Search

Adenoviral vectors are commonly used in clinical gene therapy. Apart from oncolytic adenoviruses, vector replication is highly undesired as it may pose a safety risk for the treated patient. Thus, careful monitoring for the formation of replication-competent adenoviruses (RCA) during vector manufacturing is required. To render adenoviruses replication deficient, their genomic E1 region is deleted. However, it has been known for a long time that during their propagation, some viruses will regain their replication capability by recombination in production cells, most commonly HEK293. Recently developed RCA assays have revealed that many clinical batches contain more RCA than previously assumed and allowed by regulatory authorities. The clinical significance of the higher RCA content has yet to be thoroughly evaluated. In this review, we summarize the biology of adenovirus vectors, their manufacturing methods, and the origins of RCA formed during HEK293-based vector production. Lastly, we share our experience using minimally RCA-positive serotype 5 adenoviral vectors based on observations from our clinical cardiovascular gene therapy studies. [ABSTRACT FROM AUTHOR]

Immune checkpoint inhibitors have clinical success in prolonging the life of many cancer patients. However, only a minority of patients benefit from such therapy, calling for further improvements. Currently, most PD-L1 checkpoint inhibitors in the clinic do not elicit Fc effector mechanisms that would substantially increase their efficacy. To gain potency and circumvent off-target effects, we previously designed an oncolytic adenovirus (Ad-Cab) expressing an Fc fusion peptide against PD-L1 on a cross-hybrid immunoglobulin GA (IgGA) Fc. Ad-Cab elicited antibody effector mechanisms of IgG1 and IgA, which led to higher tumor killing compared with each isotype alone and with clinically approved PD-L1 checkpoint inhibitors. In this study, we further improved the therapy to increase the IgG1 Fc effector mechanisms of the IgGA Fc fusion peptide (Ad-Cab FT) by adding four somatic mutations that increase natural killer (NK) cell activation. Ad-Cab FT was shown to work better at lower concentrations compared with Ad-Cab in vitro and in vivo and to have better tumor- and myeloid-derived suppressor cell killing, likely because of higher NK cell activation. Additionally, the biodistribution of the Fc fusion peptide demonstrated targeted release in the tumor microenvironment with minimal or no leakage to the peripheral blood and organs in mice. These data demonstrate effective and safe use of Ad-Cab FT, bidding for further clinical investigation.

Oncolytic viruses hold great promise for cancer treatment but their practical applications are seriously impaired by a series of limitations. Herein, an engineered oncolytic adenovirus (OA) is constructed that can boost both the direct oncolysis and antitumor immune response of OA attributed to the increased tumor targeting and low‐pH responsive fusogenic activity. The tumor cell membranes are decorated with vesicular stomatitis virus glycoprotein (VSVG) via vesicular stomatitis virus (VSV) infection and then used to mask OA (V‐M@OA). After systemic administration, the engineered OA can target homologous tumors owing to the homing ability of tumor membranes. Then the unique low‐pH responsive fusogenic activity of VSVG significantly enhances the replication of OA by promoting the whole virus infection process, resulting in remarkable virus‐mediated tumoricidal effects and thus abundant in situ released tumor‐associated antigens (TAAs). Meanwhile, VSVG on V‐M@OA augments the adjuvanticity of OA and thus significantly enhancing the antitumor immune response. The synergism of virus‐mediated killing and immune effects leads to significant tumor inhibition with no obvious side effects. [ABSTRACT FROM AUTHOR]

Adenoviral vectors are commonly used in clinical gene therapy. Apart from oncolytic adenoviruses, vector replication is highly undesired as it may pose a safety risk for the treated patient. Thus, careful monitoring for the formation of replication-competent adenoviruses (RCA) during vector manufacturing is required. To render adenoviruses replication deficient, their genomic E1 region is deleted. However, it has been known for a long time that during their propagation, some viruses will regain their replication capability by recombination in production cells, most commonly HEK293. Recently developed RCA assays have revealed that many clinical batches contain more RCA than previously assumed and allowed by regulatory authorities. The clinical significance of the higher RCA content has yet to be thoroughly evaluated. In this review, we summarize the biology of adenovirus vectors, their manufacturing methods, and the origins of RCA formed during HEK293-based vector production. Lastly, we share our experience using minimally RCA-positive serotype 5 adenoviral vectors based on observations from our clinical cardiovascular gene therapy studies.

Simple Summary: Immunotherapy has revolutionized cancer treatment, as demonstrated by the tremendous success of checkpoint inhibitors in different tumor types. Unfortunately, most patients, particularly patients with non-responsive "cold" tumors, do not benefit from checkpoint inhibitors. Enter "armed" oncolytic viruses, which "cooperate" with checkpoint inhibitors to improve anticancer responses. These are genetically engineered viruses that selectively infect, replicate in, and kill cancer cells but not cells from healthy tissues; in the process, oncolytic viruses express the therapeutic proteins that they are armed with or carry. This effectively turns the infected tumors "hot" and makes them suitable for treatment with checkpoint inhibitors. The most well-studied of all the oncolytic viruses are adenoviruses. These are agents of the common cold, which makes them remarkably safe for clinical use. This review article summarizes the oncolytic adenoviruses in advanced clinical trials and presents strategies to improve their anticancer activity. Oncolytic viruses, colloquially referred to as "living drugs", amplify themselves and the therapeutic transgenes that they carry to stimulate an immune response both locally and systemically. Remarkable exceptions aside, such as the recent 14-patient trial with the PD-1 inhibitor, dostarlimab, in mismatch repair (MMR) deficient rectal cancer, where the complete response rate was 100%, checkpoint inhibitors are not cure-alls, which suggests the need for a combination partner like oncolytic viruses to prime and augment their activity. This review focuses on adenoviruses, the most clinically investigated of all the oncolytic viruses. It covers specific design features of clinical adenoviral candidates and highlights their potential both alone and in combination with checkpoint inhibitors in clinical trials to turn immunologically "cold" and unresponsive tumors into "hotter" and more responsive ones through a domino effect. Finally, a "mix-and-match" combination of therapies based on the paradigm of the cancer-immunity cycle is proposed to augment the immune responses of oncolytic adenoviruses. [ABSTRACT FROM AUTHOR]

