British zyxwvutsrqponml Journal zyxwvutsrqponm of HaematoZogy, 1995, 91, 647-651 High frequency of homozygous deletions of CDK4I gene in childhood acute lymphoblastic leukaemia ACHILLE IOLASCON, MARIA FELICIA FAIENZA, BRIGIDA COPPOLA, FULVIO DELLA RAGIONE,* NICOLA SANTORO AND FRANCESCO SCHETTINI Dipartimento di Biomedicina dell'Etu Evolutiva, Universith di Bari, and *Dipartimento di Biochimica delle Macromolecole, zyxwvuts 11 Universita di Napoli, Italy Received zyxwvutsrq 7 9 April 1 995; accepted for publication 7 July 1995 Summary. To determine the incidence of homozygous deletions of the newly identified tumour suppressor gene, CDK41, molecular genomic DNA analyses by PCR technique were performed on primary neoplastic cells from 22 child- hood acute leukaemias obtained at presentation. The blast cells derived in all the analysed cases from bone marrow. We found that none of acute myeloblastic leukaemias (four cases) showed the OK41 alteration, whereas 6/13 (46%) common acute lymphoblastic leukaemias (ALLs) displayed homozygous deletions. Moreover, and even more important, all the blasts purified from zyxwvuts ALLs derived from early lymphoid precursors (three early-T AILS and two pre-B ALLs) showed the absence of CDK4I gene. When the entire coding sequence of the CDK4I gene from samples without homo- zygous deletions was analysed by the single-strand con- formational polymorphism method, no point mutations were identified. These results demonstrate that CDK4.I gene deletions are very frequent and probably early events in childhood acute leukaemias of lymphoid origin and espe- cially in early-T and pre-B ALLs. Moreover, the molecular mechanism of the loss of function of the gene is correlated, at least in childhood ALLs, almost exclusively to deletions and not to point mutations. Keywords: leukaemia, acute leukaemia, p 16, tumour suppressor gene, CDK inhibitor, deletion. Recent advances in the molecular biology of human malignancies have demonstrated conclusively that the genesis and the progression of a tumour are due to the accumulation of multiple genetic alterations, including the gain of function of proto-oncogenes and the inactivation of the so-called tumour suppressor genes (TSGs) (Fearon zyxwvuts & Vogelstein, 1990: Knudson, 1993). In particular, increasing evidence has clearly demonstrated that cancer transformation is mostly due to the loss of function of specific TSGs (Rb, p53 and CDK4I genes) controlling the cell division cycle (Hartwell & Kastan, 1994). Thus, studies on the characterization of cell-cycle-related genes in human malignancies might extend signillcantlyour knowledge of early and late pivotal steps involved in the development of specific types of cancer. These investigations might also give a rich harvest from a prognostic and diagnostic point of view and might be envisioned as the only rationale base to develop new efficacious therapies. Aberrations of the short arm of chromosome 9, which were first reported by Knowalczyk & Sandberg (1983), are Correspondence: Dr Achille Iolascon, Dipartimento di Biomedicina dell'Eta Evolutiva. Piazza Giulio Cesare 11, 70124 Bari, Italy. 0 1995 Blackwell Science Ltd observable in 7-12% of childhood acute lymphoblastic leukaemias (ALL), as well as in a large number of additional neoplasms (Diaz et zyxwvu al, 1990). On these bases, it has been suggested that such abnormalities are related to the loss of function of one (or more) TSG(s). Recently, this putative TSG was identified as the ~ 1 6 ~ ~ gene (also named OK41 or MTS1, multiple tumour suppressor gene 1) by means of positional cloning studies performed on the basis of deletion analyses occurring in malignant cell lines at level of chromosome 9p21 (Kamb et al, 1994; Nobori et al, 1994). The CDK4I gene consists of three coding exons of 12 5 bp, 307bp and 12 bp (Serrano et al, 1993; Kamb et zyx RI, 1994; Nobori et al, 1994). Interestingly, exon 2 codifies for an acidic proteic domain which is 97% homologous to the corresponding exon of an additional gene, named p l SINK* gene (MTSZ) (Hannon & Beach, 1994). This ast gene is localized on chromosome 9p2 1 immediately adjacent (approximately 25kb) to the p161NK4 gene. The protein which is codified by the p161NK4 gene is a key factor in the control of cell proliferation in that it inhibits the activity of two cyclin-dependent serine-threonine kinases (CDKs), namely CDK4 and CDK6. These CDKs are the catalytic subunit of quaternary complexes specifically activated 647