Oncolytic adenoviruses (OAds) are very promising for the treatment of lung cancer. However, OAd-based monotherapeutics have not been effective during clinical trials. Therefore, the effectiveness of virotherapy must be enhanced by combining OAds with other therapies. In this study, the therapeutic potential of OAd in combination with temozolomide (TMZ) was evaluated in lung cancer cells in vitro and in vivo. The combination of OAd and TMZ therapy synergistically enhanced cancer cell death; this enhanced cancer cell death may be explained via three related mechanisms: apoptosis, virus replication, and autophagy. Autophagy inhibition partially protected cancer cells from this combined therapy. This combination significantly suppressed the growth of subcutaneous H441 lung cancer xenograft tumors in athymic nude mice. In this study, we have provided an experimental rationale to test OAds in combination with TMZ in a lung cancer clinical trial.

The SA-4-1BBL, an oligomeric novel form of the natural ligand for the 4-1BB co-stimulatory receptor of the tumor necrosis factor (TNF) superfamily, as a recombinant protein has potent pleiotropic effects on cells of innate, adaptive, and regulatory immunity with demonstrated therapeutic efficacy in several tumor models. However, the production of soluble form of SA-4-1BBL protein and quality control is time and resource intensive and face various issues pertinent to clinical development of biologics. The present study sought to take advantage of the simplicity and translatability of DNA-based vaccines for the production and delivery of SA-4-1BBL for cancer immune prevention and therapy. A chimeric HPV-16 E7 DNA vaccine (SP-SA-E7-4-1BBL) was constructed that contains the signal peptide (SP) of calreticulin (CRT), streptavidin (SA) domain of SA-4-1BBL, HPV-16 E7 double mutant gene, and the extracellular domain of mouse 4-1BBL. Immunization by gene gun with SP-SA-E7-4-1BBL induced greater prophylactic as well as therapeutic effects in C57BL/6 mice against TC-1 tumor model compared with immunization with E7wt, SP-SA-4-1BBL or reference-positive control CRT-E7wt. The therapeutic efficacy of the DNA vaccine was associated with increased frequency of E7-specific T cells producing interferon (IFN)-γ. Overall, our data suggest that this DNA-based vaccine strategy might represent a translational approach because it provides a simpler and versatile alternative to a subunit vaccine based on SA-4-1BBL and E7 proteins.

Fluorescent proteins are useful reporter molecules for a variety of biological systems. We present an alternative strategy for cloning reporter genes that are regulated by the nisin-controlled gene expression (NICE) system. Lactoccocus lactis was genetically engineered to express green fluorescent protein (GFP), mCherry or near-infrared fluorescent protein (iRFP). The reporter gene sequences were optimized to be expressed by L. lactis using inducible promoter pNis within the pNZ8048 vector. Expression of constructions that carry mCherry or GFP was observed by fluorescence microscopy 2 h after induction with nisin. Expression of iRFP was evaluated at 700 nm using an infrared scanner; cultures induced for 6 h showed greater iRFP expression than non-induced cultures or those expressing GFP. We demonstrated that L. lactis can express efficiently GFP, mCherry and iRFP fluorescent proteins using an inducible expression system. These strains will be useful for live cell imaging studies in vitro or for imaging studies in vivo in the case of iRFP.

Auranofin (AF) is a gold compound approved in 1985 as a main treatment against rheumatoid arthritis. However, as new anti-rheumatoid agents displaced the use of AF, there have been studies aiming to repurpose the drug to treat other diseases, including cancer. In this study, we investigated the potentiality of AF to impair the growth of high-grade serous ovarian cancer (HGSOC) cells regardless of platinum (CDDP) sensitivity. Preliminary studies utilizing a colorimetric proliferation assay revealed that HGSOC cells that are clinically sensitive (PEO1) or resistant (PEO4) to CDDP-based chemotherapy are both equally sensitive to the growth inhibition induced by AF. The resistant factor estimated for AF-defined as the ratio between IC50 values calculated for the resistant cells and for the sensitive ones—was close to one, suggesting that CDDP-resistant PEO4 cells are equally sensitive to AF than CDDP-sensitive PEO1 cells. As expected, the resistant factor for CDDP was 12-fold higher in PEO4 compared to PEO1. The toxicity of AF against HGSOC cells was further studied exposing PEO1 to various concentrations of AF and assessing cell number, cell viability, and cell cycle traverse after 72 h. Results showed that AF, in a dose-dependent manner, blocked cell proliferation and caused accumulation of cells with hypo-diploid DNA content, suggesting that AF-induced cytotoxicity was associated not only with inhibition of cell proliferation but also with cell death likely driven via apoptosis. We further studied the long-term or residual toxicity of AF by exposing PEO1 cells that were alive, after 72 h treatment with AF, to a clonogenic survival assay. The results clearly demonstrated that even if not dying within 72 h, the cells were affected in a manner not compatible with long-term cell cycle progression as demonstrated by the formation of ‘abortive' colonies (i.e. colonies with few cells that show abnormal phenotypes). Finally, as AF is a pro-oxidant agent, we evaluated whether AF-induced cell death was mediated by oxidative stress by incubating PEO1 cells with AF in the presence or absence of the anti-oxidant N-acetyl-cysteine (NAC). PEO1 cells treated with AF and NAC showed no decrease in cell viability and cell number in comparison to PEO1 cells treated with AF alone. This data reveals that AF most likely induces cell death in a reactive oxygen species (ROS)-dependent manner. In conclusion, our results suggest that AF can equally impair the growth of CDDP-sensitive or -resistant HGSOC cells, and that its toxicity is likely mediated by oxidative stress. Citation Format: Farah H. Abdalbari, Alicia A. Goyeneche, Elvis Martinez-Jaramillo, Siham Sabri, Carlos M. Telleria. Repurposing the anti-rheumatic gold compound auranofin for high-grade serous ovarian cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1014.

Conditionally replicative adenoviruses (CRAds) replicate poorly in murine cancer cells; however, E1b-deleted CRAds may replicate effectively in HPV16-E6/E7-positive murine cancer cells (TC-1). The HPV16 E7 open reading frame encodes functions analogous to these deleted adenovirus E1 proteins. In this study, an E1b-deleted CRAd (Adhz60) was evaluated for its ability to replicate and induce oncolysis in TC-1 cells. Adhz60-mediated oncolysis was similar in TC-1 and HeLa cells. Productive viral replication was evident based on expression of E1A and hexon, production of infectious virus progeny, and Adhz60-induced apoptosis. The results suggest that TC-1 murine cancer cells allow Adhz60 replication and oncolysis.

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