Absi Alfaro"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9139",title:"Topics in Primary Care Medicine",subtitle:null,isOpenForSubmission:!1,hash:"ea774a4d4c1179da92a782e0ae9cde92",slug:"topics-in-primary-care-medicine",bookSignature:"Thomas F. Heston",coverURL:"https://cdn.intechopen.com/books/images_new/9139.jpg",editors:[{id:"217926",title:"Dr.",name:"Thomas F.",middleName:null,surname:"Heston",slug:"thomas-f.-heston",fullName:"Thomas F. Heston"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8697",title:"Virtual Reality and Its Application in Education",subtitle:null,isOpenForSubmission:!1,hash:"ee01b5e387ba0062c6b0d1e9227bda05",slug:"virtual-reality-and-its-application-in-education",bookSignature:"Dragan Cvetković",coverURL:"https://cdn.intechopen.com/books/images_new/8697.jpg",editors:[{id:"101330",title:"Dr.",name:"Dragan",middleName:"Mladen",surname:"Cvetković",slug:"dragan-cvetkovic",fullName:"Dragan Cvetković"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9785",title:"Endometriosis",subtitle:null,isOpenForSubmission:!1,hash:"f457ca61f29cf7e8bc191732c50bb0ce",slug:"endometriosis",bookSignature:"Courtney Marsh",coverURL:"https://cdn.intechopen.com/books/images_new/9785.jpg",editors:[{id:"255491",title:"Dr.",name:"Courtney",middleName:null,surname:"Marsh",slug:"courtney-marsh",fullName:"Courtney Marsh"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9343",title:"Trace Metals in the Environment",subtitle:"New Approaches and Recent Advances",isOpenForSubmission:!1,hash:"ae07e345bc2ce1ebbda9f70c5cd12141",slug:"trace-metals-in-the-environment-new-approaches-and-recent-advances",bookSignature:"Mario Alfonso Murillo-Tovar, Hugo Saldarriaga-Noreña and Agnieszka Saeid",coverURL:"https://cdn.intechopen.com/books/images_new/9343.jpg",editors:[{id:"255959",title:"Dr.",name:"Mario Alfonso",middleName:null,surname:"Murillo-Tovar",slug:"mario-alfonso-murillo-tovar",fullName:"Mario Alfonso Murillo-Tovar"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7831",title:"Sustainability in Urban Planning and Design",subtitle:null,isOpenForSubmission:!1,hash:"c924420492c8c2c9751e178d025f4066",slug:"sustainability-in-urban-planning-and-design",bookSignature:"Amjad Almusaed, Asaad Almssad and Linh Truong - Hong",coverURL:"https://cdn.intechopen.com/books/images_new/7831.jpg",editors:[{id:"110471",title:"Dr.",name:"Amjad",middleName:"Zaki",surname:"Almusaed",slug:"amjad-almusaed",fullName:"Amjad Almusaed"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8468",title:"Sheep Farming",subtitle:"An Approach to Feed, Growth and Sanity",isOpenForSubmission:!1,hash:"838f08594850bc04aa14ec873ed1b96f",slug:"sheep-farming-an-approach-to-feed-growth-and-sanity",bookSignature:"António Monteiro",coverURL:"https://cdn.intechopen.com/books/images_new/8468.jpg",editors:[{id:"190314",title:"Prof.",name:"António",middleName:"Cardoso",surname:"Monteiro",slug:"antonio-monteiro",fullName:"António Monteiro"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8816",title:"Financial Crises",subtitle:"A Selection of Readings",isOpenForSubmission:!1,hash:"6f2f49fb903656e4e54280c79fabd10c",slug:"financial-crises-a-selection-of-readings",bookSignature:"Stelios Markoulis",coverURL:"https://cdn.intechopen.com/books/images_new/8816.jpg",editors:[{id:"237863",title:"Dr.",name:"Stelios",middleName:null,surname:"Markoulis",slug:"stelios-markoulis",fullName:"Stelios Markoulis"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9376",title:"Contemporary Developments and Perspectives in International Health Security",subtitle:"Volume 1",isOpenForSubmission:!1,hash:"b9a00b84cd04aae458fb1d6c65795601",slug:"contemporary-developments-and-perspectives-in-international-health-security-volume-1",bookSignature:"Stanislaw P. Stawicki, Michael S. Firstenberg, Sagar C. Galwankar, Ricardo Izurieta and Thomas Papadimos",coverURL:"https://cdn.intechopen.com/books/images_new/9376.jpg",editors:[{id:"181694",title:"Dr.",name:"Stanislaw P.",middleName:null,surname:"Stawicki",slug:"stanislaw-p.