Open access

Lentiviral Vectors in Immunotherapy

Written By

Ines Dufait, Therese Liechtenstein, Alessio Lanna, Roberta Laranga, Antonella Padella, Christopher Bricogne, Frederick Arce, Grazyna Kochan, Karine Breckpot and David Escors

Submitted: 02 June 2012 Published: 27 February 2013

DOI: 10.5772/50717

From the Edited Volume

Gene Therapy - Tools and Potential Applications

Edited by Francisco Martin Molina

Chapter metrics overview

2,473 Chapter Downloads

View Full Metrics

1. Introduction

Genetic immunotherapy can be defined as a therapeutic approach in which therapeutic genes are introduced into defined target cell types to modulate immune responses. A major challenge for this therapeutic strategy is the delivery of these genes into target cells in an efficient, stable manner. Possibly one of the best systems to achieve this is the use of lentiviral vectors (lentivectors) as gene carriers, as they are capable of transducing both dividing and resting cells [1].

Lentivectors are mainly derived from the human immunodeficiency virus (HIV-1) genome, a member of the Retroviridae family. The defining characteristic of retroviruses is their capacity to stably integrate their RNA genome into the host cell chromosomes, in the form of a cDNA copy (Figure 1). Therefore, retrovirus and lentivirus vectors have been used extensively in research since they are ideal gene carriers into target cells. Moreover, both retrovirus and lentivirus vectors have been successfully applied in human gene therapy for the treatment of several genetic/metabolic inherited diseases (Cartier et al, 2009; Cavazzana-Calvo et al, 2010; Gaspar et al, 2004; Grez et al, 2010; Ott et al, 2006; Thrasher et al, 2006).

Lentiviruses are spherical enveloped viruses with a diameter around 80 to 120 nm and contain two copies of a single-stranded RNA genome (Figure 2) [2]. The genome is enclosed within a core composed of the structural and enzymatic proteins nucleocapsid (NC), capsid (CA), reverse transcriptase (RT), integrase (IN) and protease (PR). The core is surrounded by a protein layer of matrix (MA) protein. The envelope protein (ENV) is embedded in the virion lipid envelope, and it binds to the target cellular receptor and mediates virion entry.

Figure 1.

The retrovirus life cycle.

The life cycle of retroviruses, including lentiviruses is shown in this figure as a multistep mechanism, starting with virion binding to the cellular receptor (R), leading to direct fusion or endocytosis. Then, the internal core is released and the two RNA molecules undergo reverse transcription as indicated, ending up with a single cDNA molecule. The core is then transported to the nucleus (in the case of lentiviruses) and the cDNA is integrated into the cell chromosome. The integrated genome (provirus) undergoes transcription, producing more RNA genome copies (and also spliced mRNAs, not shown here), which are also translated into structural and enzymatic proteins. These are then assembled into virions that bud out of the infected cells.

Figure 2.

In this scheme, the lentivirus virion is represented as a sphere containing a genome made of two RNA molecules associated to the nucleocapsid (NC) protein. The nucleocapsid is enclosed by a core made of capsid (CA) protein, which is surrounded by a shell of matrix (MA) protein that associates to the virion envelope. The two subunits of the HIV-1 envelope are also indicated (SU and TM). In addition, other enzymatic (IN, RT, PR), accessory (Vpr, p6) and cellular (Cyclophilin A, Cy-A) proteins are shown, which are incorporated into lentivirus particles.

Lentivectors are classified as complex retroviruses according to their genome organisation, as it contains accessory and regulatory genes absent in other retroviruses. Nevertheless, the retrovirus genome shares a common 5’ to the 3’ gene organisation, with Gag, Pol and Env genes [1, 3]. Gag encodes MA, CA and NC as a polyprotein. Pol encodes enzymatic proteins associated to reverse transcription, that is, the reverse transcriptase (RT), integrase (IN) and protease (PR). RT synthesizes a single cDNA copy from the retrovirus genomic RNA [4]. IN mediates cDNA integration in the host cell chromosome, while PR cleaves Gag and Gag-Pol polyproteins during virion maturation.

The integrated cDNA genome is flanked by two long terminal repeats (LTRs) subdivided in U3, R and U5 regions. U3 is the HIV-1 promoter. The R region marks the starting point of transcription, and U5 region is critical for reverse transcription. The other key elements are the packaging signal (Ψ) and the polypurine tract (PPT). The packaging signal, as in many other virus species, allows RNA genome encapsidation during virion assembly in the cytoplasm. The PPT element is a key element for reverse transcription [5].

Lentivectors are usually obtained following a three-plasmid co-transfection in 293T cells (Figure 3) [6, 7]. The first one, the packaging plasmid, provides the structural and RT proteins (Gag-Pol). The second one, the envelope plasmid, encodes a glycoprotein to pseudotype the lentivector particles. This process consists on the incorporation of an heterologous Env in the viral lipid envelope. This will allow the lentivector to exhibit the specific cell tropism given by the Env used in pseudotyping. One of the most used Env is the Vesicular Stomatitis Virus (VSV) Glycoprotein (G). The VSV-G confers stability to the lentivector particles and a very broad tropism for human and non-human cells [8]. Lastly, the third one, the transfer plasmid, contains the cis-acting sequences for replication/transcription and packaging (Figure 3) [9]. By including promoters within the transfer plasmid, any gene of interest can be expressed either constitutively or inducibly, in a cell type-specific of unspecific manner (Figure 3) [1]. Therefore, lentiviral vectors can also incorporate genes with immunoregulatory properties in cells from the immune system.

Two main cell types of the immune system have been preferential targets for genetic immunotherapy: antigen presenting cells (APCs) and effector T lymphocytes. These two cell types are key controllers of immune responses. By expressing transgenes of interest in APCs, such as DCs, they can be processed and presented to antigen-specific T cells in the immunological synapse. This antigen presentation is the first step in either starting of suppressing immune responses. Therefore, if genes with modulating properties of APC functions can be co-expressed with antigens, the strength and type of immune response can be controlled. In fact, genetic modification of cells from the immune system can circumvent the limitations of current immunotherapeutic protocols. Using targeted lentiviral vectors, specificity and effectiveness can be achieved by targeting key cells that modulate and polarize immune responses.

Although more challenging than DCs, T lymphocytes can also be genetically modified using lentiviral vectors. Vectors expressing T cell receptors (TCRs) specific for antigens of interest can modify the specificity of T cell populations, or expand their antigen profile. Therefore, these genetically modified T cells can be adoptively transferred in the human patients. This strategy is particularly important to generate T cells with high affinity TCRs towards tumour-associated antigens.

Figure 3.

The HIV-1 genome is shown at the top of the figure. All structural, accessory and enzymatic genes are indicated throughout the genome. The two LTRs are shown as present in an integrated provirus. The three functional regions of the HIV LTR are shown on top of the 5- LTR. Numbers indicate nucleotide positions. The HIV genome is splitted in three different plasmids to engineer a gene vector. The transfer (vector) plasmid is indicated, with only the HIV-1 LTRs containing an internal promoter driving the expression of a gene of interest. In the packaging plasmid, only the Gag-Pro-Pol and Rev, Tat genes have been retained. This increases biosafety. Transcription of these genes takes place under the control of the cytomegalovirus promoter (CMV), as indicated in the figure. Lastly, a third plasmid encoding an envelope glycoprotein is shown on the bottom of the figure. This envelope pseudotypes the lentivector particle.

Advertisement

2. Genetic modification of DCs with lentivectors

For the elimination of cancer cells and chronic infections such as HIV, hepatitits B and malaria, a strong, effective T cell response is required. To initiate these strong T cell responses, the interaction between antigen-specific T cells and antigen-presenting APCs has to be strengthened[10]. Amongst APCs, DCs are most frequently the targets of immunotherapy protocols since they are probably the most immunogenic [10, 11].

