Vaccinia virus (VACV) oncolytic therapy has been successful in a number of tumor models. In this study our goal was to generate a double recombinant vaccinia virus (VV-GMCSF-Lact) with enhanced antitumor activity that expresses exogenous proteins: the antitumor protein lactaptin and human granulocyte-macrophage colony-stimulating factor (GM-CSF). Lactaptin has previously been demonstrated to act as a tumor suppressor in mouse hepatoma as well as MDA-MB-231 human adenocarcinoma cells grafted into SCID mice. VV-GMCSF-Lact was engineered from Lister strain (L-IVP) vaccinia virus and has deletions of the viral thymidine kinase and vaccinia growth factor genes. Cell culture experiments revealed that engineered VV-GMCSF-Lact induced the death of cultured cancer cells more efficiently than recombinant VACV coding only GM-CSF (VV-GMCSF-dGF). Normal human MCF-10A cells were resistant to both recombinants up to 10 PFU/cell. The selectivity index for breast cancer cells measured in pair cultures MCF-7/MCF-10A was 200 for recombinant VV-GMCSF-Lact coding lactaptin and 100 for VV-GMCSF-dGF. Using flow cytometry we demonstrated that both recombinants induced apoptosis in treated cells but that the rate in the cells with active caspase −3 and −7 was higher after treatment with VV-GMCSF-Lact than with VV-GMCSF-dGF. Tumor growth inhibition and survival outcomes after VV-GMCSF-Lact treatment were estimated using immunodeficient and immunocompetent mice models. We observed that VV-GMCSF-Lact efficiently delays the growth of sensitive and chemoresistant tumors. These results demonstrate that recombinant VACVs coding an apoptosis-inducing protein have good therapeutic potential against chemoresistant tumors. Our data will also stimulate further investigation of coding lactaptin double recombinant VACV in clinical settings.
Enzyme immunoassay of the serum neurospecific antigens (gliofibrillar acid protein and neurospecific enolase) was used for evaluation of the resistance of the blood-brain barrier in Wistar rats with perinatal hypoxia and ischemia of the CNS. Perinatal hypoxia and ischemia of the CNS was modeled by two methods: ligation of the common carotid artery in 7-day-old rats followed by 3.5-h hypoxic hypoxia or 15-min anoxic exposure of fetuses isolated via hysterectomy on day 21 of gestation. Enzyme immunoassay of serum gliofibrillar acid protein and neurospecific enolase in control an experimental rat pups was carried out once a week during 3 months. In controls serum levels of gliofibrillar acid protein and neurospecific enolase virtually did not change during postnatal development, while in animals with cerebral hypoxia and ischemia induced in fetuses by both methods serum concentration of neurospecific enolase sharply increased 1 week after the injury and increased on weeks 6 and 10. The content of gliofibrillar acid protein was maximum on week 1 and later considerably varied, the peaks of its concentrations observed on weeks 3 and 8 preceded the increase in neurospecific enolase activity in peripheral blood.
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