Enhanced Apoptosis to Chemotherapeutic Agents Is Dependent on NFκB and Bcl2-Related Proteins but Is Independent of P53 and Bax in Burkitt's Lymphoma Cells.

Abstract

Deregulated expression of c-Myc is involved in the pathogenesis and growth of Burkitt's lymphoma cells. Aberrant NFκB activation has been linked to several forms of cancers, including Hodgkin's lymphoma, breast cancers, and B-cell malignancies. The cytoplasmic retention of NFκB is mediated by a family of inhibitor proteins, the IκBs. We showed that expression of a super-repressor form of IκBα, IκBα-SR, resulted in decreased proliferation and an increased G0/G1 population of Raji Burkitt's cells. No spontaneous apoptosis was observed, but there was increased sensitivity to apoptosis following treatment with chemotherapeutic agents. Further studies were performed to determine the mechanisms involved in the induction of apoptosis. Interference with NFκB activity decreased the levels of c-Myc and Bcl-xL, but there was no change in the levels of Bax, Bak, Mcl-1, or p53. Treatment with chemotherapeutic agents alone resulted in decreased expression of Mcl-1 with no change in the other proteins measured. Decreased expression of both Bcl-xL and Mcl-1 following inhibition of NFκB activity and chemotherapy (vincristine, 4HC or doxorubicin) treatment resulted in apoptosis. In support of the important role of Bcl-xL and Mcl-1, siRNA experiments revealed that decreased expression of either one alone did not lead to apoptosis, while decreased expression of both Bcl-xL and Mcl-1 resulted in the induction of apoptosis. Many Burkitt's lymphoma cells have mutations in p53. We found that the mutated p53 was able to bind to IκBα and Iκ Bα-SR as well as to Bcl-xL, however, no up-regulation of Bax or p53 was observed in response to a chemotherapeutic agent. This indicates that mutated p53 retains the ability to bind the other proteins but has lost transcriptional function. Introduction of high levels of wild-type p53 into Raji Burkitt's cells was able to restore the induction of Bax and Mdm2 and induce apoptosis, whereas the mutant was not. These results suggest that sensitization of Burkitt's cells to apoptotic death with chemotherapeutic agents requires interference with NFκB activity and changes in the expression of Bcl2-family members but is independent of p53. Inhibition of NFκB activity could be useful in combination with chemotherapy for the treatment of Burkitt's lymphoma.