-stawicki",fullName:"Stanislaw P. Stawicki"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7769",title:"Medical Isotopes",subtitle:null,isOpenForSubmission:!1,hash:"f8d3c5a6c9a42398e56b4e82264753f7",slug:"medical-isotopes",bookSignature:"Syed Ali Raza Naqvi and Muhammad Babar Imrani",coverURL:"https://cdn.intechopen.com/books/images_new/7769.jpg",editors:[{id:"259190",title:"Dr.",name:"Syed Ali Raza",middleName:null,surname:"Naqvi",slug:"syed-ali-raza-naqvi",fullName:"Syed Ali Raza Naqvi"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9279",title:"Concepts, Applications and Emerging Opportunities in Industrial Engineering",subtitle:null,isOpenForSubmission:!1,hash:"9bfa87f9b627a5468b7c1e30b0eea07a",slug:"concepts-applications-and-emerging-opportunities-in-industrial-engineering",bookSignature:"Gary Moynihan",coverURL:"https://cdn.intechopen.com/books/images_new/9279.jpg",editors:[{id:"16974",title:"Dr.",name:"Gary",middleName:null,surname:"Moynihan",slug:"gary-moynihan",fullName:"Gary Moynihan"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7807",title:"A Closer Look at Organizational Culture in Action",subtitle:null,isOpenForSubmission:!1,hash:"05c608b9271cc2bc711f4b28748b247b",slug:"a-closer-look-at-organizational-culture-in-action",bookSignature:"Süleyman Davut Göker",coverURL:"https://cdn.intechopen.com/books/images_new/7807.jpg",editors:[{id:"190035",title:"Associate Prof.",name:"Süleyman Davut",middleName:null,surname:"Göker",slug:"suleyman-davut-goker",fullName:"Süleyman Davut Göker"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}}],offset:12,limit:12,total:5131},hotBookTopics:{hotBooks:[],offset:0,limit:12,total:null},publish:{},publishingProposal:{success:null,errors:{}},books:{featuredBooks:[{type:"book",id:"9208",title:"Welding",subtitle:"Modern Topics",isOpenForSubmission:!1,hash:"7d6be076ccf3a3f8bd2ca52d86d4506b",slug:"welding-modern-topics",bookSignature:"Sadek Crisóstomo Absi Alfaro, Wojciech Borek and Błażej Tomiczek",coverURL:"https://cdn.intechopen.com/books/images_new/9208.jpg",editors:[{id:"65292",title:"Prof.",name:"Sadek Crisostomo Absi",middleName:"C. Absi",surname:"Alfaro",slug:"sadek-crisostomo-absi-alfaro",fullName:"Sadek Crisostomo Absi Alfaro"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9139",title:"Topics in Primary Care Medicine",subtitle:null,isOpenForSubmission:!1,hash:"ea774a4d4c1179da92a782e0ae9cde92",slug:"topics-in-primary-care-medicine",bookSignature:"Thomas F. Heston",coverURL:"https://cdn.intechopen.com/books/images_new/9139.jpg",editors:[{id:"217926",title:"Dr.",name:"Thomas F.",middleName:null,surname:"Heston",slug:"thomas-f.-heston",fullName:"Thomas F. Heston"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"8697",title:"Virtual Reality and Its Application in Education",subtitle:null,isOpenForSubmission:!1,hash:"ee01b5e387ba0062c6b0d1e9227bda05",slug:"virtual-reality-and-its-application-in-education",bookSignature:"Dragan Cvetković",coverURL:"https://cdn.intechopen.com/books/images_new/8697.jpg",editors:[{id:"101330",title:"Dr.",name:"Dragan",middleName:"Mladen",surname:"Cvetković",slug:"dragan-cvetkovic",fullName:"Dragan Cvetković"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9343",title:"Trace Metals in the Environment",subtitle:"New Approaches and Recent Advances",isOpenForSubmission:!1,hash:"ae07e345bc2ce1ebbda9f70c5cd12141",slug:"trace-metals-in-the-environment-new-approaches-and-recent-advances",bookSignature:"Mario Alfonso Murillo-Tovar, Hugo Saldarriaga-Noreña and Agnieszka Saeid",coverURL:"https://cdn.intechopen.com/books/images_new/9343.jpg",editors:[{id:"255959",title:"Dr.",name:"Mario Alfonso",middleName:null,surname:"Murillo-Tovar",slug:"mario-alfonso-murillo-tovar",fullName:"Mario Alfonso Murillo-Tovar"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9785",title:"Endometriosis",subtitle:null,isOpenForSubmission:!1,hash:"f457ca61f29cf7e8bc191732c50bb0ce",slug:"endometriosis",bookSignature:"Courtney Marsh",coverURL:"https://cdn.intechopen.com/books/images_new/9785.jpg",editors:[{id:"255491",title:"Dr.",name:"Courtney",middleName:null,surname:"Marsh",slug:"courtney-marsh",fullName:"Courtney Marsh"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7831",title:"Sustainability in Urban Planning and Design",subtitle:null,isOpenForSubmission:!1,hash:"c924420492c8c2c9751e178d025f4066",slug:"sustainability-in-urban-planning-and-design",bookSignature:"Amjad Almusaed, Asaad Almssad and Linh Truong - Hong",coverURL:"https://cdn.intechopen.com/books/images_new/7831.jpg",editors:[{id:"110471",title:"Dr.",name:"Amjad",middleName:"Zaki",surname:"Almusaed",slug:"amjad-almusaed",fullName:"Amjad Almusaed"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9376",title:"Contemporary Developments and Perspectives in International Health Security",subtitle:"Volume 1",isOpenForSubmission:!1,hash:"b9a00b84cd04aae458fb1d6c65795601",slug:"contemporary-developments-and-perspectives-in-international-health-security-volume-1",bookSignature:"Stanislaw P. Stawicki, Michael S. Firstenberg, Sagar C. Galwankar, Ricardo Izurieta and Thomas Papadimos",coverURL:"https://cdn.intechopen.com/books/images_new/9376.jpg",editors:[{id:"181694",title:"Dr.",name:"Stanislaw P.",middleName:null,surname:"Stawicki",slug:"stanislaw-p.