Vectors For Cancer Immunotherapy Research Articles

Abstract 2048: In vitro and in vivo characterization of non-oncolytic engineered arenavirus vectors for cancer immunotherapy

Abstract Cytotoxic CD8+ T cells are paramount for effective cancer immunotherapy. We engineered two distantly related arenaviruses, lymphocytic choriomeningitis virus (LCMV) and Pichinde virus (PICV), encoding the same non-oncogenic HPV16 E7E6 fusion protein to effectively mount and maintain tumor specific CD8+ T cell responses. The designated vectors, HB-201 (LCMV) and HB-202 (PICV), are currently in an ongoing Phase I/II open labelled clinical trial of HB-201 single vector therapy and HB-202/HB-201 alternating two-vector therapy in patients with treatment-refractory HPV16+ cancers (NCT04180215). To characterize the immunogenic properties of both vectors, we designed a comprehensive set of preclinical and translational experiments utilizing human PBMCs, artificial APCs and HPV16 E6 and E7 specific reporter T cells. This was done in conjunction with a preclinical model for HPV16 associated cancer (TC-1). HB-201 (LCMV) and HB-202 (PICV) surrogate vectors encoding GFP readily infected human monocytes, plasmacytoid dendritic, and conventional dendritic cells. In addition, HB-201 and HB-202 vectors successfully induced antigen presentation of E7 and E6 epitopes as evidenced by TCR reporter cells. Innate immune cell activation and cytokine responses to HB-202 were slightly lower compared to HB-201, findings which are consistent with different memory profiles of E7 specific CD8+ T cells in non-tumor bearing mice. In a therapeutic setting of the TC-1 tumor model, both vectors were comparably effective, showing major infiltration of CD8+ T cells into the tumor microenvironment, tumor growth delay and a significantly prolonged survival already after a single administration. In this data set we have confirmed the strong immunogenicity of the engineered arenavirus platform leading to efficient tumor control in a relevant mouse model for HPV16+ cancers, which further supports the clinical development of our novel arenavirus platform. Citation Format: Josipa Raguz, Theresa Kleissner, Sandra Rosskopf, Mohamed Habbeddine, Katharina Lechner, Valentin Just, Donna Edwards, Igor Matushansky, Christoph Lampert, Klaus K. Orlinger, Henning Lauterbach. In vitro and in vivo characterization of non-oncolytic engineered arenavirus vectors for cancer immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2048.

Nonreplicating Adenoviral Vectors: Improving Tropism and Delivery of Cancer Gene Therapy.

Simple SummaryThe treatment of cancer has progressed greatly with the advent of immunotherapy and gene therapy, including the use of nonreplicating adenoviral vectors to deliver genes with antitumor activity for cancer gene therapy. Even so, the successful application of these vectors may benefit from modifications in their design, including their molecular structure, so that specificity for the target cell is increased and off-target effects are minimized. With such improvements, we may find new opportunities for systemic administration of adenoviral vectors as well as the delivery of strategic antigen targets of an antitumor immune response. We propose that the improvement of nonreplicating adenoviral vectors will allow them to continue to hold a key position in cancer gene therapy and immunotherapy.Recent preclinical and clinical studies have used viral vectors in gene therapy research, especially nonreplicating adenovirus encoding strategic therapeutic genes for cancer treatment. Adenoviruses were the first DNA viruses to go into therapeutic development, mainly due to well-known biological features: stability in vivo, ease of manufacture, and efficient gene delivery to dividing and nondividing cells. However, there are some limitations for gene therapy using adenoviral vectors, such as nonspecific transduction of normal cells and liver sequestration and neutralization by antibodies, especially when administered systemically. On the other hand, adenoviral vectors are amenable to strategies for the modification of their biological structures, including genetic manipulation of viral proteins, pseudotyping, and conjugation with polymers or biological membranes. Such modifications provide greater specificity to the target cell and better safety in systemic administration; thus, a reduction of antiviral host responses would favor the use of adenoviral vectors in cancer immunotherapy. In this review, we describe the structural and molecular features of nonreplicating adenoviral vectors, the current limitations to their use, and strategies to modify adenoviral tropism, highlighting the approaches that may allow for the systemic administration of gene therapy.