Malignant mesothelioma is a rare and aggressive cancer that develops in the thin layer surrounding the mesothelium and is mainly caused by asbestos exposure. Despite improvements in patient prognosis with conventional cancer treatments, such as surgery, chemotherapy, and radiotherapy, there are still no curative treatment modalities for advanced disease. In recent years, new therapeutic avenues have been explored. Improved understanding of the mechanisms underlying the dynamic tumor interaction with the immune system has led to the development of immunotherapeutic approaches. Numerous recent clinical trials have shown a desire to develop more effective treatments that can be used to fight against the disease. Immune checkpoint inhibitors, oncolytic adenoviruses, and their combination represent a promising strategy that can be used to synergistically overcome immunosuppression in the mesothelioma tumor microenvironment. This review provides a synthesized overview of the current state of knowledge on new therapeutic options for mesothelioma with a focus on the results of clinical trials conducted in the field. [ABSTRACT FROM AUTHOR]

Intratumoral injection of various effector cells combined with oncolytic adenovirus expressing antitumor cytokines exert an effective antitumor immune effect by oncolysis and altering the tumor microenvironment. However, this combination therapy had certain limitations. When used in high concentrations, effector cells and oncolytic viruses can spread rapidly to surrounding non-target tissues. And because both therapies used in combination are immunogenic and exhibit shorter biological activity, multiple injections were required to attain an adequate therapeutic index. To overcome these drawbacks, we encapsulated gelatin-based hydrogel capable of co-deliver oncolytic adenovirus armed with IL12 and IL15 (CRAd-IL12-IL15) and CIK cells for enhancing and prolonging the antitumor effects of both therapies after a single intratumoral injection. The injectable and biodegradable hydrogel reduced the dispersion of high-dose oncolytic adenovirus and CIK cells from the injection site to the liver and other non-target tissues. In this study, a novel oncolytic adenoviral vector CRAd-IL12-IL15 was constructed to verify the cytokine expression and oncolytic ability, which can upregulate the expression levels of Bcl-2, Cish and Gzmb in tumor cells. The CRAd-IL12-IL15 + CIKs/gelatin treatment maintained sustained release of CRAd-IL12-IL15 and active CIK cells over a longer period of time, attenuating the antiviral immune response against adenovirus. In conclusion, the results suggested that hydrogel-mediated co-delivery of CRAd-IL12-IL15 and CIK cells might be a an approach to overcome limitations. Both treatments could be effectively retained in tumor tissue and sustained to induce potent anti-tumor immune responses with a single administration.

Malignant mesothelioma is a rare and aggressive cancer that develops in the thin layer surrounding the mesothelium and is mainly caused by asbestos exposure. Despite improvements in patient prognosis with conventional cancer treatments, such as surgery, chemotherapy, and radiotherapy, there are still no curative treatment modalities for advanced disease. In recent years, new therapeutic avenues have been explored. Improved understanding of the mechanisms underlying the dynamic tumor interaction with the immune system has led to the development of immunotherapeutic approaches. Numerous recent clinical trials have shown a desire to develop more effective treatments that can be used to fight against the disease. Immune checkpoint inhibitors, oncolytic adenoviruses, and their combination represent a promising strategy that can be used to synergistically overcome immunosuppression in the mesothelioma tumor microenvironment. This review provides a synthesized overview of the current state of knowledge on new therapeutic options for mesothelioma with a focus on the results of clinical trials conducted in the field.

Oncolytic virotherapy is an emerging therapeutic approach based on replication-competent viruses able to selectively infect and destroy cancer cells, inducing the release of tumor-associated antigens and thereby recruiting immune cells with a subsequent increase in antitumoral immune response. To increase the anticancer activity, we engineered a specific oncolytic adenovirus expressing a single-chain variable fragment of an antibody against PD-L1 to combine blockage of PD-1/PD-L1 interaction with the antitumoral activity of Onc.Ad5. To assess its efficacy, we infected B16.OVA cells, a murine model of melanoma, with Ad5Δ24 -anti-PD-L1-scFv and then co-cultured them with C57BL/6J naïve splenocytes. We observed that the combinatorial treatments were significantly more effective in inducing cancer cell death. Furthermore, we assessed the efficacy of intratumoral administrations of Ad5Δ24-anti-PD-L1-scFv in C57BL/6J mice engrafted with B16.OVA and compared this treatment to that of the parental Ad5Δ24 or placebo. Treatment with the scFv-expressing Onc.Ad induced a marked reduction of tumor growth concerning the parental Onc.Ad. Additionally, the evaluation of the lymphocytic population infiltrating the treated tumor reveals a favorable immune profile with an enhancement of the CD8+ population. These data suggest that Onc.Ad-mediated expression of immune checkpoint inhibitors increases oncolytic virotherapy efficacy and could be an effective and promising tool for cancer treatments, opening a new way into cancer therapy. [ABSTRACT FROM AUTHOR]