To activate T cells during antigen presentation, these T cells have to receive at least three different signals from APCs (Figure 4) [10, 12-16]. The first signal, or signal 1, is the direct recognition of the peptide-major histocompatibility molecule complex (p-MHC) by the TCR. However, this interaction is not sufficient to confer T cells with effector activities. For this, a second co-stimulatory signal (signal 2) has to be co-delivered together with p-MHC recognition. This signal 2 is the consequence of the integration of activatory and inhibitory interactions between ligands/receptors on the surface of DCs and T cells (Figure 4)[16, 17]. For example, CD80 binding to CD28 is strongly activatory, while CD80 binding with CTLA-4, or PD-L1 with PD-1, are strongly inhibitory[16, 17]. Apart from these two signals, T cells require a third signal which drives their differentiation into distinct subtypes that will regulate different types of immune responses [13, 18]. This third signal is usually provided by different combination of cytokines present within the immunological synapse (Figure 4). For example, the presence of high levels of IL12 will polarise T cell differentation towards a Th1 type (cellular cytotoxic immunity). On the other hand, high levels of IL10 will drive polarisation towards Th2 (antibody responses).

Figure 4.

In this scheme, DCs (left) present antigens to specific CD8 and CD4 T cells (as indicated) in the context of MHC class I (I) or class II (II) molecules. These T cells receive further stimuli by co-stimulation through ligand-receptor interactions between the DC and T cells (as indicated in the figure). Simultaneously, activated DCs secrete cytokines and chemokines (indicated in the figure as spheres) that will drive T cell activation, proliferation and differentation into either cytotoxic CD8 T cells or T helper cells, as shown on the right.

An ideal immunotherapeutic approach would be to use lentiviral vectors to deliver the antigen of interest together with the three signals required for the desired T cell polarisation. Lentivectors have been extensively used for this purpose, because they are particularly effective in transducing DCs without affecting their functionality, unlike other vectors such as those based on adenoviruses [8, 19-26]. In fact, the stable integration of the lentivector genome allows long-term, sustained transgene expression[1, 9]. In addition, the expressed transgene is processed and its antigen peptides loaded in MHC I and MHC II molecules [27]. This is of the outmost importance for immunotherapy, since expressed proteins can be processed and loaded onto MHC-II molecules through several pathways. While secreted proteins can enter the endocytic pathway and membrane proteins can be recycled towards the endosomal pathway, cytoplasmic proteins can still enter the MHC II pathway by autophagy[28]. Nevertheless, to improve MHC II loading of peptides from cytoplasmic proteins, endocytic localisation sequences can be fused to the transgene, with the lysosomal-associated membrane protein 1 or with the amino-terminal portion of the MHC II invariant chain [29-32].

The use of lentivectors to express whole trangenes rather than antigen petides circumvents the necessity of designing specific peptide/protein vaccines for loading into specific MHC genotypes [27]. Thus, lentivectors expressing model antigens have been extensively used as a proof of principle. Amongst others, the antigens chicken ovalbumin (OVA), tumour-tumour associated antigens such as MELAN-A, tyrosinase related protein (Trp), NY-ESO or antigens from infectious agents have been expressed in DCs. These modified DCs induced strong activation and proliferation of antigen-specific T cells. [17, 33-38].

Advertisement

3. Lentivector immunogenicity

Lentivectors have been extensively used in vaccination protocols, due to their capacity of raising strong transgene-specific immune responses [9, 17, 30, 38-40]. Interestingly, some reports suggest that lentivectors are incapable of inducing DC maturation in vitro, suggesting that some components of the lentivector preparations provide signals 2 and 3 through a mechanisms not well understood [40, 41].

DC maturation is a complex, step-wise process in which they up-regulate the surface expression of co-stimulatory molecules such as CD80, CD83, CD86, CD40, adhesion molecules such as ICAM-1 and also the expression of MHC molecules. In general terms, DC maturation can be triggered by recognition of pathogen-derived molecules by specific receptors on the DC surface, such as the family of toll-like receptors (TLRs) [42, 43]. DCs can also mature through the exposure of pro-inflammatory cytokines by a process called cytokine priming [13-15]. Matured DCs can effectively provide strong signals 1 and 2, leading to efficient T cell activation and proliferation. Thus, their administration in vivo induces DC maturation and production of type I interferon that can provide signal 3 [9, 39, 44, 45].

The capacity of lentivectors to induce DC maturation after vaccination is probably caused by either specific components of the lentivector particle or by contaminants present in the lentivector preparation. As a matter of fact, lentivector particles resemble viruses and therefore, some components have the potential to stimulate immune responses such as the RNA genome or the cDNA [46]. These are ligands for TLR7 and TLR9, respectively [41, 45]. In addition to specific components of the lentivector particle, contaminants can also alter their immunostimulatory properties. In fact, most lentivector preparations pseudotyped with VSV G contain VSV-G tubulo-vesicular structures enclosing plasmid DNA that stimulate TLR9 in vitro, leading to type I IFN production by pDCs [47]. In addition, foetal calf serum (FCS) contributes to immunogenicity by providing T cell epitopes with adjuvant capacities [48].

Advertisement

4. Control of DC maturation by expression of molecular activators with lentivectors

Lentivector preparations can induce DC maturation in vivo. However, in some circumstances this is not enough for effective therapeutic activities. This is the case for cancer immunotherapy, in which breaking tolerance towards TAAs is still a medical challenge. One possible solution is to co-express TAAs with molecular activators of DCs using lentivectors, particularly using activators of signalling cascades belonging to the TLR pathways.

This has been firstly achieved by over-expressing adaptor molecules, which associate with TLR cytoplasmic tails. These adaptor molecules recruit activatory protein kinases leading to DC maturation. Thus, lentivectors have been used to express MYD88 or TRIF1 in mouse myeloid DCs, which also increases secretion of pro-inflammatory cytokines IL-6, IL-12 and IFN-α, which enhanced T cell cytotoxicity [49]. The NF-κB pathway has also been an attractive target because it controls transcription of the majority of pro-inflammatory genes (Figure 5). Lentivectors have thus been used to express the Kaposi’s sarcoma-associated herpes virus FLICE-like inhibitory protein (vFLIP), a constitutive activator of NF-κB by direct association and activation of NF-κB essential modulator (NEMO) [50-53]. In fact, vFLIP expression has resulted to be a strong adjuvant when expressed in DCs, leading to strong DC maturation and effective CD4 and CD8 T cell responses. Lentivector expression of vFLIP significantly improves anti-tumour activities in a lymphoma mouse model and anti-parasitic efficacy in an OVA-expressing leishmania model [38, 54]. Lentivectors have also been used to inhibit negative regulators of NF-κB activation, such as the ubiqutin ligase A20. Lentivectors have successfully delivered to DCs short hairpin RNAs (shRNAs) targeting A20. The abrogation of A20 expression caused DC maturation, effective CD8 cell responses and inihibition of regulatory T cells (Tregs) [55, 56].

Other molecular activators of mitogen activated protein kinases (MAPKs), activated after TLR engagement, have also been co-expressed in DCs with antigens of interest (Figure 5). MAPKs are mainly divided in three groups, ERK, p38 and JNK. While ERK is associated to survival and immune suppression, p38 and JNK are thought to stimulate DC maturation and inflammation (Figure 5). Constitutive p38 activation was achieved by expressing the MKK6 EE mutant using lentivectors, and it resulted in CD80, CD40 and ICAM-I up-regulation without significant secretion of pro-inflammatory cytokines [17, 40, 41]. A similar result was achieved by JNK1 activation, following expression of the MKK7-JNK1 fusion gene in DCs. Interestingly, although a full DC maturation phenotype was not achieved ex vivo, co-expression of these MAPK activators with an OVA-containing transgene or MELAN-A induced significant antigen-specific CD4 and CD8 T cell responses. Moreover, these lentivectors improved survival in a murine tumour model for lymphoma, both with integrating and non-integrating lentivectors [38].

Figure 5.

Intracellular signalling pathways regulating DC functions.

