Decreased cyclin B1 expression contributes to G 2 delay in human brain tumor cells after treatment with camptothecin 1 Anna J. Janss, 2 Amit Maity, Cheng-Bi Tang, Ruth J. Muschel, W. Gillies McKenna, Leslie Sutton, and Peter C. Phillips Divisions of Neurology (A.J.J., C.B.T., P.C.P.), Oncology (A.J.J., P.C.P.), and Neurosurgery (L.S.), Children’s Hospital of Philadelphia, 324 South 34th St., Philadelphia, PA 19104-9786; and Departments of Radiation Oncology (A.M., W.G.M.) and Pathology and Laboratory Medicine (R.J.M.), University of Pennsylvania School of Medicine, Philadelphia, PA 19104 DNA damage produces delayed mitosis (G 2 /M delay) in proliferating cells, and shortening the delay sensitizes human malignant glioma and medulloblastoma cells to cytotoxic chemotherapy. Although activation of the cyclin-dependent kinase CDC2 mediates G 2 /M transi- tion in all tumor cells studied to date, regulation of CDC2 varies between tumor types. Persistent hyper- phosphorylation of kinase and reduced cyclin expres- sion have been implicated as mediators of treatment- induced G 2 delay in different tumor models. To evaluate regulation of G 2 /M transition in human brain tumors, we studied the expression and/or activity of CDC2 kinase and cyclins A and B1 in U-251 MG and DAOY medulloblastoma cells after their treatment with camp- tothecin (CPT). Synchronized cells were treated during S phase, then harvested at predetermined intervals for evaluation of cell cycle kinetics, kinase activity mRNA, and protein expression. CPT produced G 2 delay associ- ated with decreased CDC2 kinase activity and cyclin B1 expression. Kinase activity was associated with CDC2 bound to cyclin B1, not cyclin A, in both cell lines. Cyclin A mRNA and protein expression were reduced after CPT treatment; however, decreased protein expres- sion was short lived and moderate in the glioma and primitive neuroectodermal tumor/medulloblastoma cells, respectively. We conclude that G 2 delay is a com- mon response of brain tumor cells to chemotherapy with topoisomerase I inhibitors and that a mechanism of this delay may be reduced expression of cyclin B1. Neuro- Oncology 3, 11–21, 2001 (Posted to Neuro-Oncology [serial online], Doc. 00-025, November 8, 2000. URL <neuro-oncology.mc.duke.edu>) S ignals that regulate transition between cell cycle phases in proliferating cells represent potential tar- gets for new cancer treatment strategies because they modulate cell responses to DNA injury as well as progression through the cell cycle (Johnson and Walker, 1999). Inhibition of cellular division by delaying the transition from the second gap phase of the cell cycle (G 2 ) to mitosis (M) is an important cellular response to DNA-damaging agents (Lock and Ross, 1990; McKenna et al., 1991; O’Connor et al., 1993). G 2 delay may be necessary to complete DNA repair before dam- aged, potentially lethal replicons are conferred to the next generation of cells through mitosis. Cytotoxicity of radiation and chemotherapy is inversely associated with the duration of radiation- or chemotherapy-induced G 2 delay in human lymphoma (O’Connor and Kohn, 1992) and transformed rat embryo cells (McKenna et al., 1991). Furthermore, pharmacologic abrogation of radi- ation- or drug-induced G 2 delay with caffeine, stau- rosporine, or pentoxifylline increases cell death in HeLa cells (Bernhard et al., 1994), human lymphoma cells Received 5 April 2000, accepted 17 August 2000. 1 Supported by National Institutes of Health Grants GM 47439 (R.J.M.), CA-36245 (P.C.P.), and NS-31102 (P.C.P.); and a grant from the American Brain Tumor Association/Jordan Bassett Fellowship (A.J.J.). 2 Address correspondence and reprint requests to Anna J. Janss, Division of Neurology, Children’s Hospital of Philadelphia, 324 South 34th St., Philadelphia, PA 19104-9786. 3 Abbreviations used are as follows: CPT, camptothecin. Neuro-Oncology Neuro-Oncology n JANUARY 2001 11 Downloaded from https://academic.oup.com/neuro-oncology/article/3/1/11/1071870 by guest on 30 September 2022