POTENTIAL ROLE FOR CONCURRENT ABNORMALITIES OF THE CYCLIN D1, P16(CDKN2) AND P15(CDKN2B) GENES IN CERTAIN B-CELL NON-HODGKINS-LYMPHOMAS - FUNCTIONAL-STUDIES IN A CELL-LINE (GRANTA-519)

Abnormalities of several cell-cycle regulatory genes including cyclin D1, p16(CDKN2) and p15(CDKN2B) have been described in B cell non-Hodgk in's lymphoma (B-NHL). We describe a new B-NHL cell line (Granta 519), with concurrent abnormalities of the cyclin D1, p16(CDKN2) and p15(CD KN2B) genes. An independent clinical case of mantle cell NHL (Mc-NHL) with concomitant overexpression of cyclin D1, and deletion of the p16( CDKN2) gene was also identified, suggesting that this combination of o ncogenic aberration is a pathophysiologic contribution to a subset of NHL cases. More in-depth functional studies of this concept were facil itated by the availability of the cell line Granta 519 which was deriv ed from a case of high-grade NHL and has a mature B cell immunophenoty pe. Cytogenetic analysis identified translocation t(11;14)(q13;q32) an d complex rearrangements involving chromosomes 9p22, 13p21, 17p11, and 18q21. Molecular analysis identified overexpression of cyclin D1 mRNA and biallelic deletion of the p16(CDKN2) and p15(CDKN2B) genes. To el ucidate the effect of these genetic abnormalities on the G1 control of Granta 519 cells, the level and function of the major components of t he cyclinD/retinoblastoma (RE) pathway were investigated. Cyclin D1 wa s dominant among the D-type cyclins, formed abundant complexes with cy clin-dependent kinase (Cdk) Cdk4 rather than Cdk6, and the immunopreci pitated cyclin D1/Cdk4 holoenzyme was active as a PRE kinase. Electrop oration of wild-type p16(CDKN2) arrested the Granta 519 cells in G1, c onsistent with the p16(CDKN2) loss as a biologically relevant event du ring multistep evolution of the tumor, and with the expression of func tional pRB. Direct cooperation of these distinct abnormalities to cell -cycle deregulation in NHL cells was suggested by G1 acceleration upon inducible overexpression of cyclin D1 in a control breast cancer cell line lacking p16(CDKN2), an effect which could be prevented by ectopi c expression of p16(CDKN2). Taken together, these data suggest that co ncurrent overexpression of cyclin D1 and functional elimination of p16 (CDKN2) and p15(CDKN2B) may characterize certain cases of mantle cell NHL, and that cooperation of the abnormalities is likely to provide a growth advantage of the tumour cells through more efficient inactivati on of the RE tumor suppressor. Further clinicopathologic studies of th is possibility are warranted.