Tumor suppressor function of the SEMA3B gene in human lung and renal cancers

The SEMA3B gene is located in the 3p21.3 LUCA region, which is frequently affected in different types of cancer. The objective of our study was to expand our knowledge of the SEMA3B gene as a tumor suppressor and the mechanisms of its inactivation. In this study, several experimental approaches were used: tumor growth analyses and apoptosis assays in vitro and in SCID mice, expression and methylation assays and other. With the use of the small cell lung cancer cell line U2020 we confirmed the function of SEMA3B as a tumor suppressor, and showed that the suppression can be realized through the induction of apoptosis and, possibly, associated with the inhibition of angiogenesis. In addition, for the first time, high methylation frequencies have been observed in both intronic (32-39%) and promoter (44-52%) CpG-islands in 38 non-small cell lung carcinomas, including 16 squamous cell carcinomas (SCC) and 22 adenocarcinomas (ADC), and in 83 clear cell renal cell carcinomas (ccRCC). Correlations between the methylation frequencies of the promoter and the intronic CpG-islands of SEMA3B with tumor stage and grade have been revealed for SCC, ADC and ccRCC. The association between the decrease of the SEMA3B mRNA level and hypermethylation of the promoter and the intronic CpG-islands has been estimated in renal primary tumors (P < 0.01). Using qPCR, we observed on the average 10- and 14-fold decrease of the SEMA3B mRNA level in SCC and ADC, respectively, and a 4-fold decrease in ccRCC. The frequency of this effect was high in both lung (92-95%) and renal (84%) tumor samples. Moreover, we showed a clear difference (P < 0.05) of the SEMA3B relative mRNA levels in ADC with and without lymph node metastases. We conclude that aberrant expression and methylation of SEMA3B could be suggested as markers of lung and renal cancer progression

Measurement of the e+e−→KSKLπ0\mathbf{e^+e^- \rightarrow K_S K_L \pi^0} cross section in the energy range s=1.3−2.0\mathbf{\sqrt{s}=1.3-2.0} GeV

The $e^+e^- \rightarrow K_S K_L \pi^0$ cross section is measured in the center-of-mass energy range $\sqrt{s}=1.3-2.0$ GeV. The analysis is based on the data sample with an integrated luminosity of 33.5 pb$^{-1}$ collected with the SND detector at the VEPP-2000 $e^+e^-$ collider.Comment: 8 pages, 3 figures, accepted for publication in Phys. Rev.

Simultaneous down-regulation of tumor suppressor genes RBSP3/CTDSPL, NPRL2/G21 and RASSF1A in primary non-small cell lung cancer

<p>Abstract</p> <p>Background</p> <p>The short arm of human chromosome 3 is involved in the development of many cancers including lung cancer. Three bona fide lung cancer tumor suppressor genes namely <it>RBSP3 </it>(AP20 region),<it>NPRL2 </it>and <it>RASSF1A </it>(LUCA region) were identified in the 3p21.3 region. We have shown previously that homozygous deletions in AP20 and LUCA sub-regions often occurred in the same tumor (P < 10^-6).</p> <p>Methods</p> <p>We estimated the quantity of <it>RBSP3, NPRL2, RASSF1A, GAPDH, RPN1 </it>mRNA and <it>RBSP3 </it>DNA copy number in 59 primary non-small cell lung cancers, including 41 squamous cell and 18 adenocarcinomas by real-time reverse transcription-polymerase chain reaction based on TaqMan technology and relative quantification.</p> <p>Results</p> <p>We evaluated the relationship between mRNA level and clinicopathologic characteristics in non-small cell lung cancer. A significant expression decrease (≥2) was found for all three genes early in tumor development: in 85% of cases for <it>RBSP3</it>; 73% for <it>NPRL2 </it>and 67% for <it>RASSF1A </it>(P < 0.001), more strongly pronounced in squamous cell than in adenocarcinomas. Strong suppression of both, <it>NPRL2 </it>and <it>RBSP3 </it>was seen in 100% of cases already at Stage I of squamous cell carcinomas. Deregulation of <it>RASSF1A </it>correlated with tumor progression of squamous cell (P = 0.196) and adenocarcinomas (P < 0.05). Most likely, genetic and epigenetic mechanisms might be responsible for transcriptional inactivation of <it>RBSP3 </it>in non-small cell lung cancers as promoter methylation of <it>RBSP3 </it>according to NotI microarrays data was detected in 80% of squamous cell and in 38% of adenocarcinomas. With NotI microarrays we tested how often LUCA (<it>NPRL2, RASSF1A</it>) and AP20 (<it>RBSP3</it>) regions were deleted or methylated in the same tumor sample and found that this occured in 39% of all studied samples (P < 0.05).</p> <p>Conclusion</p> <p>Our data support the hypothesis that these TSG are involved in tumorigenesis of NSCLC. Both genetic and epigenetic mechanisms contribute to down-regulation of these three genes representing two tumor suppressor clusters in 3p21.3. Most importantly expression of <it>RBSP3, NPRL2 </it>and <it>RASSF1A </it>was simultaneously decreased in the same sample of primary NSCLC: in 39% of cases all these three genes showed reduced expression (P < 0.05).</p