Other molecules have been applied for DC maturation. For example, CD40 ligand expression achieved human DC activation and up-regulated the expression of CD83, CD80, MHC-I and induced IL-12 secretion [57]. This strategy increased CD4 and CD8 responses towards influenza epitopes and the TAA gp100. Co-expression of DC-promoting cytokines such as GM-CSF- and IL-4 using lentivectors resulted in long-lasting immunity against melanoma when co-expressed with TAAs Trp-2 and Mart-1 [58].

In this scheme, the main signalling pathways triggered after the engagement of a wide range of receptors on the DC surface (see the indicated receptors embedded in the membrane) with their ligands. Engagement of these receptors starts a complicated cascade of signalling pathways that will converge in a few, well-characterised ones, the NK-κB, MAPKs and interferon regulatory factors (IRFs) (as indicated below the membrane). Some of these pathways, such as NK-κB, MAPKs p38 and JNK1 are pro-inflammatory and lead to DC maturation. Others, such as ERK, are clearly immunosuppressive.

Advertisement

5. Control of DC maturation by inhibiting negative co-stimulation using lentivectors

DC maturation can also up-regulate molecules that provide negative stimulation to T cells, such as programmed cell death receptor ligand 1 (PD-L1) and PD-L2, the ligands for the PD-1 receptor on T cells. Negative co-stimulation is part of a regulatory mechanism that controls the activation state of T cells following antigen presentation [17, 59, 60]. Thus, interference with negative co-stimulation could in principle reinforce T cell activation and enhance cytotoxic activities. Therefore, lentivectors have been used to deliver shRNAs in DC against PD-L1. PD-L1 silencing in antigen-presenting DCs hyperactivated T cells by preventing the up-regulation of Casitas B-lymphoma (Cbl)-b E3 ubiquitin ligase. This strategy co- accelerated anti-tumour immune responses, particularly if combined with a p38 activator or dominant negative mutant of MEK1, the upstream kinase of ERK [17, 59].

Advertisement

6. Lentivectors and cancer immunotherapy

Lentivectors are particularly promising in cancer immunotherapy, for which conventional immunization is largely ineffective due to two major barriers. Firstly, TAAs are generally self-proteins to which there is strong immunological tolerance. Secondly, that tumours are strongly immune-suppressive and they use several mechanisms to avoid immune responses [41].

Lentivectors can be used in cancer immunotherapy in two different ways. In the first one, DCs can be generated ex vivo from the patient, followed by lentivector transduction and in vivo administration. Thus, cellular vaccination with transduced DCs expressing HLA-Cw3 induced activation and proliferation of CD8 T cells in a mouse model [37]. Similarly, lentivector transduction was shown to be superior to peptide pulsing in inducing OVA-specific T cell responses [61], protected mice from OVA-expressing tumour cells and significantly inhibited tumour growth. The second strategy is direct lentivector vaccination, taking advantage of their intrinsic immuno-stimulatory capacities and their reduced cost [24, 26, 35].

Figure 6.

A scheme of the TCR is shown embedded in the cellular membrane. Both α and β chains are shown as indicated, subdivided in variable and constant regions (V, C). The other CD3 chains that associate with the TCR are also included in the figure. On the bottom, a lentivector co-expressing α and β TCR chains is shown, under the control of the spleen focus forming virus promoter (SFFV) and an internal ribosome entry site (IRES) [67]. This particular lentivector is self-inactivating (SIN) and presents a deletion of viral enhancers in the 3’ LTR. When this construct is integrated, the 5’ LTR disappears and it is replaced with the deleted version.

As mentioned above, a major issue with cancer immunotherapy is that most TAA-specific T cells may have been eliminated during thymic clonal deletion. Thus, even if effective and strong DC maturation is achieved, no effective responses will be achieved due to lack of TAA-specific T cells. To circumvent this, TAA-specific T cells can be generated by lentivector transduction in vitro, and adoptively transfered in patients (Figure 6) [62]. Clinical efficacy has been reported for melanoma, synovial cell sarcoma, colorectal, neuroblastoma and lymphoma, but using γ-retrovirus vectors instead of lentivectors [63-66].

T cells are largely refractory to transduction by VSV G-pseudotyped lentivectors, and they require some level of T cell stimulation [67]. Treatment with IL-2 and IL-7 allows lentivector transduction and preserves a functional T cell repertoire [68, 69]. As an example, Wilms tumour antigen (WT1)-specific T cells were generated by lentivector expression of a WT1-specific TCR in the presence of IL-15 and IL-21. These modified T cells were multifunctional and exhibited the expected antigen specificity [67]. This approach of T cell modification is rather promising. In a clinical trial with 15 terminally sick melanoma patients, 2 showed complete regression and long-term survival after transfer of T cells expressing a MART-1-specific TCR using γ-retrovirus vectors [70]. Interestingly, it has been recently demonstrated that entivectors pseudotyped with measles virus H/F glycoproteins effectively transduce quiescent adult T cells in the absence of any exogenous stimulus, whether cytokines or anti-CD3/anti-CD28 stimulation. In fact, transduction with these lentivectors did not affect T cells in any way [17, 71-73].

Advertisement

7. Lentivector gene immunotherapy for the treatment of autoimmune disorders

It is relatively “straightforward” to achieve immune stimulation using lentivectors. However, the induction of immune suppression or tolerance with lentivectors is rather challenging. Nevertheless, the induction and maintenance of immunological tolerance is critical for homeostasis. The organism is permanently and closely in contact with a very wide range of antigens of many origins. A large majority of them are innocuous and do not pose a direct threat. Thus, the immune system must not respond to these antigens, as an immune response is associated with significant collateral tissue damage. The immune system should be activated only if a real threat appears. Therefore, the immune system possesses several tolerogenic mechanisms in place to keep immunological homeostasis. As mentioned before, a key one is clonal deletion of auto-reactive T cells in the thymus [74]. However, there is a significant number of auto-reactive T cells that escape from clonal deletion. Many of them will differentiate towards natural Foxp3 CD4 regulatory T cells [74-77].

In addition to clonal deletion and differentiation of natural Tregs, there are a number of tolerogenic mechanisms in place that regulate immune responses towards peripheral antigens. The organism is in permanent direct contact with many substances and commensal organisms in mucosal areas and in peripheral tissues. In these situations, inducible Tregs differentiate from naïve CD4 T cells after antigen presentation by tolerogenic DCs. These regulatory T cell types are usually classified in Tr1 (CD4 CD25 IL10 or TGF-β) and Th3 (CD4 CD25 Foxp3) cells [78-82]. Therefore, DCs can also be converted in tolerogenic by expression of immunomodulatory genes with lentivectors. This strategy opens up the application of lentivectors for the treatment of autoimmune disorders.

Advertisement

8. Induction of tolerogenic DCs using lentivectors

It is relatively “straightforward” to achieve immune stimulation using lentivectors. However, the induction of immune suppression or tolerance with lentivectors is rather challenging. However, the induction and maintenance of immunological tolerance is critical for homeostasis. The organism is permanently and closely in contact with a very wide range of antigens of many origins. A large majority of them are innocuous and do not pose a direct threat. Thus, the immune system must not respond to these antigens, as an immune response is associated with significant collateral tissue damage. The immune system should be activated only if a real threat appears. Therefore, the immune system possesses several tolerogenic mechanisms in place to keep immunological homeostasis. As mentioned before, a key one is clonal deletion of auto-reactive T cells in the thymus [74]. However, there is a significant number of auto-reactive T cells that escape from clonal deletion. Many of them will differentiate towards natural Foxp3 CD4 regulatory T cells [74-77].