-stawicki",fullName:"Stanislaw P. Stawicki"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7769",title:"Medical Isotopes",subtitle:null,isOpenForSubmission:!1,hash:"f8d3c5a6c9a42398e56b4e82264753f7",slug:"medical-isotopes",bookSignature:"Syed Ali Raza Naqvi and Muhammad Babar Imrani",coverURL:"https://cdn.intechopen.com/books/images_new/7769.jpg",editors:[{id:"259190",title:"Dr.",name:"Syed Ali Raza",middleName:null,surname:"Naqvi",slug:"syed-ali-raza-naqvi",fullName:"Syed Ali Raza Naqvi"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"9279",title:"Concepts, Applications and Emerging Opportunities in Industrial Engineering",subtitle:null,isOpenForSubmission:!1,hash:"9bfa87f9b627a5468b7c1e30b0eea07a",slug:"concepts-applications-and-emerging-opportunities-in-industrial-engineering",bookSignature:"Gary Moynihan",coverURL:"https://cdn.intechopen.com/books/images_new/9279.jpg",editors:[{id:"16974",title:"Dr.",name:"Gary",middleName:null,surname:"Moynihan",slug:"gary-moynihan",fullName:"Gary Moynihan"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}},{type:"book",id:"7807",title:"A Closer Look at Organizational Culture in Action",subtitle:null,isOpenForSubmission:!1,hash:"05c608b9271cc2bc711f4b28748b247b",slug:"a-closer-look-at-organizational-culture-in-action",bookSignature:"Süleyman Davut Göker",coverURL:"https://cdn.intechopen.com/books/images_new/7807.jpg",editors:[{id:"190035",title:"Associate Prof.",name:"Süleyman Davut",middleName:null,surname:"Göker",slug:"suleyman-davut-goker",fullName:"Süleyman Davut Göker"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}}],latestBooks:[{type:"book",id:"7434",title:"Molecular Biotechnology",subtitle:null,isOpenForSubmission:!1,hash:"eceede809920e1ec7ecadd4691ede2ec",slug:"molecular-biotechnology",bookSignature:"Sergey Sedykh",coverURL:"https://cdn.intechopen.com/books/images_new/7434.jpg",editedByType:"Edited by",editors:[{id:"178316",title:"Ph.D.",name:"Sergey",middleName:null,surname:"Sedykh",slug:"sergey-sedykh",fullName:"Sergey Sedykh"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8545",title:"Animal Reproduction in Veterinary Medicine",subtitle:null,isOpenForSubmission:!1,hash:"13aaddf5fdbbc78387e77a7da2388bf6",slug:"animal-reproduction-in-veterinary-medicine",bookSignature:"Faruk Aral, Rita Payan-Carreira and Miguel Quaresma",coverURL:"https://cdn.intechopen.com/books/images_new/8545.jpg",editedByType:"Edited by",editors:[{id:"25600",title:"Prof.",name:"Faruk",middleName:null,surname:"Aral",slug:"faruk-aral",fullName:"Faruk Aral"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9569",title:"Methods in Molecular Medicine",subtitle:null,isOpenForSubmission:!1,hash:"691d3f3c4ac25a8093414e9b270d2843",slug:"methods-in-molecular-medicine",bookSignature:"Yusuf Tutar",coverURL:"https://cdn.intechopen.com/books/images_new/9569.jpg",editedByType:"Edited by",editors:[{id:"158492",title:"Prof.",name:"Yusuf",middleName:null,surname:"Tutar",slug:"yusuf-tutar",fullName:"Yusuf Tutar"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9839",title:"Outdoor Recreation",subtitle:"Physiological and Psychological Effects on Health",isOpenForSubmission:!1,hash:"5f5a0d64267e32567daffa5b0c6a6972",slug:"outdoor-recreation-physiological-and-psychological-effects-on-health",bookSignature:"Hilde G. Nielsen",coverURL:"https://cdn.intechopen.com/books/images_new/9839.jpg",editedByType:"Edited by",editors:[{id:"158692",title:"Ph.D.",name:"Hilde G.",middleName:null,surname:"Nielsen",slug:"hilde-g.