Detection and cloning of potent transforming gene(s) from chewing tobacco-related human oral carcinomas

Saranath, Dhananjaya ; Bhoite, Leena T. ; Deo, Madhav G. ; Tandle, Anita T. ; D'Costa, Jenice ; Kolhapure, Rajendra N. ; Govardhan, Manohar K. ; Banerjee, Kalyan (1994) Detection and cloning of potent transforming gene(s) from chewing tobacco-related human oral carcinomas European Journal of Cancer: Part B - Oral Oncology, 30 (4). pp. 268-277. ISSN 0964-1955

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High molecular weight DNA isolated from 14 primary tumour tissues of human oral carcinoma patients was analysed for transforming activity by NIH3T3 co-transfection assay using pSV2neo gene as a selectable marker, followed by nude mouse tumorigenicity assay. Ten of the patient tumour tissues demonstrated molecular lesions in myc, ras or/and EGF-R genes, whereas 4 patients did not show tumour associated aberrations in these oncogenes. The G418-resistant transfected cells from 12 of 14 individual patients demonstrated transforming potential by colony formation in soft agar and tumour induction in nude mice within 25-80 days. DNAs from the transfected cells, consequent nude mice tumours and corresponding cell lines, contained human Alu sequences. Southern blot hybridisation with ras, myc, EGF-R oncogenes demonstrated the presence of human H-ras oncogene in one of the 12 sets of nude mice tumours. In contrast, DNA from the other 11 sets of nude mice tumours indicated absence of c-myc, N-myc, L-myc, H-ras, K-ras, N-ras and EGF-R genes on Southern analysis. Further, DNAs from five first cycle tumorigenic transformants were subjected to a second cycle of transfection, and induced tumours in nude mice with a shorter latency period of 21-50 days. The secondary transformants contained discrete human Alu sequences; however, the DNA did not hybridise with myc/ras/EGF-R probes. A genomic library was constructed from a second cycle nude mice tumour, using EMBL-3 as the vector. Four human Alu sequence positive clones were isolated on screening 2 × 105 plaques, and one of the recombinant clones subjected to fine restriction mapping using 16 restriction enzymes. The lack of association of the nude mice tumour DNA with myc/ras/EGF-R showing aberrations in the primary human tumour, implies activation of an alternative potent transforming gene(s) in the chewing tobacco-related oral carcinomas in India.

Item Type:Article
Source:Copyright of this article belongs to Elsevier Science.
ID Code:21745
Deposited On:22 Nov 2010 10:54
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