Vaccine strain Listeria monocytogenes abscess in a dog: a case report

BackgroundListeria monocytogenes is a promising therapeutic vaccine vector for cancer immunotherapy. Although highly attenuated, three cases of systemic listeriosis have been reported in people following treatment with Listeria-based therapeutic vaccines. This complication has thus far not been reported in canine patients.Case presentationA dog previously diagnosed with osteoblastic osteosarcoma was presented for care following administration of three doses of the Canine Osteosarcoma Vaccine-Live Listeria Vector. On routine staging chest radiographs, mild sternal lymphadenopathy and a right caudoventral thoracic mass effect were noted. Further evaluation of the mass effect with computed tomography and ultrasound revealed a cavitated mass associated with the 7th right rib. Aspirates of the mass cultured positive for Listeria monocytogenes. The mass and associated ribs were surgically removed. Histopathology was consistent with metastatic osteoblastic osteosarcoma. Treatment was continued with doxorubicin chemotherapy and at the time of publication, the dog was alive over 1 year following diagnosis with no evidence of further disease progression. Genotyping of the abscess-derived L. monocytogenes was consistent with the vaccine strain.ConclusionsThis case represents the first veterinary case to describe development of a Listeria abscess following administration of a Listeria-based therapeutic vaccine.

The features of circulating and tumor-infiltrating γδ T cells in melanoma patients display critical perturbations with prognostic impact on clinical outcome

ABSTRACT γδT cells hold a pivotal role in tumor immunosurveillance through their prompt activation and cytokine secretion, their ability to kill tumor cells in an Human Leukocyte Antigen (HLA)-unrestricted manner, and their combination of features of both innate and adaptive immunity. These unique properties and functional plasticity render them very attractive both as targets and vectors for cancer immunotherapy. Yet, these potent and fascinating antitumor effectors have not been extensively explored in melanoma. We provided here a detailed investigation of the phenotypic and functional properties of circulating and tumor-infiltrating γδT cells in melanoma patients, and their impact on clinical evolution. High proportions of circulating- and tumor-infiltrating γδT and δ2+ subset were associated with better clinical outcome. We reported however that circulating and tumor-infiltrating γδT cells from melanoma patients displayed an altered expression of NCR, KIR, and immune checkpoints, and identified NKp44, PD1, 41BB/41BBL, TIM3, and LAG3 as crucial checkpoints allowing immune escape and tumor progression. Notably, melanoma drastically impaired the ability of γδT cells to exhibit activation molecules, secrete cytokines, and display cytotoxicity toward melanoma in response to stimulation with phosphoantigens. It drove them toward regulatory and Th17 profiles associated with poor clinical outcomes. Our study highlights that melanoma hijacked γδT cells to escape from immune control, and revealed that circulating and tumor-infiltrating γδT cell features are promising potential biomarkers of clinical evolution. Such understanding of the physiopathology of γδT cells may help designing new therapeutic approaches exploiting the antitumor potential of γδT cells while counteracting their skewing by tumors to improve patient outcomes.

Abstract LB-287: Pseudocowpox: A next generation viral vector for cancer immunotherapy