In addition to clonal deletion and differentiation of natural Tregs, there are a number of tolerogenic mechanisms in place that regulate immune responses towards peripheral antigens. The organism is in permanent direct contact with many substances and commensal organisms in mucosal areas and in peripheral tissues. In these situations, inducible Tregs differentiate from naïve CD4 T cells after antigen presentation by tolerogenic DCs. These regulatory T cell types are usually classified in Tr1 (CD4 CD25 IL10 or TGF-β) and Th3 (CD4 CD25 Foxp3) cells [78-82]. Therefore, DCs can also be converted in tolerogenic by expression of immunomodulatory genes with lentivectors. This strategy opens up the application of lentivectors for the treatment of autoimmune disorders

Advertisement

9. Induction of tolerogenic DCs using lentivectors

DCs can induce immunological tolerance through a number of mechanisms. It is generally accepted that antigen presentation by immature DCs is poorly immunogenic, and results in Treg differentiation, T cell apoptosis and T cell anergy [83-86]. These immature tolerogenic DCs express low levels of co-stimulatory molecules CD80, CD86, CD83, CD40 and MHC molecules [10, 40, 41, 78, 87]. Resident mucosal DCs are intrinsically tolerogenic independently on their maturation phenotype as a consequence of the presence of retinoic-acid [88]. In addition, these DCs become strongly immunosuppressive due to contact with TLR agonists from commensal microbiota [88-90]. DCs can also become strongly immunosuppressive after treatment with lectin ligands or exposure to immunosuppressive cytokines such as IL-10, IL-4 or TGF-β[78, 87, 89-92]. Tolerogenic DCs usually express high levels of these immunosuppressive cytokines, even if they are phenotypically mature [10, 40, 78, 87, 89-93]. In this situation, they provide strong signals 1 and 2 to T cells, together with a simultaneous strong tolerogenic signal 3. For example, in the presence of bioactive TGF-β, strong antigen presentation leads to differentiation to antigen-specific Foxp3 Tregs, while secretion of IL-10 usually results in Tr1 differentiation [91, 93-95].

Tolerogenic DCs can also up-regulate molecules that provide an inhibitory signal to T cells, such as PD-L1 (or B7-H1), a member of the B7 co-stimulatory molecules [17, 96]. PD-L1 expression in DCs regulates T cell activities during antigen presentation and prevents T cell hyperactivation [17]. In addition, PD-L1-CD80 binding on T cells induces antigen-specific Treg differentiation [97]. Other members of the B7 family are immunosuppressive [98]. Immunosuppressive DCs also up-regulate aminoacid-metabolising enzymes, such as arginase or indoleamine 2,3-dioxygenase (IDO) [99-104]. It is thought that these enzymes deplete T cells of essential aminoacids.

Lentivectors can be used to confer tolerogenic activities to DCs by expression of immunoregulatory genes together with antigens of interest. The first strategy that was tested experimental was the expression of potent immunosuppressive cytokines. This approach was used with γ-retroviral vectors for inflammatory diseases [95, 105, 106], 105, 106]. Lentivectors have been applied in an experimental model of asthma by expressing IL-10, leading to expansion of IL-10-expressing Foxp3 Tregs with potent anti-inflammatory properties [107]. Alternatively, small immunosuppressive peptides can also be delivered with lentivectors, such as the vasointestinal peptide (VIP). Intraperitoneal administration of VIP-encoding lentivectors in mice effectively inhibited the development of experimental collagen-induced arthritis. This was achieved by a markedly reduction of pro-inflammatory cytokine secretion and the expansion of Foxp3 Tregs [108]. The administration of genetically modified VIP-expressing DCs also showed significant therapeutic effects in EAE and in the coecal ligation and puncture (CLP) model [109].

DCs can also be reprogrammed by direct modulation tolerogenic signalling pathways within DCs (Figure 5). Therefore, lentivector expression of a constitutively active MEK1 mutant resulted in sustained MAPK ERK phosphorylation, resulting in immunological tolerance [40, 90, 110-114]. These genetically modified DCs exhibit an immature phenotype with low levels of CD40 and secretion of bioactive TGF-β[40, 78]. Antigen presentation by these ERK-activated DCs differentiated antigen-specific Foxp3 Tregs both ex vivo and in vivo in a mouse model [78]. Direct lentivector vaccination encoding the ERK activator effectively controlled antigen-induced inflammatory arthritis in a mouse model [78].

Similarly, lentivector expression of a constitutively active IRF3 mutant induced high expression levels of IL-10, and expanded antigen-specific Foxp3 Tregs which inhibited immune responses (Figure 5) [40]. Activation of endogenous negative feedback mechanisms of DC maturation pathways has also been applied to induce immune suppression. In this way, by over-expressing the suppressor of cytokine signalling 3 (SOCS-3) in DCs, pro-inflammatory signalling pathways were severly impaired [115]. These genetically modified DCs significantly decreased secretion of pro-inflammatory cytokines IFN-γ, IL-12 and IL-23, and showed an enhanced IL-10 production, which effectively inhibited effectively inhibit experimental autoimmune encephalomyelitis (EAE) in mice [115].

An alternative strategy to generate tolerogenic DCs is the inhibition of pro-inflammatory signalling pathways instead of activating immunosuppressive pathways. As NF-κB is a critical inflammatory signalling pathway, its inhibition in promising for the induction of immunological tolerance [41]. To achieve this, Rel-B was silenced by the delivery of a shRNA targeted to Rel-B [116]. In this way, its inhibition could effectively prevent DC maturation after engagement with TLR ligands, and it was sufficient to treat autoimmune myasthenia gravis in a mouse model [116]. In an analogous manner, lentivectors have also been applied to silence B cell activating factor (BAFF) in the inflamed joint [117, 118], which was very effective for the treatment of experimental collagen-induced arthritis [119] without the need of targeting the arthritogenic antigen. These lentivectors were directly injected in the inflamed joint, where they preferentially transduced resident DCs. BAFF silencing in these DCs inhibited their maturation, and most importantly, inhibited differentiation of pathogenic Th17 [119].

Advertisement

10. Conclussions

Classical immunotherapeutic strategies for the treatment of cancer and infectious diseases rely on either administration of the antigen peptides together with adjuvants, or the inoculation with attenuated strains of pathogenic agents. This approach has been largely successful for the treatment of a wide range of infectious agents. However, for cancer immunotherapy, most potent and targeted immunotherapeutic approaches are required to break the natural tolerance towards TAAs. The targeted co-delivery of immunomodulatory genes with antigens of interests to DCs has opened the application of gene therapy for immunotherapy. Lentivectors exhibit a remarkable transduction capacity of DCs and also T cells, and thus, they are ideal tools to achieve immunodulation. In this way, the immune system can be strongly and specifically activated for the treatment of cancer and infectious diseases, but it can on the other hand be strongly immunosuppressed. This makes it possible the induction of immunological tolerance and treatment of autoimmune disorders.

Acknowledgments

Ines Dufait is funded by an ERASMUS scholarship. Alessio Lanna is funded by an University College London “bench-to-bedside” PhD scholarship. Christopher Bricogne is funded by an University College London MB-PhD scholarship. The Structural Genomics Consortium Oxfod is a registered UK charity (number 1097737) that receives funds from the Canadian Institutes of Health Research, The Canadian Foundation for Innovation, Genome Canada through the Ontario Genomics Insitute, GlaxoSmithKline, Karolinska Institutet, the Knut and Alice Wallenberg Foundations, the Ontario Innovation Trust, the Ontario Ministry for Research and Innovation, Merck & Co., Inc., the Novartis Research Foundation, the Swedish Foundation for Strategic Research and the Wellcome Trust. Karine Breckpot is funded by the Fund for Scientific Research- Flandes. David Escors is funded by an Arthritis Research UK Career Development Fellowship (18433).

