Construction of random tumor transcriptome expression library for creating and selecting novel tumor antigens.

Novel tumor antigens are necessary for the development of efficient tumor vaccines for overcoming the immunotolerance and immunosuppression induced by tumors. Here, we developed a novel strategy to create tumor antigens by construction of random tumor transcriptome expression library (RTTEL). The complementary DNA (cDNA) from S180 sarcoma was used as template for arbitrarily amplifying gene fragments with random primers by PCR, then ligated to the C-terminal of HSP65 in a plasmid pET28a-HSP for constructing RTTEL in Escherichia coli. A novel antigen of A5 was selected from RTTEL with the strongest immunotherapeutic effects on S180 sarcoma. Adoptive immunotherapy with anti-A5 sera also inhibited tumor growth, further confirming the key antitumor roles of A5-specific antibodies in mice. A5 contains a sequence similar to protein-L-isoaspartate (D-aspartate) O-methyltransferase (PCMT1). The antisera of A5 were verified to cross-react with PCMT1 by Western blotting assay and vice versa. Both anti-A5 sera and anti-PCMT1 sera could induce antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity toward S180 cells by in vitro assay. Further assay with fluorescent staining showed that PCMT1 is detectable on the surface of S180 cells. Summary, the strategy to construct RTTEL is potential for creating and screening novel tumor antigens to develop efficient tumor vaccines. By RTTEL, we successfully created a protein antigen of A5 with significant immunotherapeutic effects on S180 sarcoma by induction of antibodies targeting for PCMT1.

Anti-tumor effect investigation of obtustatin and crude Macrovipera lebetina obtusa venom in S-180 sarcoma bearing mice.

Over the last few decades, research on snake venom toxins has provided not only new tools to decipher molecular details of various physiological processes, but also inspiration to design and develop a number of therapeutic agents. Isolated from the venom of Macrovipera lebetina obutusa (MLO), obtustatin represents the shortest known snake venom monomeric disintegrin specific inhibitor of α1ß1 integrin. This low molecular weight peptide revealed a potent therapeutic effect on melanoma progression. Its oncostatic effect was related to the inhibition of angiogenesis. The aim of the proposed investigation was to study the influence of obtustatin and crude MLO venom on the S-180 sarcoma growth in vitro and in vivo. A S-180 sarcoma bearing mouse model, histological examination, DNA retardation assay were utilized to investigate the anti-tumor effects of MLO and obtustatin. In addition, some biochemical tests (chemiluminescence-ChL, TBA-test) were applied to elucidate the influence of obtustatin and crude MLO venom on the S-180 sarcoma. The size of tumor was significantly inhibited by MLO venom and obtustatin with the inhibitory rate of 50% and 33% at the doses of 10 µg/mouse and 1mg/kg/day respectively. Both ChL and MDA decrease in the two treated groups. Both obtustatin and MLO venom have an anticancer activity and might be candidates for the treatment of malignant sarcoma. All our results have shown that both obtustatin and MLO venom have an anticancer activity and might be candidates for the treatment of malignant sarcoma.

Cancer resistance as an acquired and inheritable trait.

AIM: To induce cancer resistance in wild-type mice and detect if the resistance could be inherited to the progeny of the induced resistant mice. Furthermore to investigate the spectrum and immunology of this inherited cancer resistance. MATERIALS AND METHODS: Resistance to with live S180 cancer cells in BALB/c mice was induced by immunization with inactivated S180 cancer cells. The immunization was performed by either frozen/thawed or irradiated cancer cells or cell-free ascitic fluid (CFAF). RESULTS: In all instances the induced resistance was demonstrated to be inheritable. The phenotype was named HICR (heritable induced cancer resistance) and was defined as primary resistant progeny from mice immunized with frozen/thawed or irradiated S180 cells or CFAF obtained from mice with S180 induced ascites. Notably, this resistance was transferred from both male and female mice to the offspring of the immunized mice for at least two generations. Although inheritable, the frequency of cancer-resistant pups was lost over a few generations. Cells from the J774A.1 and RAW cancer cell lines did not induce inheritable cancer resistance, and C57BL/6 mice could not pass on cancer resistance fluorescence-activated cell sorting (FACS) analyses of the peritoneal cells revealed an increased fraction of macrophages. In necropsies of resistant mice no histological signs of cancer or other disease was found. CONCLUSION: Only materials derived from S180 cells could give rise to HICR mice. The molecular basis of the resistance is unknown but may involve epigenetic mechanisms. Other examples of inheritability of acquired phenotypic changes exist but, to our knowledge, this is the first demonstration of acquired, inherited cancer resistance.