Abstract Engineered viral vectors are effective approaches to stimulate anti-tumor immunity, and change the tumor immune environment. Several viruses and strains have been developed to express tumor antigens and cytokines, and corresponding products are in advanced clinical trials. However, novel viral strains with improved immunogenic properties are sought. In this perspective, we screened a variety of poxviridae potentially usable in humans: Cowpox (CPX), Pseudocowpox (PCPV), Parapoxvirus Ovis (ORF), Myxoma virus (MYX), Swinepox (SWP), Yaba-like disease virus (YLDV), Raccoonpox (RCN), Cotia virus (CTV), and compared them to the well-established vaccine strain Modified Virus Ankara (MVA), and oncolytic Vaccinia Virus, strain Copenhagen (VV). Both in vitro with human primary immune cells, and in vivo with syngeneic mouse tumor models, PCPV proved to be a very promising vector for immunotherapy. Compared with MVA, PCPV induced a 1000-fold higher expression of IFN-alpha in human PBMCs, whereas SWPV and ORF displayed a lower 10 to 100-fold induction. Other viruses (i.e. VV, RCN, CTV, or MYX) did not raise the IFN-alpha level. PCPV was also shown to increase the level of GM-CSF, and to be safe for PBMCs, in contrast to ORF which displayed some cytotoxic effects. When tested for its capacity to trigger the expression of CD86, a co-stimulatory factor for T-cell activation, PCPV was shown to be superior than MVA in primary moDCs. Furthermore, PCPV treatment increased CD86 expression in human CD163+CD206+ “M2”-type macrophages derived from CD14+ monocytes, suggesting a shift to an antigen-presenting phenotype. In these cells, PCPV increased significantly the secretion of IL-18, IL-6 and IP-10, signing a conversion towards a less suppressive macrophage phenotype. Last, incubation of PCPV in a co-culture model overcame the immunosuppressive effect of MDSCs on human autologous CD8+ T. A GFP-encoding recombinant PCPV vector was generated, and we could demonstrate that PCPV was capable of infecting human primary immune cells, comparably to recombinant MVA vectors, except for activated T cells. A recombinant PCPV encoding for the HPV E7 protein was generated to assess the anti-tumor activity and immunogenicity in a syngeneic murine tumor model. Like MVA-E7, PCPV-E7 induced a strong cellular response (ELISPOT on splenocytes, and frequency of antigen-specific short-lived effector cells), but PCPV-E7 displayed a different cytokine/chemokine profile at the site of injection, with increased levels of pro-immune cytokines including IP-10, IFN-gamma, GM-CSF, IL-18, MIP-1 alpha, MIP-1 beta, IL-12 and IL-6. When injected intratumorally into fast growing MC-38 tumors, PCPV led to tumor control. Analysis of tumors infiltrates showed that PCPV treatment led to higher levels of neutrophils and decreased the frequency of MHCIIlo TAMs. Our data demonstrate that PCPV might display better properties than current viral vectors, in terms of local response and priming activity, of ability to induce effector T cells and to reshape the tumor infiltration profiles. It has the same capacity as other poxviruses to encode and deliver large genetic payload, which will be useful for designing advanced anti-tumor vaccines. Citation Format: Karola Rittner, Marine Ricordel, Caroline Tosch, Christine Thioudellet, Christelle Remy-Ziller, Marie-Christine Claudepierre, Chantal Hoffmann, Doris Schmitt, Benoit Sansas, Johann Foloppe, Philippe Erbs, Nathalie Silvestre, Kaïdre Bendjama, Eric Quemeneur. Pseudocowpox: A next generation viral vector for cancer immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-287.

Replicating viral vector platform exploits alarmin signals for potent CD8+ T cell-mediated tumour immunotherapy

Viral infections lead to alarmin release and elicit potent cytotoxic effector T lymphocyte (CTLeff) responses. Conversely, the induction of protective tumour-specific CTLeff and their recruitment into the tumour remain challenging tasks. Here we show that lymphocytic choriomeningitis virus (LCMV) can be engineered to serve as a replication competent, stably-attenuated immunotherapy vector (artLCMV). artLCMV delivers tumour-associated antigens to dendritic cells for efficient CTL priming. Unlike replication-deficient vectors, artLCMV targets also lymphoid tissue stroma cells expressing the alarmin interleukin-33. By triggering interleukin-33 signals, artLCMV elicits CTLeff responses of higher magnitude and functionality than those induced by replication-deficient vectors. Superior anti-tumour efficacy of artLCMV immunotherapy depends on interleukin-33 signalling, and a massive CTLeff influx triggers an inflammatory conversion of the tumour microenvironment. Our observations suggest that replicating viral delivery systems can release alarmins for improved anti-tumour efficacy. These mechanistic insights may outweigh safety concerns around replicating viral vectors in cancer immunotherapy.