References

  1. 1. Escors D. Breckpot K. 2010 Lentiviral Vectors in Gene Therapy: Their Current Status and Future Potential Arch Immunol Ther Exp 58 2 107 119
  2. 2. Vogt V. M. Simon M. N. 1999 Mass determination of rous sarcoma virus virions by scanning transmission electron microscopy J Virol 73 8 7050 7055
  3. 3. Katz R. A. Skalka A. M. 1994 The retroviral enzymes Annual review of biochemistry 63 133 173
  4. 4. Herschhorn A. Hizi A. 2010 Retroviral reverse transcriptases Cell Mol Life Sci 67 16 2717 2747
  5. 5. Charneau P. Alizon M. Clavel F. 1992 A second origin of DNA plus-strand synthesis is required for optimal human immunodeficiency virus replication J Virol 66 5 2814 20
  6. 6. Naldini L. Blomer U. Gage F. H. Trono D. Verma I. M. 1996 Efficient transfer, integration, and sustained long-term expression of the transgene in adult rat brains injected with a lentiviral vector Proc Natl Acad Sci U S A 93 21 11382 11388
  7. 7. Naldini L. Blomer U. Gallay P. Ory D. Mulligan R. Gage F. H. Verma I. M. Trono D. 1996 In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector Science 272 5259 263 267
  8. 8. Yee J. K. Friedmann T. Burns J. C. 1994 Generation of high-titer pseudotyped retroviral vectors with very broad host range Methods in cell biology 43 Pt A 99 112
  9. 9. Breckpot K. Escors D. Arce F. Lopes L. Karwacz K. Van Lint S. Keyaerts M. Collins M. 2010 HIV-1 lentiviral vector immunogenicity is mediated by Toll-like receptor 3 (TLR3) and TLR7 J. Virol. 84 5627 5636
  10. 10. Liechtenstein T. Dufait I. Lanna A. Breckpot K. Escors D. 2012 Modulating co-stimulation during antigen presentation to enhance cancer immunotherapy Immun., Endoc. & Metab. Agents in Med. Chem. 12 00 00
  11. 11. Steinman R. M. Banchereau J. 2007 Taking dendritic cells into medicine Nature 449 7161 419 26
  12. 12. Janeway C. A. Jr. Bottomly K. 1994 Signals and signs for lymphocyte responses Cell 76 2 275 85
  13. 13. Curtsinger J. M. Johnson C. M. Mescher M. F. 2003 CD8 T cell clonal expansion and development of effector function require prolonged exposure to antigen, costimulation, and signal 3 cytokine Journal of immunology 171 10 5165 71
  14. 14. Curtsinger J. M. Lins D. C. Mescher M. F. 2003 Signal 3 determines tolerance versus full activation of naive CD8 T cells: dissociating proliferation and development of effector function The Journal of experimental medicine 197 9 1141 51
  15. 15. Curtsinger J. Lins D. Mescher M. 2003 Signal 3 determines tolerance versus full activation of naive CD8 T cells: dissociating proliferation and development of effector function Journal of experimental medicine 197 1141 1151
  16. 16. Nurieva R. Thomas S. Nguyen T. Martin-Orozco N. Wang Y. Kaja M. K. Yu X. Z. Dong C. 2006 T-cell tolerance or function is determined by combinatorial costimulatory signals Embo J. 25 11 2623 2633
  17. 17. Karwacz K. Bricogne C. Macdonald D. Arce F. Bennett C. L. Collins M. Escors D. 2011 PD-L1 co-stimulation contributes to ligand-induced T cell receptor down-modulation on CD8(+) T cells EMBO molecular medicine 3 10 581 92
  18. 18. Curtsinger J. M. Schmidt C. S. Mondino A. Lins D. C. Kedl R. M. Jenkins M. K. Mescher M. F. 1999 Inflammatory cytokines provide a third signal for activation of naive CD4+ and CD8+ T cells Journal of immunology 162 6 3256 62
  19. 19. Copreni E. Castellani S. Palmieri L. Penzo M. Conese M. 2008 Involvement of glycosaminoglycans in vesicular stomatitis virus G glycoprotein pseudotyped lentiviral vector-mediated gene transfer into airway epithelial cells J Gene Med 10 12 1294 1302
  20. 20. Burns J. C. Matsubara T. Lozinski G. Yee J. K. Friedmann T. Washabaugh C. H. Tsonis P. A. 1994 Pantropic retroviral vector-mediated gene transfer, integration, and expression in cultured newt limb cells Developmental biology 165 1 285 289
  21. 21. Bouard D. Alazard-Dany D. Cosset F. L. 2009 Viral vectors: from virology to transgene expression British journal of pharmacology 157 2 153 165
  22. 22. Strang B. L. Ikeda Y. Cosset F. L. Collins M. K. Takeuchi Y. 2004 Characterization of HIV-1 vectors with gammaretrovirus envelope glycoproteins produced from stable packaging cells Gene Ther 11 7 591 598
  23. 23. Miller A. D. Garcia J. V. von Suhr. N. Lynch C. M. Wilson C. Eiden M. V. 1991 Construction and properties of retrovirus packaging cells based on gibbon ape leukemia virus J Virol 65 5 2220 2224
  24. 24. Vanden Driessche. T. Thorrez L. Naldini L. Follenzi A. Moons L. Berneman Z. Collen D. Chuah M. K. 2002 Lentiviral vectors containing the human immunodeficiency virus type-1 central polypurine tract can efficiently transduce nondividing hepatocytes and antigen-presenting cells in vivo Blood 100 3 813 22
  25. 25. Faix P. H. Feldman S. A. Overbaugh J. Eiden M. V. 2002 Host range and receptor binding properties of vectors bearing feline leukemia virus subgroup B envelopes can be modulated by envelope sequences outside of the receptor binding domain J Virol 76 23 12369 75
  26. 26. Esslinger C. Chapatte L. Finke D. Miconnet I. Guillaume P. Levy F. Mac Donald. H. R. 2003 In vivo administration of a lentiviral vaccine targets DCs and induces efficient CD8(+) T cell responses J Clin Invest 111 11 1673 1681
  27. 27. Collins M. K. Cerundolo V. 2004 Gene therapy meets vaccine development Trends Biotechnol 22 12 623 626
  28. 28. Paludan C. Schmid D. Landthaler M. Vockerodt M. Kube D. Tuschl T. Munz C. 2005 Endogenous MHC class II processing of a viral nuclear antigen after autophagy Science 307 5709 593 596
  29. 29. Gregers T. F. Fleckenstein B. Vartdal F. Roepstorff P. Bakke O. Sandlie I. 2003 MHC class II loading of high or low affinity peptides directed by Ii/peide fusion constructs: implications for T cell activation International immunology 15 11 1291 1299
  30. 30. Rowe H. M. Lopes L. Ikeda Y. Bailey R. Barde I. Zenke M. Chain B. M. Collins M. K. 2006 Immunization with a lentiviral vector stimulates both CD4 and CD8 T cell responses to an ovalbumin transgene Mol Ther 13 2 310 9
  31. 31. Sanderson S. Frauwirth K. Shastri N. 1995 Expression of endogenous peptide-major histocompatibility complex class II complexes derived from invariant chain-antigen fusion proteins Proceedings of the National Academy of Sciences of the United States of America 92 16 7217 7221
  32. 32. Wu T. C. Guarnieri F. G. Staveley-O’Carroll K. F. Viscidi R. P. Levitsky H. I. Hedrick L. Cho K. R. August J. T. Pardoll D. M. 1995 Engineering an intracellular pathway for major histocompatibility complex class II presentation of antigens Proceedings of the National Academy of Sciences of the United States of America 92 25 11671 11675
