Promotion of incisional wound repair by human mesenchymal stem cell transplantation.

The purpose of this study was to determine the effect of transplanted human mesenchymal stem cells (hMSCs) on wound healing. In this model, full-thickness cutaneous wounds were created by incision in the skin of adult New Zealand white rabbits and treated by transplanted hMSCs into the wounds. Wound healing was evaluated by histological analysis and tensiometry over time. A total of 15 New Zealand white rabbits with 10 wounds per animal were examined in this study. Animals were treated with hMSCs and euthanised at 3, 7, 14, 21 and 80 days after manipulation. The hMSCs were labelled with a fluorescent dye (CM-DiI), suspended in phosphate-buffered saline and used to treat full-thickness incisional wounds in rabbit skin. Tensiometry and histology were used to characterise the wound-healing rate of the incisional wounds. These results showed that transplanted hMSCs significantly inhibited scar formation and increased the tensile strength of the wounds. Importantly, MSCs from genetically unrelated donors did not appear to induce an immunologic response. In conclusion, human mesenchymal stem cell therapy is a viable approach to significantly affect the course of normal cutaneous wound healing and significantly increase the tensile strength.

Combining high selectivity of replication via CXCR4 promoter with fiber chimerism for effective adenoviral oncolysis in breast cancer.

Conditionally replicative adenoviruses (CRAds) represent novel therapeutic agents that have been recently applied in the context of breast cancer therapy. However, deficiencies in the ability of the adenovirus to infect target tumor cells and to specifically replicate within the tumor target represent key deficiencies preventing the realization of the full potential of this therapeutic approach. Minimal expression of the adenovirus serotype 5 (Ad5) receptor CAR (coxsackie and adenovirus receptor) on breast cancer cells represents a major limitation for Ad5-based virotherapy. Genetic fiber chimerism is a method to alter the tropism of Ad5-based CRAds to achieve CAR-independent infectivity of tumor cells. Here, we describe the use of a CRAd with cancer specific transcriptional control of the essential Ad5 E1A gene using the human CXCR4 gene promoter. We further modified the fiber protein of this agent by switching the knob domain with that of the adenovirus serotype 3. The oncolytic activity of this 5/3 fiber-modified CRAd was studied in breast cancer cell lines, primary breast cancer and human liver tissue slices from patients, and in a xenograft breast cancer mouse model. This infectivity enhanced CRAd agent showed improved replication and killing in breast cancer cells in vitro and in vivo with a remarkable specificity profile that was strongly attenuated in nonbreast cancer cells, as well as in normal human breast and liver tissues. In conclusion, utilization of a CRAd that combined infectivity enhancement strategies and transcriptional targeting improved the CRAd-based antineoplastic effects for breast cancer therapy.

Employment of liver tissue slice analysis to assay hepatotoxicity linked to replicative and nonreplicative adenoviral agents.

Whereas virotherapy has emerged as a novel and promising approach for neoplastic diseases, appropriate model systems have hampered preclinical evaluation of candidate conditionally replicative adenovirus agents (CRAds) with respect to liver toxicity. This is due to the inability of human viral agents to cross species. We have recently shown the human liver tissue slice model to be a facile means to validate adenoviral replication. On this basis, we sought to determine whether our ex vivo liver tissue slice model could be used to assess CRAd-mediated liver toxicity. We analyzed and compared the toxicity of a conditionally replicative adenovirus (AdDelta24) to that of a replication incompetent adenovirus (Adnull [E1-]) in mouse and human liver tissue slices. To accomplish this, we examined the hepatic apoptosis expression profile by DNA microarray analyses, and compared these results to extracellular release of aminotransferase enzymes, along with direct evidence of apoptosis by caspase-3 immunhistochemical staining and TUNEL assays. Human and mouse liver tissue slices demonstrated a marked increase in extracellular release of aminotransferase enzymes on infection with AdDelta24 compared to Adnull. AdDelta24-mediated liver toxicity was further demonstrated by apoptosis induction, as detected by caspase-3 immunohistochemical staining, TUNEL assay and microarray analysis. In conclusion, concordance of CRAd-mediated apoptosis in both the human and the mouse liver tissue slice models was demonstrated, despite the limited replication ability of CRAds in mouse liver slices. The results of this study, defining the CRAd-mediated apoptosis gene expression profiles in human and mouse liver, may lay a foundation for preclinical liver toxicity analysis of CRAd agents.

p53 Adenovirus as Gene Therapy for Ovarian Cancer.

Ovarian cancer arises from the accumulation of mutations in multiple combinations of genes (1). The most extensively studied tumor suppressor gene in solid tumors is p53, a 53-kD nuclear phosphoprotein that binds DNA. The p53 gene product plays a role in normal cellular proliferation by regulating gene transcription, cell cycle control, and apoptosis (2). Mutations of p53 are the most common molecular genetic abnormality to be described in human cancer, and have been identified in malignancies of the breast, colon, lung, esophagus, head and neck, and hematopoietic system (3). Mutations of the p53 gene have been identified in 30 to 79% of epithelial ovarian cancers (4,5). Most of the mutations identified in p53 are distributed throughout the open reading frame as missense mutations. We have identified a missense mutation in the p53 gene in the 2774 ovarian cancer cell line that converts an arginine residue in the DNA binding region of the protein to a histidine residue (6). The mutation in codon 273 we found in 2774 cells is one of the six major hotspots identified for p53 missense mutations (7).