Oncolytic virotherapy is an emerging therapeutic approach based on replication-competent viruses able to selectively infect and destroy cancer cells, inducing the release of tumor-associated antigens and thereby recruiting immune cells with a subsequent increase in antitumoral immune response. To increase the anticancer activity, we engineered a specific oncolytic adenovirus expressing a single-chain variable fragment of an antibody against PD-L1 to combine blockage of PD-1/PD-L1 interaction with the antitumoral activity of Onc.Ad5. To assess its efficacy, we infected B16.OVA cells, a murine model of melanoma, with Ad5Δ24 -anti-PD-L1-scFv and then co-cultured them with C57BL/6J naïve splenocytes. We observed that the combinatorial treatments were significantly more effective in inducing cancer cell death. Furthermore, we assessed the efficacy of intratumoral administrations of Ad5Δ24-anti-PD-L1-scFv in C57BL/6J mice engrafted with B16.OVA and compared this treatment to that of the parental Ad5Δ24 or placebo. Treatment with the scFv-expressing Onc.Ad induced a marked reduction of tumor growth concerning the parental Onc.Ad. Additionally, the evaluation of the lymphocytic population infiltrating the treated tumor reveals a favorable immune profile with an enhancement of the CD8+ population. These data suggest that Onc.Ad-mediated expression of immune checkpoint inhibitors increases oncolytic virotherapy efficacy and could be an effective and promising tool for cancer treatments, opening a new way into cancer therapy.

Abstract Background Conditionally replicative oncolytic adenoviruses (CRAds) display significant anti-tumor effects. However, the traditional adenovirus of serotype 5 (Ad5) entering cancer cells via coxsackie virus and adenovirus receptor (CAR) can’t be utilized for bladder cancer with low expression of CAR, which limits the application of Ad5. Methods We utilized Ad5/F11p containing the chimeric fiber gene encoding the Ad5 fiber tail domain and Ad11p fiber shaft and knob domains to construct bladder cancer-specific chimeric type viruses Ad5/F11p-PSCAE-UPII-E1A, which can infect bladder cancer cells mediated by CD46 molecule. We carried out series of experiments in vitro to research anti-tumor effect of Ad5/F11p-PSCAE-UPII-E1A and the interaction in combination with cisplatin. Results The results demonstrated Ad5/F11p-PSCAE-UPII-E1A could infect bladder cancer cells (T24, EJ and 5637) in a CAR-independent way, and exert anti-tumor effect by blocking the cancer cells in G1 phase and inducing apoptosis. Ad5/F11p-PSCAE-UPII-E1A plus cisplatin enhanced the anti-proliferative effect and increased the number of apoptotic cells compared with viruses or cisplatin alone. Ad5/F11p-PSCAE-UPII-E1A plus cisplatin could upregulate the proteins expression of p53, Bax, and cleaved caspase-3, and downregulated Bcl-2 protein expression in T24, EJ and 5637 cells. Conclusion We constructed a bladder cancer-specific oncolytic adenovirus and provided new combination treatment strategies for bladder cancer.

Abstract Extracellular vesicles (EVs), are naturally occurring cargo delivery tools with the potential to be used as drug vehicles of single agents or combination therapies. We previously demonstrated that human lung cancer cell-derived EVs could be used for the systemic delivery of oncolytic virus (OVs) and chemotherapy drugs such as paclitaxel (PTX), leading to enhanced anti-tumor effects in nude mice. In the current work, we evaluated the biodistribution of EVs by using bioluminescence and fluorescence imaging technologies, thus proving the ability of these EVs-formulations to specifically target the neoplasia, while leaving other body tissues unaffected. Moreover, in vivo imaging of NFκB activation in an immunocompetent reporter mouse model allowed to demonstrate the selective ability of EVs to induce tumor-associated inflammatory reactions, which are characterized by immunogenic cell death and CD3+/CD4+/CD8+ T-cell infiltration. While EVs have the potential to induce a systemic immune reaction by pro-inflammatory cytokines, our study provides compelling evidences of a localized inflammatory effect in the peritumoral area. Collectively, our findings strongly support the systemic administration of EVs formulations with OVs alone or in combination with chemotherapy agents as a novel strategy aimed at treating primary and metastatic cancers. Graphical abstract Unlabelled Image [ABSTRACT FROM AUTHOR]

Genetic replacement of adenoviral fiber knobs by ligands that enable tumor specific targeting of oncolytic adenoviruses is challenging because the fiber knob contributes to virus assembly. Here, we present a novel concept by describing stable recombinant adenoviruses with tumor specific infection mode. The fiber knob was replaced by endosialidaseNF (endoNF), the tailspike protein of bacteriophage K1F. EndoNF recognizes polysialic acid, an oncofetal antigen characteristic for high malignant tumors of neuroendocrine origin. An intramolecular chaperone contained in endoNF warrants folding and compensates for the knob function in virus assembly. Obtained recombinant viruses demonstrated polysialic acid dependent infection modes, strong oncolytic capacity with polysialic acid positive cells in culture and a high potential to inhibit tumor growth in a therapeutic mouse model of subcutaneous neuroblastoma. With a single genetic manipulation we achieved ablation of the fiber knob, introduction of a tumor specific ligand, and folding control over the chimeric fiber construct. [ABSTRACT FROM AUTHOR]