Abstract A016: Multipotency of a CD40L-expressing recombinant vaccinia virus

Abstract Vaccinia Virus (VV) is a viral vector used in pre-clinical and clinical studies of active cancer immunotherapeutic strategies. CD40 receptor is expressed on a broad range of immune cells. Furthermore, CD40 expression has been reported also in nearly all B-cell malignancies and solid tumors of different origins. Interestingly, CD40 ligation might result in different biological responses depending on the cell types and to the strength of CD40 ligand (CD40L; CD154)-mediated stimulation of CD40 receptor. Based on this background, we have generated a replication incompetent CD40 ligand-expressing recombinant vaccinia virus (rVV40L) and we have investigated its antitumor activity. Notably, rVV40L-infection resulted in an improvement of (i) the antigen presenting capacity of CD40+ nonprofessional CD14+ monocytes leading (ii) to the rapid expansion and differentiation of human naïve CD8+ T cells into central memory T cells (TCM) specific for tumor associated antigens (TAAs), (ii) the induction of apoptotic effects on CD40+ established tumor cell lines of different origin and (iii) the antitumor activity, TNF-mediated, of macrophages. Collectively, these data indicate that rVV40L can be considered a successful viral vector for cancer immunotherapy according to its multiple mechanisms of action. Citation Format: Emanuele Trella. Multipotency of a CD40L-expressing recombinant vaccinia virus [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr A016.

Lentiviral vectors in cancer immunotherapy.

Basic science advances in cancer immunotherapy have resulted in various treatments that have recently shown success in the clinic. Many of these therapies require the insertion of genes into cells to directly kill them or to redirect the host's cells to induce potent immune responses. Other analogous therapies work by modifying effector cells for improved targeting and enhanced killing of tumor cells. Initial studies done using γ-retroviruses were promising, but safety concerns centered on the potential for insertional mutagenesis have highlighted the desire to develop other options for gene delivery. Lentiviral vectors (LVs) have been identified as potentially more effective and safer alternative delivery vehicles. LVs are now in use in clinical trials for many different types of inherited and acquired disorders, including cancer. This review will discuss current knowledge of LVs and the applications of this viral vector-based delivery vehicle to cancer immunotherapy.

Viral Vectors in Cancer Immunotherapy: Which Vector for Which Strategy?

Gene therapy involves the transfer of genetic information to a target cell to facilitate the production of therapeutic proteins and is now a realistic prospect as a cancer treatment. Gene transfer may be achieved through the use of both viral and non-viral delivery methods and the role of this method in the gene therapy of cancer has been demonstrated. Viruses represent an attractive vehicle for cancer gene therapy due to their high efficiency of gene delivery. Many viruses can mediate long term gene expression, while some are also capable of infecting both dividing and non-dividing cells. Given the broadly differing capabilities of various viral vectors, it is imperative that the functionality of the virus meets the requirements of the specific treatment. A number of immunogene therapy strategies have been undertaken, utilising a range of viral vectors, and studies carried out in animal models and patients have demonstrated the therapeutic potential of viral vectors to carry genes to cancer cells and induce anti-tumour immune responses. This review critically discusses the advances in the viral vector mediated delivery of immunostimulatory molecules directly to tumour cells, the use of viral vectors to modify tumour cells, the creation of whole cell vaccines and the direct delivery of tumour antigens in animal models and clinical trials, specifically in the context of the suitability of vector types for specific strategies.

Sphingosine Kinase and Sphingosine-1-Phosphate Regulate Migration, Endocytosis and Apoptosis of Dendritic Cells

Dendritic cells (DC) are inducers of primary immune responses and represent an attractive vector for cancer immunotherapy. Sphingosine kinase (SphK) and its product sphingosine-1-phosphate (S1P) play an important role in the regulation of immune cells and cancer, affecting processes such as differentiation, growth or migration. We studied the role of SphK and S1P on migration of DC. RT-PCR showed mRNA expression of SphK in DC, declining from immature (iDC) to mature DC (mDC) to antigen-loaded mDC. Expression of S1P receptors was S1P1 > S1P2 = S1P3, unrelated to maturation or antigen uptake. In transwell assays, iDC migrated towards SDF-1, MIP-1α, MCP and S1P, whereby S1P combined with a chemokine had a synergistic effect. mDC migrated towards 6Ckine and MIP-3β, but not towards S1P. The SphK-inhibitor dihydro-sphingosine (DHS) reduced migration of iDC but not of mDC. In addition S1P3-inhibitor suramin inhibited DC migration in response to S1P. DHS had a reverse effect on endocytosis, enhancing the uptake of FITC dextran. We also observed an anti-apoptotic effect of S1P on mDC for the first time. This indicates that SphK/S1P may play a role in accumulation of peripheral iDC at the location of antigen and subsequent antigen-uptake. These findings may help to optimise DC-based cancer immunotherapy by modulation of SphK/S1P.

