A recombinant vaccinia virus VVdGF-ApoS24/2 expressing apoptin selectively kills human cancer cells in vitro [Kochneva et al., 2013]. We compared the oncolytic activity of this recombinant with that of the parental strain L-IVP using a model of human A431 carcinoma xenografts in nude mice. Single intratumoral injections (2×107 PFU/mouse) of the viruses produced a dramatic decrease in tumor volumes, which was higher after injection of apoptin-producing virus. The tumor dried out after the injection of recombinant while injection of L-IVP strain resulted in formation of cavities filled with cell debris and liquid. Both viruses rapidly spread in xenografts and replicate exclusively in tumor cells causing their destruction within 8 days. Both viruses induced insignificant level of apoptosis in tumors. Unlike the previously described nuclear localization of apoptin in cancer cells the apoptin produced by recombinant virus was localized to the cytoplasm. The apoptin did not induce a typical apoptosis, but it rather influenced pathway of cell death and thereby caused tumor shrinkage. The replacement of destroyed cells by filamentous material is the main feature of tumor regression caused by the VVdGF-ApoS24/2 virus. The study points the presence of complicated mechanisms of apoptin effects at the background of vaccinia virus replication.

BackgroundHepatocellular carcinoma (HCC) is one of the most common malignancies worldwide with limited therapeutic options. Since HCC has been shown to be immunogenic, immunotherapy is considered a promising therapeutic approach. Small interfering RNAs (siRNAs), depending on their structure and sequence, can trigger the innate immune system, which can potentially enhance the adaptive anticancer immune response in the tumor-bearing subjects. Immunostimulatory properties of nucleic acids can be applied to develop adjuvants for HCC treatment.MethodsThe transplantable HCC G-29 tumor in male CBA/LacSto (CBA) mice was used to study the effects of immunostimulatory RNA on tumor growth. Tumor size, metastases area in different organs of mice and mouse survival rate were analyzed. Furthermore the mouse serum IFN-α levels were measured using ELISA.ResultsIn the present study, we found that a 19-bp RNA duplex (ImmunoStimulattory RNA or isRNA) with 3-nt overhangs at the 3′-ends of specific sequence displays immunostimulatory, antitumor, and antimetastatic activities in mice bearing HCC G-29. Our results demonstrate that isRNA strongly increases the level of interferon-α (IFN-α) by up to 25-fold relative to the level in mice injected with Lipofectamine alone (Mock), and to a lesser extent increases the level of proinflammatory cytokine interleukin-6 (IL-6) (by up to 5.5-fold relative to the Mock level), in mice blood serum. We showed that isRNA reliably (P < 0.05) inhibits primary tumor growth in mice compared to the mock group. Furthermore, injections of isRNA significantly enhanced necrotic processes in the center of the primary tumor, and decreased by twofold the width of the undifferentiated peripheral zone and the number of mitotic cells in this zone. The results showed that isRNA efficiently reduces the area of metastases in the liver, kidneys, and heart of CBA/LacSto mice with HCC.ConclusionsThe obtained results clearly demonstrate immunostimulatory and antimetastatic properties of the isRNAs in mice with HCC. Consequently, this short double-stranded RNA can be considered as a potential adjuvant for the therapy of HCC.

Oncolytic abilities of vaccinia virus (VACV) served as a basis for the development of various recombinants for treating cancer; however, “natural” oncolytic properties of the virus are not examined in detail. Our study was conducted to know how the genetically unmodified L-IVP strain of VACV produces its antitumor effect. Human A431 carcinoma xenografts in nude mice and murine Ehrlich carcinoma in C57Bl mice were used as targets for VACV, which was injected intratumorally. A set of virological methods, immunohistochemistry, light and electron microscopy was used in the study. We found that in mice bearing A431 carcinoma, the L-IVP strain was observed in visceral organs within two weeks, but rapidly disappeared from the blood. The L-IVP strain caused decrease of sizes in both tumors, however, in different ways. Direct cell destruction by replicating virus plays a main role in regression of A431 carcinoma xenografts, while in Ehrlich carcinoma, which poorly supported VACV replication, the virus induced decrease of mitoses by pushing tumor cells into S-phase of cell cycle. Our study showed that genetically unmodified VACV possesses at least two mechanisms of antitumor effect: direct destruction of tumor cells and suppression of mitoses in tumor cells.