-nielsen",fullName:"Hilde G. Nielsen"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7802",title:"Modern Slavery and Human Trafficking",subtitle:null,isOpenForSubmission:!1,hash:"587a0b7fb765f31cc98de33c6c07c2e0",slug:"modern-slavery-and-human-trafficking",bookSignature:"Jane Reeves",coverURL:"https://cdn.intechopen.com/books/images_new/7802.jpg",editedByType:"Edited by",editors:[{id:"211328",title:"Prof.",name:"Jane",middleName:null,surname:"Reeves",slug:"jane-reeves",fullName:"Jane Reeves"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8063",title:"Food Security in Africa",subtitle:null,isOpenForSubmission:!1,hash:"8cbf3d662b104d19db2efc9d59249efc",slug:"food-security-in-africa",bookSignature:"Barakat Mahmoud",coverURL:"https://cdn.intechopen.com/books/images_new/8063.jpg",editedByType:"Edited by",editors:[{id:"92016",title:"Dr.",name:"Barakat",middleName:null,surname:"Mahmoud",slug:"barakat-mahmoud",fullName:"Barakat Mahmoud"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"10118",title:"Plant Stress Physiology",subtitle:null,isOpenForSubmission:!1,hash:"c68b09d2d2634fc719ae3b9a64a27839",slug:"plant-stress-physiology",bookSignature:"Akbar Hossain",coverURL:"https://cdn.intechopen.com/books/images_new/10118.jpg",editedByType:"Edited by",editors:[{id:"280755",title:"Dr.",name:"Akbar",middleName:null,surname:"Hossain",slug:"akbar-hossain",fullName:"Akbar Hossain"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9157",title:"Neurodegenerative Diseases",subtitle:"Molecular Mechanisms and Current Therapeutic Approaches",isOpenForSubmission:!1,hash:"bc8be577966ef88735677d7e1e92ed28",slug:"neurodegenerative-diseases-molecular-mechanisms-and-current-therapeutic-approaches",bookSignature:"Nagehan Ersoy Tunalı",coverURL:"https://cdn.intechopen.com/books/images_new/9157.jpg",editedByType:"Edited by",editors:[{id:"82778",title:"Ph.D.",name:"Nagehan",middleName:null,surname:"Ersoy Tunalı",slug:"nagehan-ersoy-tunali",fullName:"Nagehan Ersoy Tunalı"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9961",title:"Data Mining",subtitle:"Methods, Applications and Systems",isOpenForSubmission:!1,hash:"ed79fb6364f2caf464079f94a0387146",slug:"data-mining-methods-applications-and-systems",bookSignature:"Derya Birant",coverURL:"https://cdn.intechopen.com/books/images_new/9961.jpg",editedByType:"Edited by",editors:[{id:"15609",title:"Dr.",name:"Derya",middleName:null,surname:"Birant",slug:"derya-birant",fullName:"Derya Birant"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8686",title:"Direct Torque Control Strategies of Electrical Machines",subtitle:null,isOpenForSubmission:!1,hash:"b6ad22b14db2b8450228545d3d4f6b1a",slug:"direct-torque-control-strategies-of-electrical-machines",bookSignature:"Fatma Ben Salem",coverURL:"https://cdn.intechopen.com/books/images_new/8686.jpg",editedByType:"Edited by",editors:[{id:"295623",title:"Associate Prof.",name:"Fatma",middleName:null,surname:"Ben Salem",slug:"fatma-ben-salem",fullName:"Fatma Ben Salem"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},subject:{topic:{id:"13",title:"Immunology and Microbiology",slug:"immunology-and-microbiology",parent:{title:"Life Sciences",slug:"life-sciences"},numberOfBooks:65,numberOfAuthorsAndEditors:1808,numberOfWosCitations:762,numberOfCrossrefCitations:741,numberOfDimensionsCitations:1754,videoUrl:null,fallbackUrl:null,description:null},booksByTopicFilter:{topicSlug:"immunology-and-microbiology",sort:"-publishedDate",limit:12,offset:0},booksByTopicCollection:[{type:"book",id:"9742",title:"Ubiquitin",subtitle:"Proteasome Pathway",isOpenForSubmission:!1,hash:"af6880d3a5571da1377ac8f6373b9e82",slug:"ubiquitin-proteasome-pathway",bookSignature:"Xianquan Zhan",coverURL:"https://cdn.intechopen.com/books/images_new/9742.jpg",editedByType:"Edited by",editors:[{id:"223233",title:"Prof.",name:"Xianquan",middleName:null,surname:"Zhan",slug:"xianquan-zhan",fullName:"Xianquan Zhan"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8967",title:"Bacterial Biofilms",subtitle:null,isOpenForSubmission:!1,hash:"e692b520263526cca2b37092c3e8d0b4",slug:"bacterial-biofilms",bookSignature:"Sadik Dincer, Melis Sümengen Özdenefe and Afet Arkut",coverURL:"https://cdn.intechopen.com/books/images_new/8967.jpg",editedByType:"Edited by",editors:[{id:"188141",title:"Prof.",name:"Sadik",middleName:null,surname:"Dincer",slug:"sadik-dincer",fullName:"Sadik Dincer"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9294",title:"Fluorescence Methods for Investigation of Living Cells and Microorganisms",subtitle:null,isOpenForSubmission:!1,hash:"a97a566a3a19eb9e0c9ba61042bb06c5",slug:"fluorescence-methods-for-investigation-of-living-cells-and-microorganisms",bookSignature:"Natalia