Depletion of the transcriptional coactivators CREB-binding protein or EP300 downregulates CD20 in diffuse large B-cell lymphoma cells and impairs the cytotoxic effects of anti-CD20 antibodies

Abstract Monoclonal antibodies targeting CD20 are central in the treatment of B-cell lymphomas. In diffuse large B-cell lymphoma (DLBCL), inactivating mutations of the histone acetyltransferases CREB-binding protein (CBP) and EP300 are common. Moreover, knockdown of CBP in DLBCL has been shown to result in aberrant transcriptional silencing. Expression of CD20 is sensitive to epigenetic manipulation, and histone deacetylase inhibitors has been shown to potentiate treatment with anti-CD20 antibodies. Therefore, we studied the role of CBP and EP300 depletion on CD20 expression and effects of the anti-CD20 antibodies rituximab and obinutuzumab in DLBCL cells. Levels of CBP and EP300 were reduced by shRNA in the germinal centre-derived diffuse large B-cell lymphoma cell line SU-DHL4. The levels of CD20 mRNA and protein were determined by qPCR, Western blot and flowcytometry. Binding of the transcription factors PU.1 and FOXO1 to the CD20 promoter was determined by ChIP-qPCR. Response to the monoclonal anti-CD20 antibodies rituximab and obinutuzumab in CBP or EP300 depleted cells was assessed by complement dependent cell death (CDC), direct cell death (DCD), and antibody-dependent cellular cytotoxicity (ADCC). Our results suggest that depletion of CBP and EP300 levels leads to a strong reduction of CD20 expression, accompanied by reduced binding of PU.1 to the CD20 promoter. In CBP depleted, but not EP300 depleted, cells an increased binding of FOXO1 to the CD20 promoter was observed. Interestingly, CBP or EP300 depletion leads to decreased CDC and DCD in response to rituximab and obinutuzumab, which was most pronounced in response to rituximab in CBP depleted cells. Our data suggest that inactivating mutations of CBP, and to a lesser extent EP300, may impair the response to anti-CD20 antibodies. However, these observations should be analyzed in future clinical trials.