  33. 33. Miller D. G. Miller A. D. 1994 A family of retroviruses that utilize related phosphate transporters for cell entry J Virol 68 12 8270 8276
  34. 34. Lopes L. Fletcher K. Ikeda Y. Collins M. 2006 Lentiviral vector expression of tumour antigens in dendritic cells as an immunotherapeutic strategy Cancer Immunol Immunother 55 8 1011 1016
  35. 35. Palmowski M. J. Lopes L. Ikeda Y. Salio M. Cerundolo V. Collins M. K. 2004 Intravenous injection of a lentiviral vector encoding NY-ESO-1 induces an effective CTL response J Immunol 172 3 1582 7
  36. 36. Christodoulopoulos I. Cannon P. M. 2001 Sequences in the cytoplasmic tail of the gibbon ape leukemia virus envelope protein that prevent its incorporation into lentivirus vectors J Virol 75 9 4129 4138
  37. 37. Rasko J. E. Battini J. L. Gottschalk R. J. Mazo I. Miller A. D. 1999 The RD114/simian type D retrovirus receptor is a neutral amino acid transporter Proc Natl Acad Sci U S A 96 5 2129 2134
  38. 38. Karwacz K. Mukherjee S. Apolonia L. Blundell M. P. Bouma G. Escors D. Collins M. K. Thrasher A. J. 2009 Nonintegrating lentivector vaccines stimulate prolonged T-cell and antibody responses and are effective in tumor therapy J Virol 83 7 3094 103
  39. 39. Goold H. D. Escors D. Conlan T. J. Chakraverty R. Bennett C. L. 2011 Conventional dendritic cells are required for the activation of helper-dependent CD8 T cell responses to a model antigen after cutaneous vaccination with lentiviral vectors J Immunol,186 8 4565 4572
  40. 40. Escors D. Lopes L. Lin R. Hiscott J. Akira S. Davis R. J. Collins M. K. 2008 Targeting dendritic cell signalling to regulate the response to immunisation Blood 111 6 3050 3061
  41. 41. Breckpot K. Escors D. 2009 Dendritic Cells for Active Anti-cancer Immunotherapy: Targeting Activation Pathways Through Genetic Modification Endocrine, metabolic & immune disorders drug targets 9 328 343
  42. 42. Pasare C. Medzhitov R. 2005 Toll-like receptors: linking innate and adaptive immunity Adv Exp Med Biol 560 11 18
  43. 43. Takeda K. Akira S. 2005 Toll-like receptors in innate immunity Int Immunol 17 1 1 14
  44. 44. Brown B. D. Sitia G. Annoni A. Hauben E. Sergi Sergi. L. Zingale A. Roncarolo M. G. Guidotti L. G. Naldini L. 2006 In vivo administration of lentiviral vectors triggers a type I interferon response that restricts hepatocyte gene transfer and promotes vector clearance Blood
  45. 45. Rossetti M. Gregori S. Hauben E. Brown B. D. Sergi L. S. Naldini L. Roncarolo M. G. 2011 HIV-1-derived lentiviral vectors directly activate plasmacytoid dendritic cells, which in turn induce the maturation of myeloid dendritic cells Hum Gene Ther 22 2 177 188
  46. 46. Harman A. N. Wilkinson J. Bye C. R. Bosnjak L. Stern J. L. Nicholle M. Lai J. Cunningham A. L. 2006 HIV induces maturation of monocyte-derived dendritic cells and Langerhans cells J Immunol 177 10 7103 7113
  47. 47. Pichlmair A. Diebold S. S. Gschmeissner S. Takeuchi Y. Ikeda Y. Collins M. K. Reis e Sousa C. 2007 Tubulovesicular structures within vesicular stomatitis virus G protein-pseudotyped lentiviral vector preparations carry DNA and stimulate antiviral responses via Toll-like receptor 9 J Virol 81 2 539 47
  48. 48. Bao L. Guo H. Huang X. Tammana S. Wong M. Mc Ivor R. S. Zhou X. 2009 High-titer lentiviral vectors stimulate fetal calf serum-specific human CD4 T-cell responses: implications in human gene therapy Gene Ther 16 6 788 795
  49. 49. Akazawa T. Shingai M. Sasai M. Ebihara T. Inoue N. Matsumoto M. Seya T. 2007 Tumor immunotherapy using bone marrow-derived dendritic cells overexpressing Toll-like receptor adaptors FEBS Lett 581 18 3334 3340
  50. 50. Bagneris C. Ageichik A. V. Cronin N. Wallace B. Collins M. Boshoff C. Waksman G. Barrett T. 2008 Crystal structure of a vFlip-IKKgamma complex: insights into viral activation of the IKK signalosome Molecular cell 30 5 620 31
  51. 51. Efklidou S. Bailey R. Field N. Noursadeghi M. Collins M. K. 2008 vFLIP from KSHV inhibits anoikis of primary endothelial cells J Cell Sci 121 Pt 4 450 7
  52. 52. Field N. Low W. Daniels M. Howell S. Daviet L. Boshoff C. Collins M. 2003 KSHV vFLIP binds to IKK-gamma to activate IKK J Cell Sci 116 Pt 18 3721 3728
  53. 53. Shimizu A. Baratchian M. Takeuchi Y. Escors D. Macdonald D. Barrett T. Bagneris C. Collins M. Noursadeghi M. 2011 Kaposi’s sarcoma-associated herpesvirus vFLIP and human T cell lymphotropic virus type 1 Tax oncogenic proteins activate IkappaB kinase subunit gamma by different mechanisms independent of the physiological cytokine-induced pathways J Virol 85 14 7444 8
  54. 54. Rowe H. M. Lopes L. Brown N. Efklidou S. Smallie T. Karrar S. Kaye P. M. Collins M. K. 2009 Expression of vFLIP in a lentiviral vaccine vector activates NF-{kappa}B, matures dendritic cells, and increases CD8+ T-cell responses J Virol 83 4 1555 62
  55. 55. Breckpot K. Aerts-Toegaert C. Heirman C. Peeters U. Beyaert R. Aerts J. L. Thielemans K. 2009 Attenuated expression of A20 markedly increases the efficacy of double-stranded RNA-activated dendritic cells as an anti-cancer vaccine J Immunol, 182 2 860 870
  56. 56. Song X. T. Evel-Kabler K. Shen L. Rollins L. Huang X. F. Chen S. Y. 2008 A20 is an antigen presentation attenuator, and its inhibition overcomes regulatory T cell-mediated suppression Nat Med, 14 3 258 265
  57. 57. Koya R. C. Kasahara N. Favaro P. M. Lau R. Ta H. Q. Weber J. S. Stripecke R. 2003 Potent maturation of monocyte-derived dendritic cells after CD40L lentiviral gene delivery J Immunother 26 5 451 60
  58. 58. Koya R. C. Kimura T. Ribas A. Rozengurt N. Lawson G. W. Faure-Kumar E. Wang H. J. Herschman H. Kasahara N. Stripecke R. 2007 Lentiviral vector-mediated autonomous differentiation of mouse bone marrow cells into immunologically potent dendritic cell vaccines Mol Ther 15 5 971 80
  59. 59. Karwacz K. Arce F. Bricogne C. Kochan G. Escors D. 2012 PD-L1 co-stimulation, ligand-induced TCR down-modulation and anti-tumor immunotherapy Oncoimmunology 1 1 86 88
  60. 60. Escors D. Bricogne C. Arce F. Kochan G. Karwacz K. 2011 On the Mechanism of T cell receptor down-modulation and its physiological significance The journal of bioscience and medicine 1 1
  61. 61. He Y. Zhang J. Mi Z. Robbins P. Falo L. D. Jr. 2005 Immunization with lentiviral vector-transduced dendritic cells induces strong and long-lasting T cell responses and therapeutic immunity J Immunol 174 6 3808 17
  62. 62. Park T. S. Rosenberg S. A. Morgan R. A. 2011 Treating cancer with genetically engineered T cells Trends Biotechnol
  63. 63. Kochenderfer J. N. Yu Z. Frasheri D. Restifo N. P. Rosenberg S. A. 2010 Adoptive transfer of syngeneic T cells transduced with a chimeric antigen receptor that recognizes murine CD19 can eradicate lymphoma and normal B cells Blood 116 19 3875 3886