Grigoryeva",coverURL:"https://cdn.intechopen.com/books/images_new/9294.jpg",editedByType:"Edited by",editors:[{id:"239430",title:"Dr.",name:"Natalia",middleName:"Yu.",surname:"Grigoryeva",slug:"natalia-grigoryeva",fullName:"Natalia Grigoryeva"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9352",title:"Proteoforms",subtitle:"Concept and Applications in Medical Sciences",isOpenForSubmission:!1,hash:"0f0288da2d32c0c0fcda6be0d4d45d67",slug:"proteoforms-concept-and-applications-in-medical-sciences",bookSignature:"Xianquan Zhan",coverURL:"https://cdn.intechopen.com/books/images_new/9352.jpg",editedByType:"Edited by",editors:[{id:"223233",title:"Prof.",name:"Xianquan",middleName:null,surname:"Zhan",slug:"xianquan-zhan",fullName:"Xianquan Zhan"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8997",title:"Microorganisms",subtitle:null,isOpenForSubmission:!1,hash:"d4bb9c77b89f8baf2716d1fb84c5bd9f",slug:"microorganisms",bookSignature:"Miroslav Blumenberg, Mona Shaaban, Abdelaziz Elgaml",coverURL:"https://cdn.intechopen.com/books/images_new/8997.jpg",editedByType:"Edited by",editors:[{id:"31610",title:"Dr.",name:"Miroslav",middleName:null,surname:"Blumenberg",slug:"miroslav-blumenberg",fullName:"Miroslav Blumenberg"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"9025",title:"Parasitology and Microbiology Research",subtitle:null,isOpenForSubmission:!1,hash:"d9a211396d44f07d2748e147786a2c8b",slug:"parasitology-and-microbiology-research",bookSignature:"Gilberto Antonio Bastidas Pacheco and Asghar Ali Kamboh",coverURL:"https://cdn.intechopen.com/books/images_new/9025.jpg",editedByType:"Edited by",editors:[{id:"238219",title:"Dr.",name:"Gilberto Antonio",middleName:null,surname:"Bastidas Pacheco",slug:"gilberto-antonio-bastidas-pacheco",fullName:"Gilberto Antonio Bastidas Pacheco"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8590",title:"Macrophage Activation",subtitle:"Biology and Disease",isOpenForSubmission:!1,hash:"e15abd1b0e08f1b67d33592999c52c32",slug:"macrophage-activation-biology-and-disease",bookSignature:"Khalid Hussain Bhat",coverURL:"https://cdn.intechopen.com/books/images_new/8590.jpg",editedByType:"Edited by",editors:[{id:"162478",title:"Dr.",name:"Khalid Hussain",middleName:null,surname:"Bhat",slug:"khalid-hussain-bhat",fullName:"Khalid Hussain Bhat"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8422",title:"Metagenomics",subtitle:"Basics, Methods and Applications",isOpenForSubmission:!1,hash:"82c6409553747ffccd6075f9420e3175",slug:"metagenomics-basics-methods-and-applications",bookSignature:"Wael N. Hozzein",coverURL:"https://cdn.intechopen.com/books/images_new/8422.jpg",editedByType:"Edited by",editors:[{id:"189233",title:"Prof.",name:"Wael N.",middleName:"Nabil",surname:"Hozzein",slug:"wael-n.-hozzein",fullName:"Wael N. Hozzein"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"6967",title:"Prebiotics and Probiotics",subtitle:"Potential Benefits in Nutrition and Health",isOpenForSubmission:!1,hash:"11781d6b1c070edcf204518e632033be",slug:"prebiotics-and-probiotics-potential-benefits-in-nutrition-and-health",bookSignature:"Elena Franco-Robles and Joel Ramírez-Emiliano",coverURL:"https://cdn.intechopen.com/books/images_new/6967.jpg",editedByType:"Edited by",editors:[{id:"219102",title:"Dr.",name:"Elena",middleName:null,surname:"Franco-Robles",slug:"elena-franco-robles",fullName:"Elena Franco-Robles"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"7013",title:"Immunohistochemistry",subtitle:"The Ageless Biotechnology",isOpenForSubmission:!1,hash:"cd11a72871d4b30ec4855a33d49adf3f",slug:"immunohistochemistry-the-ageless-biotechnology",bookSignature:"Charles F. Streckfus",coverURL:"https://cdn.intechopen.com/books/images_new/7013.jpg",editedByType:"Edited by",editors:[{id:"29033",title:"Prof.",name:"Charles",middleName:"F.",surname:"Streckfus",slug:"charles-streckfus",fullName:"Charles Streckfus"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8019",title:"Alginates",subtitle:"Recent Uses of This Natural Polymer",isOpenForSubmission:!1,hash:"61ea5c1aef462684a3b2215631b7dbf2",slug:"alginates-recent-uses-of-this-natural-polymer",bookSignature:"Leonel Pereira",coverURL:"https://cdn.intechopen.com/books/images_new/8019.jpg",editedByType:"Edited by",editors:[{id:"279788",title:"Dr.",name:"Leonel",middleName:null,surname:"Pereira",slug:"leonel-pereira",fullName:"Leonel