Oncolytic virotherapy has been an attractive drug platform for targeted therapy of cancer over the past few years. Viral vectors can be used to target and lyse cancer cells, but achieving good efficacy and specificity with this treatment approach is a major challenge. Here, we assessed the ability of a novel dual-specific anti-tumor oncolytic adenovirus, expressing the hemagglutinin-neuraminidase (HN) gene from the Newcastle disease virus under the human telomerase reverse transcriptase (hTERT) promoter (Ad-hTERTp-E1a-HN), to inhibit esophageal cancer EC-109 cells in culture and to reduce tumor burden in xenografted BALB/c nude mice. In vitro, infection with Ad-hTERT-E1a-HN could inhibit the growth of EC-109 cells significantly and also protect normal human liver cell line L02 from growth suppression in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. Ad-hTERT-E1a-HN also effectively and selectively decreased the sialic acid level on EC-109 cells, but not on L02 cells. Furthermore, Ad-hTERT-E1a-HN was shown to induce the apoptosis pathway via acridine orange and ethidium bromide staining (AO/EB staining), increase reactive oxygen species (ROS), reduce mitochondrial membrane potential and release cytochrome c. In vivo, xenografted BALB/c nude mice were treated via intratumoral or intravenous injections of Ad-hTERT-E1a-HN. Although both treatments showed an obvious suppression in tumor volume, only Ad-hTERT-E1a-HN delivered via intratumoral injection elicited a complete response to treatment. These results reinforced previous findings and highlighted the potential therapeutic application of Ad-hTERT-E1a-HN for treatment of esophageal cancer in clinical trials.

Adoptive immunotherapy and targeted gene therapy have been extensively used to eliminate tumor cells. The combination treatment is capable of efficiently generating an effective antitumor immune response and disrupting tumor-induced tolerance. Moreover, effective antitumor immune responses are dependent on coordinate interaction among various effector cells. This study focused on whether the combination of cytotoxic effector cell-based adoptive immunotherapy and CCL20/IL15-armed oncolytic adenoviruses could induce enhanced antitumor activity. The CCL20/IL15-armed oncolytic adenovirus was constructed using homologous recombination with shuttle plasmids and full-length Ad backbones. We chose the telomerase reverse transcriptase promoter (TERTp) to replace the E1A promoter to drive the oncolytic adenoviral E1A gene. Thus, this CRAd-CCL20-IL15 could induce apoptosis in TERTp-positive tumor cells due to viral propagation, but these viruses could not replicate efficiently in normal cells. The combination of cytotoxic effector cells and CRAd-CCL20-IL15 showed greater antitumor potential than that of cytotoxic effector cells or CRAd-CCL20-IL15 alone. Moreover, the combined treatment could induce tumor-specific cytotoxicity of CTLs in vitro. Further analysis demonstrated that this combined treatment resulted in significant tumor regression in mouse models. This study has provided preclinical evidence that combined treatment with cytotoxic effector cells and CRAd-CCL20-IL15 may offer alternative treatment options for tumor therapy. [ABSTRACT FROM AUTHOR]

The effective antitumor immune responses are dependent on coordinate interaction of various effector cells. Thus, the combination of adoptive immunotherapy and target gene therapy is capable of efficiently generating a productive antitumor immune response. We investigated whether combination of cytokine-induced killer (CIK) cells adoptive immunotherapy and CCL21/IL15 armed oncolytic adenovirus could induce the enhanced antitumor activity. The CCL21/IL15 co-expression oncolytic adenoviruses were constructed by using the AdEasy system, which uses homologous recombination with shuttle plasmids and full length Ad backbones. This conditionally replicating adenoviruses CRAd-CCL21-IL15 could induce apoptosis in TERTp-positive tumor cells for viral propagation, but do not replicate efficiently in normal cells, because the E1A promoter was replaced by telomerase reverse transcriptase promoter (TERTp). Our results showed that the combination of CIK cells and CRAd-CCL21-IL15 could induce higher antitumor activity than either CIK cells or CRAd-CCL21-IL15 alone. This combined treatment could induce the tumor specific cytotoxicity of CTLs (cytotoxic T lymphocytes) in vitro. Moreover, the treatment of established tumors with the combined therapy of CIK cells and CRAd-CCL21-IL15 resulted in tumor regression. This study suggests that the combined treatment by adoptive immunotherapy and gene therapy is a promising strategy for the therapy of tumor. [ABSTRACT FROM AUTHOR]

Multigene-armed oncolytic adenoviruses are capable of efficiently generating a productive antitumor immune response. The chemokine (C-C motif) ligand 21 (CCL21) binds to CCR7 on naïve T cells and dendritic cells (DCs) to promote their chemoattraction to the tumor and resultant antitumor activity. Interleukin 21 (IL21) promotes survival of naïve T cells while maintaining their CCR7 surface expression, which increases their capacity to transmigrate in response to CCL21 chemoattraction. IL21 is also involved in NK cell differentiation and B cell activation and proliferation. The generation of effective antitumor immune responses is a complex process dependent upon coordinated interactions of various subsets of effector cells. Using the AdEasy system, we aimed to construct an oncolytic adenovirus co-expressing CCL21 and IL21 that could selectively replicate in TERTp-positive tumor cells (Ad-CCL21-IL21 virus). The E1A promoter of these oncolytic adenoviruses was replaced by telomerase reverse transcriptase promoter (TERTp). Ad-CCL21-IL21 was constructed from three plasmids, pGTE-IL21, pShuttle-CMV-CCL21 and AdEasy-1 and was homologously recombined and propagated in the Escherichia coli strain BJ5183 and the packaging cell line HEK-293, respectively. Our results showed that our targeted and armed oncolytic adenoviruses Ad-CCL21-IL21 can induce apoptosis in TERTp-positive tumor cells to give rise to viral propagation, in a dose-dependent manner. Importantly, we confirm that these modified oncolytic adenoviruses do not replicate efficiently in normal cells even under high viral loads. Additionally, we investigate the role of Ad-CCL21-IL21 in inducing antitumor activity and tumor specific cytotoxicity of CTLs in vitro. This study suggests that Ad-CCL21-IL21 is a promising targeted tumor-specific oncolytic adenovirus. [ABSTRACT FROM AUTHOR]