Differential Expression of CHL1 Gene during Development of Major Human Cancers

CHL1 gene (also known as CALL) on 3p26.3 encodes a one-pass trans-membrane cell adhesion molecule (CAM). Previously CAMs of this type, including L1, were shown to be involved in cancer growth and metastasis.We used Clontech Cancer Profiling Arrays (19 different types of cancers, 395 samples) to analyze expression of the CHL1 gene. The results were further validated by RT-qPCR for breast, renal and lung cancer. Cancer Profiling Arrays revealed differential expression of the gene: down-regulation/silencing in a majority of primary tumors and up-regulation associated with invasive/metastatic growth. Frequent down-regulation (>40% of cases) was detected in 11 types of cancer (breast, kidney, rectum, colon, thyroid, stomach, skin, small intestine, bladder, vulva and pancreatic cancer) and frequent up-regulation (>40% of cases)--in 5 types (lung, ovary, uterus, liver and trachea) of cancer. Using real-time quantitative PCR (RT-qPCR) we found that CHL1 expression was decreased in 61% of breast, 60% of lung, 87% of clear cell and 89% papillary renal cancer specimens (P<0.03 for all the cases). There was a higher frequency of CHL1 mRNA decrease in lung squamous cell carcinoma compared to adenocarcinoma (81% vs. 38%, P = 0.02) without association with tumor progression.Our results suggested that CHL1 is involved in the development of different human cancers. Initially, during the primary tumor growth CHL1 could act as a putative tumor suppressor and is silenced to facilitate in situ tumor growth for 11 cancer types. We also suggested that re-expression of the gene on the edge of tumor mass might promote local invasive growth and enable further metastatic spread in ovary, colon and breast cancer. Our data also supported the role of CHL1 as a potentially novel specific biomarker in the early pathogenesis of two major histological types of renal cancer

High Mutability of the Tumor Suppressor Genes RASSF1 and RBSP3 (CTDSPL) in Cancer

BACKGROUND:Many different genetic alterations are observed in cancer cells. Individual cancer genes display point mutations such as base changes, insertions and deletions that initiate and promote cancer growth and spread. Somatic hypermutation is a powerful mechanism for generation of different mutations. It was shown previously that somatic hypermutability of proto-oncogenes can induce development of lymphomas. METHODOLOGY/PRINCIPAL FINDINGS:We found an exceptionally high incidence of single-base mutations in the tumor suppressor genes RASSF1 and RBSP3 (CTDSPL) both located in 3p21.3 regions, LUCA and AP20 respectively. These regions contain clusters of tumor suppressor genes involved in multiple cancer types such as lung, kidney, breast, cervical, head and neck, nasopharyngeal, prostate and other carcinomas. Altogether in 144 sequenced RASSF1A clones (exons 1-2), 129 mutations were detected (mutation frequency, MF = 0.23 per 100 bp) and in 98 clones of exons 3-5 we found 146 mutations (MF = 0.29). In 85 sequenced RBSP3 clones, 89 mutations were found (MF = 0.10). The mutations were not cytidine-specific, as would be expected from alterations generated by AID/APOBEC family enzymes, and appeared de novo during cell proliferation. They diminished the ability of corresponding transgenes to suppress cell and tumor growth implying a loss of function. These high levels of somatic mutations were found both in cancer biopsies and cancer cell lines. CONCLUSIONS/SIGNIFICANCE:This is the first report of high frequencies of somatic mutations in RASSF1 and RBSP3 in different cancers suggesting it may underlay the mutator phenotype of cancer. Somatic hypermutations in tumor suppressor genes involved in major human malignancies offer a novel insight in cancer development, progression and spread