Pereira"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"8806",title:"Biology of Trypanosoma cruzi",subtitle:null,isOpenForSubmission:!1,hash:"514ab85661e01a47575e845792ef5bdc",slug:"biology-of-em-trypanosoma-cruzi-em-",bookSignature:"Wanderley De Souza",coverURL:"https://cdn.intechopen.com/books/images_new/8806.jpg",editedByType:"Edited by",editors:[{id:"161922",title:"Dr.",name:"Wanderley",middleName:null,surname:"De Souza",slug:"wanderley-de-souza",fullName:"Wanderley De Souza"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}],booksByTopicTotal:65,mostCitedChapters:[{id:"39599",doi:"10.5772/50046",title:"Encapsulation Technology to Protect Probiotic Bacteria",slug:"encapsulation-technology-to-protect-probiotic-bacteria",totalDownloads:11213,totalCrossrefCites:23,totalDimensionsCites:53,book:{slug:"probiotics",title:"Probiotics",fullTitle:"Probiotics"},signatures:"María Chávarri, Izaskun Marañón and María Carmen Villarán",authors:[{id:"150285",title:"Dr.",name:"María",middleName:null,surname:"Chávarri Hueda",slug:"maria-chavarri-hueda",fullName:"María Chávarri Hueda"},{id:"151613",title:"MSc.",name:"Izaskun",middleName:null,surname:"Marañon",slug:"izaskun-maranon",fullName:"Izaskun Marañon"},{id:"151621",title:"Dr.",name:"Mª Carmen",middleName:null,surname:"Villarán",slug:"ma-carmen-villaran",fullName:"Mª Carmen Villarán"}]},{id:"39607",doi:"10.5772/50121",title:"Recent Application of Probiotics in Food and Agricultural Science",slug:"recent-application-of-probiotics-in-food-and-agricultural-science",totalDownloads:9076,totalCrossrefCites:17,totalDimensionsCites:48,book:{slug:"probiotics",title:"Probiotics",fullTitle:"Probiotics"},signatures:"Danfeng Song, Salam Ibrahim and Saeed Hayek",authors:[{id:"107905",title:"Prof.",name:"Salam",middleName:null,surname:"Ibrahim",slug:"salam-ibrahim",fullName:"Salam Ibrahim"},{id:"150202",title:"Dr.",name:"Danfeng",middleName:null,surname:"Song",slug:"danfeng-song",fullName:"Danfeng Song"},{id:"151025",title:"MSc.",name:"Saeed",middleName:null,surname:"Hayek",slug:"saeed-hayek",fullName:"Saeed Hayek"}]},{id:"33740",doi:"10.5772/35797",title:"Interferences in Immunoassays",slug:"interference-in-immunoassays",totalDownloads:16951,totalCrossrefCites:13,totalDimensionsCites:42,book:{slug:"advances-in-immunoassay-technology",title:"Advances in Immunoassay Technology",fullTitle:"Advances in Immunoassay Technology"},signatures:"Johan Schiettecatte, Ellen Anckaert and Johan Smitz",authors:[{id:"105883",title:"Mr.",name:"Johan",middleName:null,surname:"Schiettecatte",slug:"johan-schiettecatte",fullName:"Johan Schiettecatte"},{id:"113099",title:"Dr.",name:"Ellen",middleName:null,surname:"Anckaert",slug:"ellen-anckaert",fullName:"Ellen Anckaert"},{id:"113100",title:"Prof.",name:"Johan",middleName:null,surname:"Smitz",slug:"johan-smitz",fullName:"Johan Smitz"}]}],mostDownloadedChaptersLast30Days:[{id:"54154",title:"Staphylococcus aureus: Overview of Bacteriology, Clinical Diseases, Epidemiology, Antibiotic Resistance and Therapeutic Approach",slug:"staphylococcus-aureus-overview-of-bacteriology-clinical-diseases-epidemiology-antibiotic-resistance-",totalDownloads:5104,totalCrossrefCites:5,totalDimensionsCites:9,book:{slug:"frontiers-in-i-staphylococcus-aureus-i-",title:"Frontiers in Staphylococcus aureus",fullTitle:"Frontiers in Staphylococcus aureus"},signatures:"Arumugam Gnanamani, Periasamy Hariharan and Maneesh Paul-\nSatyaseela",authors:[{id:"192829",title:"Dr.",name:"Arumugam",middleName:null,surname:"Gnanamani",slug:"arumugam-gnanamani",fullName:"Arumugam Gnanamani"},{id:"204388",title:"Dr.",name:"Periasamy",middleName:null,surname:"Hariharan",slug:"periasamy-hariharan",fullName:"Periasamy Hariharan"},{id:"204389",title:"Dr.",name:"Maneesh",middleName:null,surname:"Paul-Satyaseela",slug:"maneesh-paul-satyaseela",fullName:"Maneesh Paul-Satyaseela"}]},{id:"62553",title:"Antibiotic Use in Poultry Production and Its Effects on Bacterial Resistance",slug:"antibiotic-use-in-poultry-production-and-its-effects-on-bacterial-resistance",totalDownloads:4918,totalCrossrefCites:15,totalDimensionsCites:36,book:{slug:"antimicrobial-resistance-a-global-threat",title:"Antimicrobial Resistance",fullTitle:"Antimicrobial Resistance - A Global Threat"},signatures:"Christian Agyare, Vivian Etsiapa Boamah, Crystal Ngofi Zumbi and\nFrank Boateng