Background: Gene-based virotherapy mediated by oncolytic viruses is currently experiencing a renaissance in cancer therapy. However, relatively little attention has been given to the potentiality of dual gene virotherapy strategy as a novel therapeutic approach to mediate triplex anticancer combination effects, particularly if the two suitable genes are well chosen. Both tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and interleukin-12 (IL-12) have been emerged as promising pharmacological candidates in cancer therapy; however, the combined efficacy of TRAIL and IL-12 genes for treatment of human hepatocellular carcinoma (HCC) remains to be determined. Methods: Herein, we investigated the therapeutic efficacy of concurrent therapy with two armed oncolytic adenoviruses encoding human TRAIL gene (Ad-ΔB/TRAIL) and IL-12 gene (Ad-ΔB/IL-12), respectively, on preclinical models of human HCC, and also elucidated the possible underlying mechanisms. The effects of Ad-ΔB/TRAIL+Ad-ΔB/ IL-12 combination therapy were assessed both in vitro on Hep3B and HuH7 human HCC cell lines and in vivo on HCC-orthotopic model established in the livers of athymic nude mice by intrahepatic implantation of human Hep3B cells. Results: Compared to therapy with non-armed control Ad-ΔB, combined therapy with Ad-ΔB/TRAIL+Ad-ΔB/IL-12 elicited profound anti-HCC killing effects on Hep3B and HuH7 cells and on the transplanted Hep3B-orthotopic model. Efficient viral replication and TRAIL and IL-12 expression were also confirmed in HCC cells and the harvested tumor tissues treated with this combination therapy. Mechanistically, co-therapy with Ad-ΔB/TRAIL+Ad-ΔB/IL-12 exhibited an enhanced effect on apoptosis promotion, activation of caspase-3 and-8, generation of anti-tumor immune response evidenced by upregulation of interferon gamma (IFN-γ) production and infiltration of natural killer-and antigen presenting cells, and remarkable repression of intratumor vascular endothelial growth factor (VEGF) and cluster of differentiation 31 (CD31) expression and tumor microvessel density. Conclusions: Overall, our data showed a favorable therapeutic effect of Ad-ΔB/TRAIL+Ad-ΔB/IL-12 combination therapy against human HCC, and may therefore constitute a promising and effective therapeutic strategy for treating human HCC. However, further studies are warranted for its reliable clinical translation. [ABSTRACT FROM AUTHOR]

Immune checkpoint inhibitors have clinical success in prolonging the life of many cancer patients. However, only a minority of patients benefit from such therapy, calling for further improvements. Currently, most PD-L1 checkpoint inhibitors in the clinic do not elicit Fc effector mechanisms that would substantially increase their efficacy. To gain potency and circumvent off-target effects, we previously designed an oncolytic adenovirus (Ad-Cab) expressing an Fc fusion peptide against PD-L1 on a cross-hybrid immunoglobulin GA (IgGA) Fc. Ad-Cab elicited antibody effector mechanisms of IgG1 and IgA, which led to higher tumor killing compared with each isotype alone and with clinically approved PD-L1 checkpoint inhibitors. In this study, we further improved the therapy to increase the IgG1 Fc effector mechanisms of the IgGA Fc fusion peptide (Ad-Cab FT) by adding four somatic mutations that increase natural killer (NK) cell activation. Ad-Cab FT was shown to work better at lower concentrations compared with Ad-Cab in vitro and in vivo and to have better tumor- and myeloid-derived suppressor cell killing, likely because of higher NK cell activation. Additionally, the biodistribution of the Fc fusion peptide demonstrated targeted release in the tumor microenvironment with minimal or no leakage to the peripheral blood and organs in mice. These data demonstrate effective and safe use of Ad-Cab FT, bidding for further clinical investigation., Competing Interests: V.C. is a co-founder and shareholder of Valo Therapeutics LTD (not related to this study)., (© 2023.)

Renal cell carcinoma is the most frequent kidney malignancy and patients with metastatic disease have a poor prognosis. Suppressed apoptosis and marked invasiveness are distinctive features of renal cell carcinoma. In the present study, a dual-regulated oncolytic adenovirus expressing the interluekin (IL)-24 gene (Ki67-ZD55-IL-24) was constructed utilizing the Ki67 promoter to replace the native viral promoter of the E1A gene. Whether the combination of Ki67-ZD55-IL-24-mediated gene virotherapy and radiotherapy produced increased cytotoxicity in renal cell carcinoma cells via mitochondrial apoptotic cell death was investigated. The data indicated that this novel strategy has the potential to be further developed into an effective approach to treat renal cell carcinoma. The results showed that the combination of Ki67-ZD55-IL-24 and radiotherapy significantly enhanced anti-tumour activity via increasing the induction of apoptosis in melanoma cells compared with the other agents. [ABSTRACT FROM AUTHOR]