  64. 64. Parkhurst M. R. Yang J. C. Langan R. C. Dudley M. E. Nathan D. A. Feldman S. A. Davis J. L. Morgan R. A. Merino M. J. Sherry R. M. Hughes M. S. Kammula U. S. Phan G. Q. Lim R. M. Wank S. A. Restifo N. P. Robbins P. F. Laurencot C. M. Rosenberg S. A. 2011 T cells targeting carcinoembryonic antigen can mediate regression of metastatic colorectal cancer but induce severe transient colitis Mol Ther 19 3 620 626
  65. 65. Pule M. A. Savoldo B. Myers G. D. Rossig C. Russell H. V. Dotti G. Huls M. H. Liu E. Gee A. P. Mei Z. Yvon E. Weiss H. L. Liu H. Rooney C. M. Heslop H. E. Brenner M. K. 2008 Virus-specific T cells engineered to coexpress tumor-specific receptors: persistence and antitumor activity in individuals with neuroblastoma Nat Med 14 11 1264 1270
  66. 66. Till B. G. Jensen M. C. Wang J. Chen E. Y. Wood B. L. Greisman H. A. Qian X. James S. E. Raubitschek A. Forman S. J. Gopal A. K. Pagel J. M. Lindgren C. G. Greenberg P. D. Riddell S. R. Press O. W. 2008 Adoptive immunotherapy for indolent non-Hodgkin lymphoma and mantle cell lymphoma using genetically modified autologous CD 20-specific T cellsBlood 112 6 2261 2271
  67. 67. Perro M. Tsang J. Xue S. A. Escors D. Cesco-Gaspere M. Pospori C. Gao L. Hart D. Collins M. Stauss H. Morris E. C. 2010 Generation of multi-functional antigen-specific human T-cells by lentiviral TCR gene transfer Gene Ther 17 721 732
  68. 68. Cavalieri S. Cazzaniga S. Geuna M. Magnani Z. Bordignon C. Naldini L. Bonini C. 2003 Human T lymphocytes transduced by lentiviral vectors in the absence of TCR activation maintain an intact immune competence Blood 102 2 497 505
  69. 69. Ducrey-Rundquist O. Guyader M. Trono D. 2002 Modalities of interleukin-7-induced human immunodeficiency virus permissiveness in quiescent T lymphocytes J Virol 76 18 9103 9111
  70. 70. Morgan R. A. Dudley M. E. Wunderlich J. R. Hughes M. S. Yang J. C. Sherry R. M. Royal R. E. Topalian S. L. Kammula U. S. Restifo N. P. Zheng Z. Nahvi A. de Vries C. R. Rogers-Freezer L. J. Mavroukakis S. A. Rosenberg S. A. 2006 Cancer regression in patients after transfer of genetically engineered lymphocytes. Science 314 5796 126 129
  71. 71. Frecha C. Costa C. Negre D. Gauthier E. Russell S. J. Cosset F. L. Verhoeyen E. 2008 Stable transduction of quiescent T cells without induction of cycle progression by a novel lentiviral vector pseudotyped with measles virus glycoproteins Blood 112 13 4843 4852
  72. 72. Frecha C. Costa C. Levy C. Negre D. Russell S. J. Maisner A. Salles G. Peng K. W. Cosset F. L. Verhoeyen E. 2009 Efficient and stable transduction of resting B lymphocytes and primary chronic lymphocyte leukemia cells using measles virus gp displaying lentiviral vectors Blood 114 15 3173 3180
  73. 73. Frecha C. Levy C. Cosset F. L. Verhoeyen E. 2010 Advances in the field of lentivector-based transduction of T and B lymphocytes for gene therapy Mol Ther 18 10 1748 1757
  74. 74. Griesemer A. D. Sorenson E. C. Hardy M. A. 2010 The role of the thymus in tolerance. Transplantation 90 5 465 474
  75. 75. Hori S. Nomura T. Sakaguchi S. 2003 Control of regulatory T cell development by the transcription factor Foxp3 Science 299 5609 1057 1061
  76. 76. Sakaguchi S. 2003 The origin of FOXP 3-expressing CD4+ regulatory T cells: thymus or periphery J Clin Invest 112 9 1310 1312
  77. 77. Sakaguchi S. Yamaguchi T. Nomura T. Ono M. 2008 Regulatory T cells and immune tolerance Cell 133 5 775 787
  78. 78. Arce F. Breckpot K. Stephenson H. Karwacz K. Ehrenstein M. R. Collins M. Escors D. 2011 Selective ERK activation differentiates mouse and human tolerogenic dendritic cells, expands antigen-specific regulatory T cells, and suppresses experimental inflammatory arthritis Arthritis and rheumatism 63 84 95
  79. 79. Mahnke K. Qian Y. Knop J. Enk A. H. 2003 Induction of CD4+/CD25+ regulatory T cells by targeting of antigens to immature dendritic cells Blood 101 12 4862 9
  80. 80. O’Garra A. Vieira P. 2004 Regulatory T cells and mechanisms of immune system control Nat Med 10 8 801 5
  81. 81. O’Garra A. Vieira P. L. Vieira P. Goldfeld A. E. 2004 IL-10-producing and naturally occurring CD4+ Tregs: limiting collateral damage J Clin Invest 114 10 1372 1378
  82. 82. Peng Y. Laouar Y. Li M. O. Green E. A. Flavell R. A. 2004 TGF-beta regulates in vivo expansion of Foxp3-expressing CD4+CD25+ regulatory T cells responsible for protection against diabetes Proc Natl Acad Sci U S A 101 13 4572 4577
  83. 83. Bonifaz L. Bonnyay D. Mahnke K. Rivera M. Nussenzweig M. C. Steinman R. M. 2002 Efficient targeting of protein antigen to the dendritic cell receptor DEC-205 in the steady state leads to antigen presentation on major histocompatibility complex class I products and peripheral CD8+ T cell tolerance J Exp Med 196 12 1627 38
  84. 84. Dhodapkar M. V. Steinman R. M. 2002 Antigen-bearing immature dendritic cells induce peptide-specific CD8(+) regulatory T cells in vivo in humans Blood 100 1 174 177
  85. 85. Hawiger D. Inaba K. Dorsett Y. Guo M. Mahnke K. Rivera M. Ravetch J. V. Steinman R. M. Nussenzweig M. C. 2001 Dendritic cells induce peripheral T cell unresponsiveness under steady state conditions in vivo J Exp Med 194 6 769 779
  86. 86. Kretschmer K. Apostolou I. Hawiger D. Khazaie K. Nussenzweig M. C. von Boehmer H. 2005 Inducing and expanding regulatory T cell populations by foreign antigen Nat Immunol 6 12 1219 27
  87. 87. Rutella S. Danese S. Leone G. 2006 Tolerogenic dendritic cells: cytokine modulation comes of age Blood 108 5 1435 40
  88. 88. Manicassamy S. Ravindran R. Deng J. Oluoch H. Denning T. L. Kasturi S. P. Rosenthal K. M. Evavold B. D. Pulendran B. 2009 Toll-like receptor 2-dependent induction of vitamin A-metabolizing enzymes in dendritic cells promotes T regulatory responses and inhibits autoimmunity Nat Med 15 4 401 409
  89. 89. Ilarregui J. M. Croci D. O. Bianco G. A. Toscano M. A. Salatino M. Vermeulen M. E. Geffner J. R. Rabinovich G. A. 2009 Tolerogenic signals delivered by dendritic cells to T cells through a galectin-1-driven immunoregulatory circuit involving interleukin 27 and interleukin 10. Nat Immunol 10 9 981 991
  90. 90. Dillon S. Agrawal S. Banerjee K. Letterio J. Denning T. L. Oswald-Richter K. Kasprowicz D. J. Kellar K. Pare J. van Dyke T. Ziegler S. Unutmaz D. Pulendran B. 2006 Yeast zymosan, a stimulus for TLR2 and dectin-1, induces regulatory antigen-presenting cells and immunological tolerance J Clin Invest 116 4 916 928
  91. 91. Corinti S. Albanesi C. la Sala A. Pastore S. Girolomoni G. 2001 Regulatory activity of autocrine IL-10 on dendritic cell functions J Immunol 166 7 4312 8