Osei",authors:[{id:"182058",title:"Dr.",name:"Christian",middleName:null,surname:"Agyare",slug:"christian-agyare",fullName:"Christian Agyare"},{id:"261271",title:"MSc.",name:"Crystal Ngofi",middleName:null,surname:"Zumbi",slug:"crystal-ngofi-zumbi",fullName:"Crystal Ngofi Zumbi"},{id:"261272",title:"MSc.",name:"Frank Boateng",middleName:null,surname:"Osei",slug:"frank-boateng-osei",fullName:"Frank Boateng Osei"},{id:"261273",title:"Dr.",name:"Vivian Etsiapa",middleName:null,surname:"Boamah",slug:"vivian-etsiapa-boamah",fullName:"Vivian Etsiapa Boamah"}]},{id:"65613",title:"The Methods for Detection of Biofilm and Screening Antibiofilm Activity of Agents",slug:"the-methods-for-detection-of-biofilm-and-screening-antibiofilm-activity-of-agents",totalDownloads:6467,totalCrossrefCites:3,totalDimensionsCites:6,book:{slug:"antimicrobials-antibiotic-resistance-antibiofilm-strategies-and-activity-methods",title:"Antimicrobials, Antibiotic Resistance, Antibiofilm Strategies and Activity Methods",fullTitle:"Antimicrobials, Antibiotic Resistance, Antibiofilm Strategies and Activity Methods"},signatures:"Sahra Kırmusaoğlu",authors:[{id:"179460",title:"Dr.",name:"Sahra",middleName:null,surname:"Kırmusaoğlu",slug:"sahra-kirmusaoglu",fullName:"Sahra Kırmusaoğlu"}]},{id:"56849",title:"Physiology and Pathology of Innate Immune Response Against Pathogens",slug:"physiology-and-pathology-of-innate-immune-response-against-pathogens",totalDownloads:4842,totalCrossrefCites:9,totalDimensionsCites:14,book:{slug:"physiology-and-pathology-of-immunology",title:"Physiology and Pathology of Immunology",fullTitle:"Physiology and Pathology of Immunology"},signatures:"José Luis Muñoz Carrillo, Flor Pamela Castro García, Oscar\nGutiérrez Coronado, María Alejandra Moreno García and Juan\nFrancisco Contreras Cordero",authors:[{id:"214236",title:"Dr.",name:"Jose Luis",middleName:null,surname:"Muñoz-Carrillo",slug:"jose-luis-munoz-carrillo",fullName:"Jose Luis Muñoz-Carrillo"},{id:"216080",title:"Dr.",name:"Alejandra",middleName:null,surname:"Moreno-García",slug:"alejandra-moreno-garcia",fullName:"Alejandra Moreno-García"},{id:"216081",title:"Dr.",name:"Oscar",middleName:null,surname:"Gutiérrez-Coronado",slug:"oscar-gutierrez-coronado",fullName:"Oscar Gutiérrez-Coronado"},{id:"216082",title:"Dr.",name:"Pamela",middleName:null,surname:"Castro-García",slug:"pamela-castro-garcia",fullName:"Pamela Castro-García"},{id:"220717",title:"Dr.",name:"Juan Francisco",middleName:null,surname:"Contreras Cordero",slug:"juan-francisco-contreras-cordero",fullName:"Juan Francisco Contreras Cordero"}]},{id:"65914",title:"Introductory Chapter: The Action Mechanisms of Antibiotics and Antibiotic Resistance",slug:"introductory-chapter-the-action-mechanisms-of-antibiotics-and-antibiotic-resistance",totalDownloads:2848,totalCrossrefCites:1,totalDimensionsCites:0,book:{slug:"antimicrobials-antibiotic-resistance-antibiofilm-strategies-and-activity-methods",title:"Antimicrobials, Antibiotic Resistance, Antibiofilm Strategies and Activity Methods",fullTitle:"Antimicrobials, Antibiotic Resistance, Antibiofilm Strategies and Activity Methods"},signatures:"Sahra Kırmusaoğlu, Nesrin Gareayaghi and Bekir S. Kocazeybek",authors:[{id:"179460",title:"Dr.",name:"Sahra",middleName:null,surname:"Kırmusaoğlu",slug:"sahra-kirmusaoglu",fullName:"Sahra Kırmusaoğlu"}]},{id:"74640",title:"Alternative Methods to Animal Use for Monoclonal Antibody Generation and Production",slug:"alternative-methods-to-animal-use-for-monoclonal-antibody-generation-and-production",totalDownloads:107,totalCrossrefCites:0,totalDimensionsCites:0,book:{slug:null,title:"Monoclonal Antibodies",fullTitle:"Monoclonal Antibodies"},signatures:"Jane Zveiter de Moraes, Barbara Hamaguchi, Camila Braggion, Enzo Speciale, Fernanda Cesar, Gabriela Soares, Juliana Osaki, Rodrigo Aguiar and Tauane Pereira",authors:null},{id:"74660",title:"Analytical Characterization of Monoclonal Antibodies with Novel Fc Receptor-Based Chromatography Technique",slug:"analytical-characterization-of-monoclonal-antibodies-with-novel-fc-receptor-based-chromatography-tec",totalDownloads:101,totalCrossrefCites:0,totalDimensionsCites:0,book:{slug:null,title:"Monoclonal Antibodies",fullTitle:"Monoclonal Antibodies"},signatures:"Atis Chakrabarti, Jukka Kervinen, Egbert Müller, Toru Tanaka and Kazuaki Muranaka",authors:null},{id:"50488",title:"Biosynthesis of Vitamins by Probiotic