Intratumoral injection of various effector cells combined with oncolytic adenovirus expressing antitumor cytokines exert an effective antitumor immune effect by oncolysis and altering the tumor microenvironment. However, this combination therapy had certain limitations. When used in high concentrations, effector cells and oncolytic viruses can spread rapidly to surrounding non-target tissues. And because both therapies used in combination are immunogenic and exhibit shorter biological activity, multiple injections were required to attain an adequate therapeutic index. To overcome these drawbacks, we encapsulated gelatin-based hydrogel capable of co-deliver oncolytic adenovirus armed with IL12 and IL15 (CRAd-IL12-IL15) and CIK cells for enhancing and prolonging the antitumor effects of both therapies after a single intratumoral injection. The injectable and biodegradable hydrogel reduced the dispersion of high-dose oncolytic adenovirus and CIK cells from the injection site to the liver and other non-target tissues. In this study, a novel oncolytic adenoviral vector CRAd-IL12-IL15 was constructed to verify the cytokine expression and oncolytic ability, which can upregulate the expression levels of Bcl-2, Cish and Gzmb in tumor cells. The CRAd-IL12-IL15 + CIKs/gelatin treatment maintained sustained release of CRAd-IL12-IL15 and active CIK cells over a longer period of time, attenuating the antiviral immune response against adenovirus. In conclusion, the results suggested that hydrogel-mediated co-delivery of CRAd-IL12-IL15 and CIK cells might be a an approach to overcome limitations. Both treatments could be effectively retained in tumor tissue and sustained to induce potent anti-tumor immune responses with a single administration. [Display omitted] • We constructed a IL12 and IL15 co-expression conditionally replicating oncolytic adenoviruses (CRAd-IL12-IL15). • We encapsulated gelatin-based hydrogel capable of co-deliver CRAd-IL12-IL15 and CIK cells. • The injectable and biodegradable hydrogel reduced the dispersion of high-dose oncolytic adenovirus and CIK cells. [ABSTRACT FROM AUTHOR]

Abstract: Oncolytic adenoviruses have been used in different preclinical and clinical studies, showing their capacity to kill tumor cells without major adverse events. However, these studies also underline the limitations of this approach. The efficacy of oncolytic adenoviruses is hampered by their limited ability to transduce some tumor types, their lack of selectivity, and their poor dissemination within tumors. In addition, the host immune response may limit oncolytic adenovirus efficacy. Combining oncolytic adenoviruses with chemotherapeutics constitutes an appealing strategy to increase their potency. The first part of this review describes the molecular basis of oncolytic adenoviruses, their use in preclinical studies and clinical trials, their limitations, and strategies to circumvent these limitations. The second part will focus on studies combining oncolytic adenoviruses with chemotherapeutic drugs, including standard chemotherapeutic drugs, molecularly targeted drugs, and other drugs that have been combined with oncolytic adenoviruses. Finally, based on these studies, we describe future directions and general rules that could be followed to identify chemotherapeutic drugs displaying additive/synergistic effects when combined with oncolytic adenoviruses. [Copyright &y& Elsevier]

Introduction: Oncolytic adenoviruses (Ads) are promising therapeutics to enhance anti-tumor immune responses and modulate the immune-suppressive tumor microenvironment (TME). Due to their potent ability to deliver genes in vivo , oncolytic Ads have been armed with a variety of immune stimulatory payloads to boost tumor immunotherapy., Areas Covered: We describe current knowledge about engineering oncolytic Ads to insert transgene payloads, including methods to increase its genome capacity for transgene insertion. We also review several categories of immune stimulatory payloads that have been used in oncolytic Ads to combat different barriers to effective immunotherapy., Expert Opinion: We anticipate that multi-armed oncolytic Ads alone or in combination with other types of immunotherapies will greatly improve the efficacy of oncolytic virotherapy in the future by targeting several immune barriers. However, the production and testing of multiple payload-armed Ads can be complex and time-consuming due to the limitations of current tools. Given this, we should develop new tools for rapid construction of armed Ads, improve animal models or new systems to compare the efficacy of multiple payloads, and carefully monitor the immunological toxicity induced by payloads or by systemic delivery. Most importantly, we should pursue payload-arming approaches with great care to ensure safety.

Malignant melanoma is one of the most lethal and aggressive human malignancies. Suppressed apoptosis and extraordinary invasiveness are the distinctive features that contribute to malignant melanoma. The alkylating agent temozolomide (TMZ) is one of the most effective single chemotherapeutic agents for patients with malignant melanoma, but resistance develops quickly and with high frequency. We constructed a dual-regulated oncolytic adenovirus expressing interleukin 24 (IL-24) gene (Ki67-ZD55-IL-24) by utilizing the Ki67 promoter to replace the native viral promoter of E1A gene. We investigated whether a combination of Ki67-ZD55-IL-24-mediated gene virotherapy and chemotherapy using TMZ produces increased cytotoxicity against human melanoma cells via the induction of apoptosis. Our data indicate that this novel strategy thus holds promising potentials for further developing an effective approach to treat malignant melanoma. [ABSTRACT FROM AUTHOR]

Lytic virus infection results in production of a virus progeny and lysis of the infected cell. Tumor cells are usually more sensitive to virus infection. Studies indicate that viral oncolysis provides a promising alternative approach to cancer therapy. The ability of viruses to selectively kill cancer cells is long known, but construction of virus variants with an improved therapeutic potential was impossible until recent advances in virus and cell molecular biology and the development of modern methods for directed modification of viruses. Adenoviruses are one of the best studied models of oncolytic viruses. These DNA viruses are convenient for genetic manipulation and show minimal pathogenicity. The review summarizes the data on the directions and approaches to generation of highly efficient variants of oncolytic adenoviruses. The approaches include introduction of directed genetic modifications into the virus genome, accelerated selection of oncolytic virus variants following treatment with mutagens, the use of adenoviruses as vectors to introduce therapeutic gene products, optimization of viral delivery systems, minimization of the negative effects from the host immune system, etc. The dynamic development of studies in the field holds promise that many variants of oncolytic adenoviruses will find clinical application in the nearest future. [ABSTRACT FROM AUTHOR]

