Results were then analyzed with the ABI Prism 7700 sequence detection system software (Applied Biosystems)
Results were then analyzed with the ABI Prism 7700 sequence detection system software (Applied Biosystems). leukemias in adults. The disease is usually characterized by the accumulation of monoclonal CD5+ B lymphocytes arrested in the G0/G1 phases of the cell cycle. The primary pathogenic event that causes the generation of aberrant B cells remains to be decided, but impaired apoptosis results in increased survival of circulating CLL cells. In contrast SMIP004 to most other hematological malignancies, CLL cells have a negligibly low fraction of growing cells. The clinical course of the disease is usually highly variable and ranges from indolent disease requiring no treatment over years to highly aggressive disease that needs immediate therapy. So far, all accepted treatment regimens for CLL are neither curative nor associated with prolonged survival (1). CLL therefore presents unique challenges to the development of new treatment strategies. Apoptosis can be induced via two main pathways: interference with mitochondrial function, also called the intrinsic pathway, or triggering of cell surface death receptors, also called the extrinsic pathway (2,3). The intrinsic pathway is usually triggered by stress or chemotherapeutic brokers and is regulated by the balance of pro- and antiapoptotic members of the Bcl-2 family of proteins. The extrinsic or receptor-dependent pathway is usually induced by members of the tumor necrosis factor (TNF) family of proteins, where a ligand binds to one of the death receptors that subsequently recruit the death-inducing signaling complex (DISC) and the apical caspase 8 (4,5). Activation of apical caspases then leads to cleavage and activation of the effector caspases 3, 6, and 7 and subsequently to apoptosis (6). Some cell SMIP004 types produce insufficient amounts of active caspase 8 at the receptor level. Thus, the induction of apoptosis via the extrinsic pathway requires additional cleavage of the proapoptotic Bcl-2 family member BID through caspase 8, which activates the intrinsic pathway and subsequently apoptosis (type II cell mechanism) (7,8). For decades, chemotherapy was considered to be the only approach in the treatment of patients with advanced or accelerated CLL. Nonspecific cytotoxic drugs such as alkylating agents or purine analogs have commonly been used to induce apoptosis via the intrinsic pathway. The use of chemotherapy in CLL is limited, however, because of rapidly developing resistance and partial unresponsiveness of patients with p53 abnormalities (9,10). Therefore, the implementation of novel, highly selective, and less toxic agents gains more and more importance in the treatment of CLL. Valproic acid (VPA) is a short-chain fatty acid that belongs to a relatively new class of agents used for anticancer therapy, the histone deacetylase inhibitors (HDAC-I). VPA has been used as an anticonvulsant and mood-stabilizing drug for decades. Even when taken over a long time, VPA is usually well-tolerated, although it is contraindicated during pregnancy because of its teratogenic effects (11). HDAC-Is exert pleiotropic antitumor effects by inducing growth arrest, differentiation, and apoptosis, both in vitro and in vivo (12). Growth inhibition goes along with cell cycle arrest as a consequence of transcriptional activation of p21WAF1/CIP1 and p27KIP1 and/or inhibition of cyclin A, cyclin D, and thymidylate synthetase (13). In malignant cells, HDACIs induce apoptosis SMIP004 by upregulation of proapoptotic and repression of antiapoptotic genes (14C16). In this study, we investigated the effect of VPA on CLL cells ex vivo. It was our goal to further elucidate the effects of VPA on apoptosis in these SMIP004 cells. MATERIALS AND METHODS Patients and Control Group Peripheral blood samples were obtained from 50 patients (34 men, 16 women, median age 67.4 years) with B – C L L a n d f ro m 5 h e Mouse Monoclonal to Rabbit IgG a l t h y d o n o r s (2 men, 3 women, median age 32.2 years). Informed consent was given. The patient characteristics are given in Table 1. Table 1 Patient characteristics and individual response in vitro. was mea- with the TaqMan Gene Expression (Applied Biosystems). The ther- cycling conditions were 95C for 2 min, by 40 cycles at 95C for 15 s and for 1 min. Real-time PCR was per- in triplicate.