Abstract: Conventional dendritic cells (cDC) are ex vivo differentiated professional antigen presenting cells capable of potently stimulating naïve T cells and with vast potential for immunotherapeutic applications. The manufacture of clinical-grade cDC is relatively complex and requires several days for completion. Clinical trials showed poor trafficking of cDC from subcutaneous injection sites to lymph nodes (LN), where DC can optimally stimulate naïve lymphocytes for long-lasting memory responses. We demonstrated in mouse and human systems that a single overnight ex vivo lentiviral (LV) gene transfer into DC precursors for production of combination of cytokines and antigens was capable to induce autonomous self-differentiation of antigen-loaded DC in vitro and in vivo. These highly viable induced DC (iDC) effectively migrated from the injected skin to LN, where they effectively activated de novo antigen-specific effector memory T cells. Two iDC modalities were validated in relevant animal models and are now in clinical development: Self-differentiated Myeloid-derived Antigen-presenting-cells Reactive against Tumors co-expressing GM-CSF/IL-4/TRP2 for melanoma immunotherapy in the autologous setting (SmartDCtrp2), and Self-differentiated Myeloid-derived Lentivirus-induced against human cytomegalovirus as an allogeneic matched adoptive cell after stem cell transplantation (SmyleDCpp65). The lentiviral vector design and packaging methodology has “evolved” continuously in order to simplify and optimize function and biosafety of in vitro and in vivo genetic reprogramming of iDC. Here, we address the challenges seeking for new creations of genetically programmed iDC and integrase-defective LV vaccines for immune regeneration.
Abstract: The immune system has acquired increasing importance as a key player in cancer maintenance and growth. Thus, modulating anti-tumor immune mediators has become an attractive strategy for cancer treatment. Toll-like receptors (TLRs) have gradually emerged as potential targets of newer immunotherapies. TLR-9 is preferentially expressed on endosome membranes of B-cells and plasmacytoid dendritic cells (pDC) and is known for its ability to stimulate specific immune reactions through the activation of inflammation-like innate responses. Several synthetic CpG oligonucleotides (ODNs) have been developed as TLR-9 agonists with the aim of enhancing cancer immune surveillance. In many preclinical models, CpG ODNs were found to suppress tumor growth and proliferation both in monotherapy and in addition to chemotherapies or target therapies. TLR-9 agonists have been also tested in several clinical trials in patients with solid tumors. These agents showed good tolerability and usually met activity endpoints in early phase trials. However, they have not yet been demonstrated to significantly impact survival, neither as single agent treatments, nor in combination with chemotherapies or cancer vaccines. Further investigations in larger prospective studies are required.
Abstract: Integration of viral vectors into a host genome is associated with insertional mutagenesis and subjects in clinical gene therapy trials must be monitored for this adverse event. Several PCR based methods such as ligase-mediated (LM) PCR, linear-amplification-mediated (LAM) PCR and non-restrictive (nr) LAM PCR were developed to identify sites of vector integration. Coupling the power of next-generation sequencing technologies with various PCR approaches will provide a comprehensive and genome-wide profiling of insertion sites and increase throughput. In this bioinformatics study, we aimed to develop and apply quality metrics to viral insertion data obtained using next-generation sequencing. We developed five simple metrics for assessing next-generation sequencing data from different PCR products and showed how the metrics can be used to objectively compare runs performed with the same methodology as well as data generated using different PCR techniques. The results will help researchers troubleshoot complex methodologies, understand the quality of sequencing data, and provide a starting point for developing standardization of vector insertion site data analysis.
Abstract: Oncolytic virotherapy has advanced since the days of its conception but therapeutic efficacy in the clinics does not seem to reach the same level as in animal models. One reason is premature oncolytic virus clearance in humans, which is a reasonable assumption considering the immune-stimulating nature of the oncolytic agents. However, several studies are beginning to reveal layers of restriction to oncolytic virotherapy that are present before an adaptive neutralizing immune response. Some of these barriers are present constitutively halting infection before it even begins, whereas others are raised by minute cues triggered by virus infection. Indeed, we and others have noticed that delivering viruses to tumors may not be the biggest obstacle to successful therapy, but instead the physical make-up of the tumor and its capacity to mount antiviral defenses seem to be the most important efficacy determinants. In this review, we summarize the constitutive and innate barriers to oncolytic virotherapy and discuss strategies to overcome them.
Abstract: Cancer has been, from the beginning, a target of intense research for gene therapy approaches. Currently, more than 60% of all on-going clinical gene therapy trials worldwide are targeting cancer. Indeed, there is a clear unmet medical need for novel therapies. This is further urged by the fact that current conventional cancer therapies are frequently troubled by their toxicities. Different gene therapy strategies have been employed for cancer, such as pro-drug activating suicide gene therapy, anti-angiogenic gene therapy, oncolytic virotherapy, gene therapy-based immune modulation, correction/compensation of gene defects, genetic manipulation of apoptotic and tumor invasion pathways, antisense, and RNAi strategies. Cancer types, which have been targeted with gene therapy, include brain, lung, breast, pancreatic, liver, colorectal, prostate, bladder, head and neck, skin, ovarian, and renal cancer. Currently, two cancer gene therapy products have received market approval, both of which are in China. In addition, the stimulation of the host’s immune system, using gene therapeutic approaches, has gained vast interest. The intention of this review is to point out the most commonly viral and non-viral vectors and methods used in cancer gene therapy, as well as highlight some key results achieved in clinical trials.
Abstract: Helper-dependent adenoviral (HDAd) vectors that are devoid of all viral coding sequences are promising non-integrating vectors for gene therapy because they efficiently transduce a variety of cell types in vivo, have a large cloning capacity, and drive long-term transgene expression without chronic toxicity. The main obstacle preventing clinical applications of HDAd vectors is the host innate inflammatory response against the vector capsid proteins that occurs shortly after intravascular vector administration and result in acute toxicity, the severity of which is dose dependent. Intense efforts have been focused on elucidating adenoviral vector–host interactions and the factors involved in the acute toxicity. This review focuses on the recent acquisition of data on such interactions and on strategies investigated to improve the therapeutic index of HDAd vectors.