From pathogen to medicine: HIV-1-derived lentiviral vectors as vehicles for dendritic cell based cancer immunotherapy

Over the years, the unique capacity of dendritic cells (DC) for efficient activation of naive T cells has led to their extensive use in cancer immunotherapy protocols. In order to be able to fulfil their role as antigen-presenting cells, the antigen of interest needs to be efficiently introduced and subsequently correctly processed and presented by the DC. For this purpose, a variety of both viral and non-viral antigen-delivery systems have been evaluated. Amongst those, HIV-1-derived lentiviral vectors have been used successfully to transduce DC. This review considers the use of HIV-1-derived lentiviral vectors to transduce human and murine DC for cancer immunotherapy. Lentivirally transduced DC have been shown to present antigenic peptides, prime transgene-specific T cells in vitro and elicit a protective cytotoxic T-lymphocyte (CTL) response in animal models. Different parameters determining the efficacy of transduction are considered. The influence of lentiviral transduction on the DC phenotype and function is described and the induction of immune responses by lentivirally transduced DC in vitro and in vivo is discussed in detail. In addition, direct in vivo administration of lentiviral vectors aiming at the induction of antigen-specific immunity is reviewed. This strategy might overcome the need for ex vivo generation and antigen loading of DC. Finally, future perspectives towards the use of lentiviral vectors in cancer immunotherapy are presented.

Sphingosine-kinase and sphingosine-1-phosphate regulate migration of immature dendritic cells

Introduction Currently, dendritic cells (DC) are tested as vectors for cancer immunotherapy. However, little is known about the mechanisms regulating DC migration. Sphingosinekinase (SphK) and its catalytic product Sphingosine-1phosphate (S1P) play a central role in processes such as cellular differentiation, survival and migration, which are often disregulated in cancer. Here, we examined the role of SphK and S1P in migration of DC.

Disabled infectious single-cycle herpes simplex virus as an oncolytic vector for immunotherapy of colorectal cancer.

New modalities of treatment for colorectal cancer are required to support and improve those currently available. One such approach is immunotherapy by transfer of immunostimulatory genes to tumor cells. Here, we report the use of a herpes simplex virus (HSV) vector that is capable of a single round of infection (disabled infectious single-cycle [DISC]-HSV) as a gene transfer vehicle for colorectal cancer. This vector has potential advantages over other vectors for cancer immunotherapy in that it lyses infected tumor cells. Infection with DISC-HSV inhibited tumor cell growth both in vitro and in vivo. In addition, DISC-HSV-mediated cell killing occurs by both apoptotic and necrotic mechanisms. A range of colorectal tumor cell lines could be rapidly transduced with DISC-HSV/lacZ (14-90% in 4 hr). Both tumor prevention and tumor therapy protocols showed clear antitumor effects with DISC-HSV/mGM-CSF. In the prophylactic approach, an infected/irradiated whole cell vaccine protected up to 80% of mice from rechallenge. In addition, intratumoral injection of established tumors with DISC-HSV/GM-CSF caused rejection in 40% of mice and generated some protection from subsequent rechallenge. In both cases, however, it is clear that a dominant therapeutic effect of the DISC-HSV vector derives from its oncolytic properties, irrespective of the transduced gene. As a prelude to taking these studies forward to human clinical trials, we demonstrate that tumor cells could be successfully grown from freshly obtained human colorectal cancer resections (within 1 week of surgery), were transduced with DISC-HSV/hGM-CSF, and secreted the cytokine. This study provides the preclinical basis for trials of immunotherapy of colorectal cancer using DISC-HSV.