A study on anti-tumour effect of Solanum lyratum Thunb. extract in S180 tumour-bearing mice.

UNLABELLED: The objective of the study was to investigate the anti-tumour effect of ethanol extract of Solanum lyratum Thunb. in S180 tumour-bearing mice, and to preliminarily explore its mechanism of action. METHODS: Mice were made into S180 solid tumour model, grouped and administered. Tumour inhibition rate was measured by harvesting the tumours. Serum IL-2, TNF-a contents were measured by taking blood samples, and thymus index and spleen index were measured by harvesting the thymus and spleen. The results showed that the Solanum lyratum Thunb. extract had certain tumour inhibitory effect, which can elevate the serum IL-2, TNF-a contents, and increase the thymus and spleen indices to a certain extent. The study concluded that Solanum lyratum Thunb. extract has certain in vivo anti-tumour effect which may be exerted through enhancing the body immunity.

Study on anti-tumor effect of total glycosides from Radix paeoniae rubra in S180 tumor-bearing mice.

The objective of the paper was to study the anti-tumor effect of total glycosides from Radix paeoniae rubra in S180 tumor-bearing mice, and to preliminarily explore its mechanism of action. Mice were made into S180 solid tumor model, grouped and administered with the extracts; tumor inhibition rate was measured by harvesting the tumors, and serum IL-2 and IL-4 levels were measured by taking blood samples. Total glycosides of Radix paeoniae rubra significantly inhibited the growth of tumor cells in tumor-bearing organisms, enhanced the cytotoxic activity of NK cells, and increased the serum IL-2 and IL-4 levels. Total glycosides of Radix paeoniae rubra have some anti-tumor effect in vivo, which might have been accomplished through the regulation of the immune system.

Effects of black yeast-derived ß-1,3-1,6-glucan on serum cytokine and microRNA expression in transplanted sarcoma in mice.

ß-1,3-1,6-glucans are the most abundant glucose polymers in the cell walls of fungi. Previous studies have shown that ß-1,3-1,6-glucans derived from fungi possess immunomodulating activitivies. Antitumor effects of these compounds have also been reported in animal models. Current studies mainly focus on the direct effects of ß-1,3-1,6-glucans on immune systems, but no data are available to address the underlying molecular events in tumor cells. ß-1,3-1,6-glucan purified from black yeast at 5 mg/100 g body weight (study group) or saline (control group) was intragastrically administered on a daily basis to subcutaneously-injected mice with mouse S180 sarcoma cells. Tumor sizes, tumor weights, serum concentrations of cytokines and levels of microRNAs (miRNAs) in transplanted tumors were compared between the treated and control groups. The volumes and weights of transplanted tumors were significantly lower in the treatment groups compared to the control groups by ∼150% and 70%, respectively. The treated mice demonstrated significantly higher levels of cytokines, including IL-2, IL-4, IL-6, IL-8, IL-10 and IL-12, compared to the control mice. Notably, the expression of several miRNAs in transplanted tumor tissues also markedly changed. These data suggest that black yeast-derived ß-1,3-1,6-glucan, not only stimulates cytokine release from immune cells, but also changes the expression profiles of miRNAs in transplanted tumors.