Bacteria",slug:"biosynthesis-of-vitamins-by-probiotic-bacteria",totalDownloads:4725,totalCrossrefCites:8,totalDimensionsCites:12,book:{slug:"probiotics-and-prebiotics-in-human-nutrition-and-health",title:"Probiotics and Prebiotics in Human Nutrition and Health",fullTitle:"Probiotics and Prebiotics in Human Nutrition and Health"},signatures:"Qing Gu and Ping Li",authors:[{id:"180415",title:"Prof.",name:"Qing",middleName:null,surname:"Gu",slug:"qing-gu",fullName:"Qing Gu"},{id:"180891",title:"Dr.",name:"Ping",middleName:null,surname:"Li",slug:"ping-li",fullName:"Ping Li"}]},{id:"74385",title:"Precision Medicine of Autoimmune Diseases",slug:"precision-medicine-of-autoimmune-diseases",totalDownloads:100,totalCrossrefCites:0,totalDimensionsCites:0,book:{slug:null,title:"Innate Immunity in Health and Disease",fullTitle:"Innate Immunity in Health and Disease"},signatures:"Ayodeji Ajayi, Oluwadunsin Adebayo and Emmanuel Adebayo",authors:null},{id:"55860",title:"Biotechnologies Applied in Biomedical Vaccines",slug:"biotechnologies-applied-in-biomedical-vaccines",totalDownloads:2055,totalCrossrefCites:2,totalDimensionsCites:3,book:{slug:"vaccines",title:"Vaccines",fullTitle:"Vaccines"},signatures:"Yuan‐Chuan Chen, Hwei‐Fang Cheng, Yi‐Chen Yang and Ming‐\nKung Yeh",authors:[{id:"180299",title:"Dr.",name:"Ming-Kung",middleName:null,surname:"Yeh",slug:"ming-kung-yeh",fullName:"Ming-Kung Yeh"},{id:"185559",title:"Dr.",name:"Yuan-Chuan",middleName:null,surname:"Chen",slug:"yuan-chuan-chen",fullName:"Yuan-Chuan Chen"},{id:"185560",title:"Dr.",name:"Hwei-Fang",middleName:null,surname:"Cheng",slug:"hwei-fang-cheng",fullName:"Hwei-Fang Cheng"},{id:"185561",title:"Dr.",name:"Yi-Chen",middleName:null,surname:"Yang",slug:"yi-chen-yang",fullName:"Yi-Chen Yang"}]}],onlineFirstChaptersFilter:{topicSlug:"immunology-and-microbiology",limit:3,offset:0},onlineFirstChaptersCollection:[{id:"74842",title:"Role of Dendritic Cells in Pathogen Infections: A Current Perspective",slug:"role-of-dendritic-cells-in-pathogen-infections-a-current-perspective",totalDownloads:7,totalDimensionsCites:0,doi:"10.5772/intechopen.95551",book:{title:"Cell Interaction - Regulation of Immune Responses, Disease Development and Management Strategies"},signatures:"José Luis Muñoz-Carrillo, Juan Francisco Contreras-Cordero, Oscar Gutiérrez-Coronado, Paola Trinidad Villalobos-Gutiérrez, Luis Guillermo Ramos-Gracia and Jazmín Monserrat Vargas-Barboza"},{id:"74788",title:"SARS-CoV-2 and Coronavirus Ancestors under a Molecular Scope",slug:"sars-cov-2-and-coronavirus-ancestors-under-a-molecular-scope",totalDownloads:6,totalDimensionsCites:0,doi:"10.5772/intechopen.95102",book:{title:"Cell Interaction - Regulation of Immune Responses, Disease Development and Management Strategies"},signatures:"Maram Adel Abdelghany, Sarah Abdullah Gozai Alghamdi and Jehane Ibrahim Eid"},{id:"74782",title:"Study of Various Chemically and Structurally Diverse Currently Clinically Used and Recently Developed Antimycobacterial Drugs",slug:"study-of-various-chemically-and-structurally-diverse-currently-clinically-used-and-recently-develope",totalDownloads:11,totalDimensionsCites:0,doi:"10.5772/intechopen.95538",book:{title:"Molecular Epidemiology Study of Mycobacterium Tuberculosis Complex"},signatures:"Saad Alghamdi and Mohammad Asif"}],onlineFirstChaptersTotal:29},preDownload:{success:null,errors:{}},aboutIntechopen:{},privacyPolicy:{},peerReviewing:{},howOpenAccessPublishingWithIntechopenWorks:{},sponsorshipBooks:{sponsorshipBooks:[{type:"book",id:"10176",title:"Microgrids and Local Energy Systems",subtitle:null,isOpenForSubmission:!0,hash:"c32b4a5351a88f263074b0d0ca813a9c",slug:null,bookSignature:"Prof. Nick Jenkins",coverURL:"https://cdn.intechopen.com/books/images_new/10176.jpg",editedByType:null,editors:[{id:"55219",title:"Prof.",name:"Nick",middleName:null,surname:"Jenkins",slug:"nick-jenkins",fullName:"Nick Jenkins"}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,productType:{id:"1",chapterContentType:"chapter"}}],offset:8,limit:8,total:1},route:{name:"profile.detail",path:"/profiles/172839/ralf-basner",hash:"",query:{},params:{id:"172839",slug:"ralf-basner"},fullPath:"/profiles/172839/ralf-basner",meta:{},from:{name:null,path:"/",hash:"",query:{},params:{},fullPath:"/",meta:{}}}},function(){var e;(e=document.currentScript||document.scripts[document.scripts.length-1]).parentNode.removeChild(e)}()