Treatment of metastatic tumors with engineered adenoviruses that replicate selectively in tumor cells is a new therapeutic approach in cancer. Systemic administration of these oncolytic adenoviruses lack metastatic targeting ability. The tumor stroma engrafting property of intravenously injected mesenchymal stem cells (MSCs) may allow the use of MSCs as cellular vehicles for targeted delivery. In this work, we study the safety and the efficacy of infusing autologous MSCs infected with ICOVIR-5, a new oncolytic adenovirus, for treating metastatic neuroblastoma. Four children with metastatic neuroblastoma refractory to front-line therapies received several doses of autologous MSCs carrying ICOVIR-5, under an approved preliminary study. The tolerance to the treatment was excellent. A complete clinical response was documented in one case, and the child is in complete remission 3 years after this therapy. We postulate that MSCs can deliver oncolytic adenoviruses to metastatic tumors with very low systemic toxicity and with beneficial antitumor effects. [ABSTRACT FROM AUTHOR]

Abstract: Head and neck squamous cell carcinoma (HNSCC) is a common and often devastating disease without curative treatment when advanced or recurrent. The aim of this study was to assess whether capsid-modified oncolytic adenoviruses have therapeutic efficacy in HNSCC low passage tumour cell cultures and if it could be further improved by combination with cetuximab, radiotherapy and chemotherapy. We investigated which adenoviral capsid modifications allow best gene transfer and cell killing of HNSCC substrates. Gene transfer was assessed using replication-deficient adenoviruses expressing luciferase. Cell killing was studied in vitro and in vivo using oncolytic adenoviruses, which kill tumour cell by viral replication. The most effective capsid-modified oncolytic adenoviruses were combined with HNSCC standard therapies and their efficacy was assessed in vitro as well as in vivo. Cell killing was assessed in vitro by MTS assay and in vivo by HNSCC subcutaneous tumour growth follow-up in nude mice. Cetuximab treatment was found to enrich CD133+ and CD44+ tumour-initiating type cells in tumours grown in mice. Capsid-modified viruses showed increased transduction and oncolysis of HNSCC substrates in comparison to Ad5-based agents. Polylysine (pK7)-modified oncolytic virus resulted in significant tumour growth reduction in vivo. Combination of chemotherapy (cisplatin and 5-fluorouracil), radiotherapy and cetuximab with oncolytic adenovirus therapy resulted in further increases in cell killing effect in vitro and complete eradication of tumours in vivo. Our pre-clinical data suggest that it is feasible and efficacious to combine oncolytic adenoviruses with HNSCC standard therapies into a multimodality treatment regimen for clinical testing in HNSCC patients. [Copyright &y& Elsevier]

The ability of oncolytic adenoviruses to replicate in and lyse cancer cells offers a potential therapeutic approach. However, selectivity and efficacy of adenovirus replication need to be improved. In this study, we present that loss of p21WAF1 promotes adenovirus replication and more effective cell killing. To test our hypothesis, we took HCT116 colon cancer cell lines carrying deletions of either p21WAF1 or p53, and infected these cell lines with wild-type adenovirus (WtD) or the oncolytic adenoviruses, ONYX-015 and Delta-24. We found that WtD, ONYX-015 and Delta-24 induced stronger cytopathic effects in HCT116 p21−/− cells compared with HCT116-WT cells. This was accompanied by increased virus production. siRNA-mediated knockdown of p21WAF1, and similarly of p27KIP1, in HCT116-WT cells also enhanced replication of and cell killing by these viruses. Furthermore, we found that TE7, an esophageal carcinoma cell line, also showed a strong cell-killing effect and virus production when p21WAF1 expression was suppressed by RNA interference before adenoviruses infection. Also, H1299 and DU-145 cells transfected with p21WAF1 siRNA showed higher virus production after ONYX-015 and Delta-24 infections. These observations suggest that p21WAF1 plays a role in mediating replication of oncolytic viruses with potential implications for adenoviral therapy of cancer. [ABSTRACT FROM AUTHOR]

The use of viruses to treat cancer has been studied for decades. With the advancement of molecular biology, viruses have been modified and genetically engineered to optimize their ability to target cancer cells. Canine viruses, such as distemper virus and adenovirus, are being exploited for the treatment of canine cancer as the dog has proven to be a good comparative model for human cancer research and proof of concept investigations. In this review, we introduce the concept of oncolytic viruses and describe some of the preliminary attempts to use oncolytic viruses for the treatment of canine cancer. [ABSTRACT FROM AUTHOR]

Treatment options for pancreatic cancer have limited success and it is therefore an appropriate target for the development of new strategies, including gene therapy. Gene therapy approaches include inhibition of activated oncogenes (KRAS, LSM1) with antisense and RNA interference strategies, replacement of inactivated tumour suppressor genes (TP53, CDKN2A, CDKN1A), targeting of cell signalling pathways, gene-directed prodrug-activation therapies and the use of replication-competent oncolytic viruses. Angiogenesis and apoptosis have also been targeted for gene therapy. Clinical trials of gene therapy have shown only moderate anti-tumour effects. As there are many genetic abnormalities in pancreatic cancer, strategies combining different targets or indeed different modalities of treatment, may be more successful. Identification of new targets and improvements in delivery and targeting may further improve the efficacy of gene therapy in pancreatic cancer. [Copyright &y& Elsevier]