  92. 92. Ghiringhelli F. Puig P. E. Roux S. Parcellier A. Schmitt E. Solary E. Kroemer G. Martin F. Chauffert B. Zitvogel L. 2005 Tumor cells convert immature myeloid dendritic cells into TGF-beta-secreting cells inducing CD4+CD25+ regulatory T cell proliferation J Exp Med 202 7 919 929
  93. 93. Saraiva M. O’Garra A. 2010 The regulation of IL-10 production by immune cells Nature reviews 10 3 170 181
  94. 94. Kuhn R. Lohler J. Rennick D. Rajewsky K. Muller W. 1993 Interleukin-10-deficient mice develop chronic enterocolitis Cell 75 2 263 274
  95. 95. Takayama T. Nishioka Y. Lu L. Lotze M. T. Tahara H. Thomson A. W. 1998 Retroviral delivery of viral interleukin-10 into myeloid dendritic cells markedly inhibits their allostimulatory activity and promotes the induction of T-cell hyporesponsiveness Transplantation 66 12 1567 74
  96. 96. Sakuishi K. Apetoh L. Sullivan J. M. Blazar B. R. Kuchroo V. K. Anderson A. C. 2010 Targeting Tim-3 and PD-1 pathways to reverse T cell exhaustion and restore anti-tumor immunity J Exp Med 207 10 2187 2194
  97. 97. Wang L. Pino-Lagos K. de Vries V. C. Guleria I. Sayegh M. H. Noelle R. J. 2008 Programmed death 1 ligand signaling regulates the generation of adaptive Foxp3+CD4+ regulatory T cells Proc Natl Acad Sci U S A 105 27 9331 9336
  98. 98. Sica G. L. Choi I. H. Zhu G. Tamada K. Wang S. D. Tamura H. Chapoval A. I. Flies D. B. Bajorath J. Chen L. 2003 B7-H4, a molecule of the B7 family, negatively regulates T cell immunity Immunity 18 6 849 861
  99. 99. Belladonna M. L. Orabona C. Grohmann U. Puccetti P. 2009 TGF-beta and kynurenines as the key to infectious tolerance Trends in molecular medicine 15 2 41 9
  100. 100. Cobbold S. P. Adams E. Farquhar C. A. Nolan K. F. Howie D. Lui K. O. Fairchild P. J. Mellor A. L. Ron D. Waldmann H. 2009 Infectious tolerance via the consumption of essential amino acids and mTOR signaling Proc Natl Acad Sci U S A 106 29 12055 12060
  101. 101. Fallarino F. Vacca C. Orabona C. Belladonna M. L. Bianchi R. Marshall B. Keskin D. B. Mellor A. L. Fioretti M. C. Grohmann U. Puccetti P. 2002 Functional expression of indoleamine 2,3-dioxygenase by murine CD8 alpha(+) dendritic cells Int Immunol 14 1 65 68
  102. 102. Mellor A. L. Munn D. H. 2004 IDO expression by dendritic cells: tolerance and tryptophan catabolism Nature reviews 4 10 762 74
  103. 103. Munder M. 2009 Arginase: an emerging key player in the mammalian immune system British journal of pharmacology 158 3 638 651
  104. 104. Norian L. A. Rodriguez P. C. O’Mara L. A. Zabaleta J. Ochoa A. C. Cella M. Allen P. M. 2009 Tumor-infiltrating regulatory dendritic cells inhibit CD8+ T cell function via L-arginine metabolism Cancer Res 69 7 3086 3094
  105. 105. Lee W. C. Zhong C. Qian S. Wan Y. Gauldie J. Mi Z. Robbins P. D. Thomson A. W. Lu L. 1998 Phenotype, function, and in vivo migration and survival of allogeneic dendritic cell progenitors genetically engineered to express TGF-beta Transplantation 66 12 1810 1817
  106. 106. Morita Y. Yang J. Gupta R. Shimizu K. Shelden E. A. Endres J. Mule J. J. Mc Donagh K. T. Fox D. A. 2001 Dendritic cells genetically engineered to express IL-4 inhibit murine collagen-induced arthritis J Clin Invest 107 10 1275 1284
  107. 107. Henry E. Desmet C. J. Garze V. Fievez L. Bedoret D. Heirman C. Faisca P. Jaspar F. J. Gosset P. Jacquet A. P. Desmecht D. Thielemans K. Lekeux P. Moser M. Bureau F. 2008 Dendritic cells genetically engineered to express IL-10 induce long-lasting antigen-specific tolerance in experimental asthma J Immunol 181 10 7230 7242
  108. 108. Delgado M. Toscano M. G. Benabdellah K. Cobo M. O’Valle F. Gonzalez-Rey E. Martin F. 2008 In vivo delivery of lentiviral vectors expressing vasoactive intestinal peptide complementary DNA as gene therapy for collagen-induced arthritis Arthritis and rheumatism 58 4 1026 1037
  109. 109. Toscano M. G. Delgado M. Kong W. Martin F. Skarica M. Ganea D. 2010 Dendritic cells transduced with lentiviral vectors expressing VIP differentiate into VIP-secreting tolerogenic-like DCs Mol Ther, 18 5 1035 1045
  110. 110. Agrawal A. Dillon S. Denning T. L. Pulendran B. 2006 ERK1-/- mice exhibit Th1 cell polarization and increased susceptibility to experimental autoimmune encephalomyelitis J Immunol 176 10 5788 5796
  111. 111. Anastasaki C. Estep A. L. Marais R. Rauen K. A. Patton E. E. 2009 Kinase-activating and kinase-impaired cardio-facio-cutaneous syndrome alleles have activity during zebrafish development and are sensitive to small molecule inhibitors Human molecular genetics 18 14 2543 2554
  112. 112. Caparros E. Munoz P. Sierra-Filardi E. Serrano-Gomez D. Puig-Kroger A. Rodriguez-Fernandez J. L. Mellado M. Sancho J. Zubiaur M. Corbi A. L. 2006 DC-SIGN ligation on dendritic cells results in ERK and PI3K activation and modulates cytokine production Blood 107 10 3950 3958
  113. 113. Pages G. Brunet A. L’Allemain G. Pouyssegur J. 1994 Constitutive mutant and putative regulatory serine phosphorylation site of mammalian MAP kinase kinase (MEK1) Embo J 13 13 3003 3010
  114. 114. Raingeaud J. Whitmarsh A. J. Barrett T. Derijard B. Davis R. J. 1996 MKK3- and MKK6-regulated gene expression is mediated by the 38 mitogen-activated protein kinase signal transduction pathway. Mol Cell Biol 16 3 1247 1255
  115. 115. Li Y. Chu N. Rostami A. Zhang G. X. 2006 Dendritic cells transduced with SOCS-3 exhibit a tolerogenic/DC2 phenotype that directs type 2 Th cell differentiation in vitro and in vivo J Immunol 177 3 1679 88
  116. 116. Zhang Y. Yang H. Xiao B. Wu M. Zhou W. Li J. Li G. Christadoss P. 2009 Dendritic cells transduced with lentiviral-mediated RelB-specific ShRNAs inhibit the development of experimental autoimmune myasthenia gravis Molecular immunology 46 4 657 667
  117. 117. Yang M. Sun L. Wang S. Ko K. H. Xu H. Zheng B. J. Cao X. Lu L. 2010 Novel function of B cell-activating factor in the induction of IL-10-producing regulatory B cells. J Immunol 184 7 3321 3325
  118. 118. Batten M. Groom J. Cachero T. G. Qian F. Schneider P. Tschopp J. Browning J. L. Mackay F. 2000 BAFF mediates survival of peripheral immature B lymphocytes J Exp Med 192 10 1453 1466
  119. 119. Lai Kwan. Lam Q. King Hung. Ko O. Zheng B. J. Lu L. 2008 Local BAFF gene silencing suppresses Th17-cell generation and ameliorates autoimmune arthritis Proc Natl Acad Sci U S A 105 39 14993 14998

Written By

Ines Dufait, Therese Liechtenstein, Alessio Lanna, Roberta Laranga, Antonella Padella, Christopher Bricogne, Frederick Arce, Grazyna Kochan, Karine Breckpot and David Escors

Submitted: 02 June 2012 Published: 27 February 2013