Positional cloning of tumor suppressor genes from 3p21.3 involved in major human cancers

Abstract

We have performed a comprehensive deletion survey of 3p on more than 400 lung, renal, breast, cervical and ovarian carcinomas (major epithelial cancers) using a defined set of markers, combining conventional LOH with quantitative real-time PCR (QPCR), NotI linking and jumping libraries, comparative genomic and NotI microarrays hybridizations. We identified two most frequently affected 3p21.3 regions, LUCA (LUng CAncer) at the centromeric and AP20 at the telomeric border of 3p21.3. Aberrations of either region were detected in more than 90% of the studied tumors suggesting they harbor multiple tumor suppressor genes (TSG). Homozygous deletions (HD) were frequently detected in all tumors in both the LUCA and AP20 regions. To facilitate the identification of tumor suppressor genes (TSGs) in the chromosome 3p21.3 AP20 and LUCA sub-regions, we constructed physical and gene map of these segments. More than 30 genes were localized in these two regions and among them at least 12 TSGs were identified: RBSP3, ITGA9, MLH1, VILL, APRG1, RASSF1, HYAL1, HYAL2, SEMA3B, SEMA3F, NPRL2 and CACNA2D2. Among these TSGs were representatives of new types of TSGs: e.g. HYAL1 showing growth inhibiting activity only in vivo and RASSF1A and RASSF1C that are alternative forms of the same gene showing different tissue specificity. We found that several tumor suppressor genes in AP20 and LUCA 3p21.3 regions were co-regulated in tumors. These results supported the hypothesis on simultaneous inactivation of clusters cancer-causing genes in AP20 and LUCA regions during the development and progression of lung cancer and other epithelial tumors. Moreover we found an exceptionally high incidence of single-base mutations in the tumor suppressor genes RASSF1 and RBSP3 (CTDSPL) in major epithelial tumors. These mutations functionally inactivated tumor suppressor activity of these genes. Somatic hypermutations in tumor suppressor genes involved in major human malignancies offer a novel insight in cancer development, progression and spread. The data could be important for development of specific biomarker sets for early cancer diagnosis and new therapeutic approaches/strategies for cancer treatment. For example we selected a set of 23 markers (BHLHB2, FBLN2, EPHB1, GATA2, GORASP1, PRICKLE2, Hmm61490, ITGA9, LOC285205, LRRC3B, MINA, MITF, MRPS17P3, NKIRAS1, PLCL2, TRH, UBE2E2, WNT7A, RARB2, p20-CGGBP, GNAI2, RPL32, THRB) that would discriminate/diagnose the majority of NSCLC cases with P > 95% and most complicated cases with a probability of more than 80%.