Tumor-endothelial interaction contributes to local prostate tumor growth and distant metastasis. In this communication, we designed a novel approach to target both cancer cells and their“crosstalk”with surrounding microvascular endothelium in an experimental hormone refractory human prostate cancer model. We evaluated the in vitro and in vivo synergistic and/or additive effects of a combination of conditional oncolytic adenovirus plus an adenoviral-mediated antiangiogenic therapy. In the in vitro study, we demonstrated that human umbilical vein endothelial cells (HUVEC) and human C4-2 androgen-independent (AI) prostate cancer cells, when infected with an antiangiogenic adenoviral (Ad)-Flk1-Fc vector secreting a soluble form of Flk1, showed dramatically inhibited proliferation, migration and tubular formation of HUVEC endothelial cells. C4-2 cells showed maximal growth inhibition when coinfected with Ad-Flk1-Fc and Ad-hOC-E1, a conditional replication-competent Ad vector with viral replication driven by a human osteocalcin (hOC) promoter targeting both prostate cancer epithelial and stromal cells. Using a three-dimensional (3D) coculture model, we found that targeting C4-2 cells with Ad-hOC-E1 markedly decreased tubular formation in HUVEC, as visualized by confocal microscopy. In a subcutaneous C4-2 tumor xenograft model, tumor volume was decreased by 40-60%in animals treated with Ad-Flk1-Fc or Ad-hOC-E1 plus vitamin D3 alone and by 90%in a combined treatment group, compared to untreated animals in an 8-week treatment period. Moreover, three of 10 (30%) pre-established tumors completely regressed when animals received combination therapy. Cotargeting tumor and tumor endothelium could be a promising gene therapy strategy for the treatment of both localized and metastatic human prostate cancer.Cancer Gene Therapy (2005) 12, 257-267. doi:10.1038/sj.cgt.7700790 Published online 26 November 2004 [ABSTRACT FROM AUTHOR]

Preclinical and clinical studies have validated the antitumor effects of several oncolytic viruses (OVs). However, the efficacy of OVs is limited when they are administered as monotherapies. Combination therapy is a promising direction for oncolytic virotherapy in the future. A high dose of vitamin C (VitC) exerts anticancer effects by triggering the accretion of substantial amounts of reactive oxygen species (ROS). OVs can induce immunogenic tumor cell death and elicit an antitumor immune response. ROS play an important role in immunogenic cell death (ICD). This study aimed to explore whether high-dose VitC in combination with oncolytic adenoviruses (oAds) exhibited a synergistic antitumor effect. High-dose VitC synergized with oAds against tumor by enhancing immunogenic tumor cell death. Combination therapy with high-dose VitC and oAds significantly increased the number of T cells in the tumor microenvironment (TME) and promoted the activation of T cells. Furthermore, the antitumor effect of the combination therapy was CD8 + T cell dependent. In addition, combination therapy with high-dose VitC and oAds reprogramed the immunosuppressive TME. Our study provides a new strategy for combination therapy of OVs., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

Extracellular vesicles (EVs) are naturally occurring cargo delivery vesicles that have recently received considerable attention for their roles in intercellular communication in many physiological and pathological processes, including tumourigenesis. EVs generated by different tissues demonstrated specific homing: in particular, cancer-derived EVs showed a selective tropism for the tumor tissue from which the vesicles originated. For this property, EVs have been proposed as drug delivery tools for anti-cancer therapies, although the limited knowledge about their in vivo tropism hinders their therapeutic applications. The current study aimed to characterize the targeting properties of cancer-derived EVs in vitro and their biodistribution in vivo , by using an imaging approach. Methods : EVs were generated from: i) murine lung (LL/2) and colon (MC-38) cancer lines, ii) human lung cancer cell line (A549) and iii) human liver biopsy samples from healthy individuals. EVs were loaded with fluorescent dyes alone or in combination with a biopharmaceutical agent, the oncolytic adenovirus (OV), characterized for charge and size and tested for their activity in cancer cell lines. Finally, optical imaging was extensively applied to study in vivo and ex vivo the biodistribution of EVs originated from different sources in different mouse models of cancer, including xenograft, syngeneic graft and the MMTV-NeuT genetically modified animal. Results : We initially demonstrated that even loading EVs even with a large biopharmaceutical oncolytic viruses (OVs) did not significantly change their charge and dimension properties, while increasing their anti-neoplastic activity compared to the virus or EVs alone. Interestingly, this activity was observed even if the EVs derived from lung cancer were applied to colon carcinoma cell lines and vice versa , suggesting that the EV uptake occurred in vitro without any specificity for the cancer cells from which the vesicles originated. When administered i.v (intravenously) to the mouse models of cancer, the tumour-derived EVs, but not the EVs derived from a healthy tissue, demonstrated a selective accumulation of the fluorescence at the tumour site 24 h after injection; adding OVs to the formulation did not change the tumour-specific tropism of the EVs also in vivo . Most interestingly, the in vivo experiments confirmed the in vitro observation of the generalized tropism of tumour-derived EVs for any neoplastic tissue, independent of the tumour type or even the species originating the vesicles. Conclusions : Taken together, our in vitro and in vivo data demonstrate for the first time a heterologous, cross-species tumour-tropism for cancer-derived EVs. This finding challenges our current view on the homing properties of EVs and opens new avenues for the selective delivery of diagnostic/therapeutic agents to solid tumours., Competing Interests: Competing Interests: The authors have declared that no competing interest exists.