Development of the Nanobody display technology to target lentiviral vectors to antigen-presenting cells

Goyvaerts, Cleo; Groeve, Kurt De; Dingemans, Jozef; Lint, Sandra Van; Robays, Lander; Heirman, Carlo; Reiser, Jakob; Xy, Zhang; Thielemans, Kris; Baetseĺier, Patrick De; Raes, Geert; Breckpot, Karine

doi:10.1038/gt.2011.206

Cited by 52 publications

(63 citation statements)

References 47 publications

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“…To allow in vivo bioluminescence imaging (BLI) of intracranial tumors, MO4 cells were transduced with a lentiviral construct encoding both tNGFR and FLuc (pHR trip CMV luc2-Ires-tNGFR SIN, described in ref. 43). 43 Stable and highly FLuc positive MO4 cells were subsequently enriched via flow cytometry using the expression levels of the tNGFR protein as a reference.…”

Section: Tumor Cell Linesmentioning

confidence: 99%

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Axitinib increases the infiltration of immune cells and reduces the suppressive capacity of monocytic MDSCs in an intracranial mouse melanoma model

et al. 2015

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Section: Tumor Cell Linesmentioning

confidence: 99%

“…43). 43 Stable and highly FLuc positive MO4 cells were subsequently enriched via flow cytometry using the expression levels of the tNGFR protein as a reference. We refer to the stable luciferase-expressing cell line used as MO4-FLuc.…”

Section: Tumor Cell Linesmentioning

confidence: 99%

Axitinib increases the infiltration of immune cells and reduces the suppressive capacity of monocytic MDSCs in an intracranial mouse melanoma model

et al. 2015

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“…Furthermore, the lentivectors' envelopes are well suited for engineering, enabling the design of targeted lentivectors. Several methods have been described to redirect lentivectors to specific antigen-presenting cells (9)(10)(11)(12). However, to our knowledge subset-specific delivery of transgenes has not been described.…”

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confidence: 99%

“…This innovative approach exploits the budding mechanism of lentivectors to incorporate both Nbs and the binding-defective but fusion-competent vesicular stomatitis virus glycoprotein (VSV.Gs) on the viral surface (10,14). Producer cells that were modified to express Nb BCII10, Nb DC2.1 or Nb R3_13 on their cell membrane were generated by transducing HEK293T cells with lentivectors encoding the membrane-bound form of these Nbs (10).…”

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confidence: 99%

Targeting of Human Antigen-Presenting Cell Subsets

Goyvaerts

Dingemans

Groeve

et al. 2013

J Virol

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dAntigen-presenting cells are a heterogeneous group of cells that are characterized by their functional specialization. Consequently, targeting specific antigen-presenting cell subsets offers opportunities to induce distinct T cell responses. Here we report on the generation and use of nanobodies (Nbs) to target lentivectors specifically to human lymph node-resident myeloid dendritic cells, demonstrating that Nbs represent a powerful tool to redirect lentivectors to human antigen-presenting cell subsets. Since the discovery of dendritic cells (DCs) in 1973, these antigen-presenting cells have been at the center of attention in vaccine development (1). In the past decade, an extra dimension of complexity was introduced by revealing an unforeseen diversity of DC subsets with distinct functions and locations (2-4). Since DC subsets induce distinct immune responses, it might be beneficial to develop strategies to target particular DC subsets and as such exploit the immune system to its full potential (5).Numerous animal studies have proven the efficiency and safety of lentivectors (LVs) as vaccination moieties (6, 7). As lentivectors are intrinsically immunogenic, they deliver both antigens and activation signals to antigen-presenting cells (8). Furthermore, the lentivectors' envelopes are well suited for engineering, enabling the design of targeted lentivectors. Several methods have been described to redirect lentivectors to specific antigen-presenting cells (9-12). However, to our knowledge subset-specific delivery of transgenes has not been described.Targeting myeloid DCs could be advantageous as they are considered to be important mediators of antigen-specific immunity. They are able to induce proper and oriented stimulation of CD4 ϩ T helper 1 and CD8 ϩ cytotoxic T cells. In addition, targeting may reduce the risk of adverse reactions such as autoimmune responses or induction of tolerance due to transgene expression and presentation by non-antigen-presenting cells or tolerogenic DC subtypes. Finally, as myeloid DCs have a limited life span, their targeting should result in a natural clearance of the lentivector and as such in a reduction of the risk of insertional mutagenesis.We recently delivered a proof of concept on the use of nanobodies (Nbs) to target lentivectors to antigen-presenting cells (10). Nbs or V H H fragments are antibody fragments of about 12 to 25 kDa that are engineered from heavy-chain-only antibodies found in Camelidae. Because of their size and target affinity, they are of particular interest as targeting moieties. In the present study, we further refined the transduction profile of lentivectors, targeting them to human myeloid DCs using Nbs.Two Nb libraries, derived from peripheral blood lymphocytes of llamas that were immunized with immature or lipopolysaccharide-stimulated murine bone marrow-derived DCs, were at our disposal (13). These were screened for cross-reactivity with hu-

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“…As shown, the latter can be accomplished by the use of target cell-specific promoters. Moreover, strategies to direct the transduction pattern of LVs using cell-specific nanobodies will further improve the safety and potential efficacy of LVs as an off-the-shelf therapeutic (44,45).…”

Section: Discussionmentioning

confidence: 99%

Proinflammatory Characteristics of SMAC/DIABLO-Induced Cell Death in Antitumor Therapy

et al. 2012

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Molecular mimetics of the caspase activator second mitochondria-derived activator of caspase (SMAC) are being investigated for use in cancer therapy, but an understanding of in vivo effects remains incomplete. In this study, we offer evidence that SMAC mimetics elicit a proinflammatory cell death in cancer cells that engages an adaptive antitumor immune response. Cancer cells of different histologic origin underwent apoptosis when transduced with lentiviral vectors encoding a cytosolic form of the SMAC mimetic LVtSMAC. Strikingly, treatment of tumor-bearing mice with LV-tSMAC resulted in the induction of apoptosis, activation of antitumor immunity, and enhanced survival. Antitumor immunity was accompanied by an increase of tumor-infiltrating lymphocytes displaying low PD-1 expression, high lytic capacity, and high levels of IFN-g when stimulated. We also noted in vivo a decrease in regulatory T cells along with in vitro activation of tumor-specific CD8 þ T cells by dendritic cells (DC) isolated from tumor draining lymph nodes. Last, tumorspecific cytotoxic T cells were also found to be activated in vivo. Mechanistic analyses showed that transduction of cancer cells with LV-tSMAC resulted in exposure of calreticulin but not release of HMGB1 or ATP. Nevertheless, DCs were activated upon engulfment of dying cancer cells. Further validation of these findings was obtained by their extension in a model of human melanoma using transcriptionally targeted LV-tSMAC. Together, our findings suggest that SMAC mimetics can elicit a proinflammatory cell death that is sufficient to activate adaptive antitumor immune responses in cancer. Cancer Res; 72(6);

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Development of the Nanobody display technology to target lentiviral vectors to antigen-presenting cells

Cited by 52 publications

References 47 publications

Axitinib increases the infiltration of immune cells and reduces the suppressive capacity of monocytic MDSCs in an intracranial mouse melanoma model

Axitinib increases the infiltration of immune cells and reduces the suppressive capacity of monocytic MDSCs in an intracranial mouse melanoma model

Targeting of Human Antigen-Presenting Cell Subsets

Proinflammatory Characteristics of SMAC/DIABLO-Induced Cell Death in Antitumor Therapy

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