Unintended target effect of anti-BCL-2 DNAi
Authors Ebrahim AS, Kandouz M, Emara N, Sugalski AB, Lipovich L, Al-Katib AM
Received 7 April 2017
Accepted for publication 15 July 2017
Published 22 September 2017 Volume 2017:9 Pages 427—432
Checked for plagiarism Yes
Review by Single-blind
Peer reviewers approved by Dr Akshita Wason
Peer reviewer comments 2
Editor who approved publication: Dr Antonella D'Anneo
Abdul Shukkur Ebrahim,1 Mustapha Kandouz,2 Nada Emara,1 Amara B Sugalski,3 Leonard Lipovich,3 Ayad M Al-Katib1
1Lymphoma Research Laboratory, 2Department of Pathology, School of Medicine, 3Center for Molecular Medicine and Genetics, Wayne State University, Detroit, MI, USA
Introduction: Previous research suggested that a novel compound PNT2258 inhibits B-cell lymphoma 2 (BCL-2) transcription by DNA interference (DNAi) and demonstrated its activity in preclinical xenograft models and in a pilot Phase II clinical trial in non-Hodgkin’s lymphoma (NHL). While the drug downregulates BCL-2 at the promoter, mRNA, and protein levels, there is a significant homology (13–16 bases) between PNT100 and a number of promoters of genes involved in cell cycle regulation and survival. In this study, we identify cyclin-dependent kinase-4 (CDK4) as an unintended target gene of PNT2258 and examine its relevance to NHL.
Methods: We performed a Basic Local Alignment Search Tool (BLAST) homology search using PNT100 DNAi sequences. Also, we conducted CDK4 promoter assay in K562 cells and studied the protein expression of CDK4 in Wayne State University (WSU)-follicular small cleaved cell lymphoma (FSCCL), WSU-diffuse large cell lymphoma, and WSU-Waldenström’s macroglobulinemia (WM) lymphoma cells.
Results: BLAST homology search showed that PNT100 completely binds to BCL-2 gene as expected. However, there was 100% homology in a stretch of 14 bases (8–21) between PNT100 and CDK4. PNT2258 strongly inhibited CDK4 promoter activity in K562 cells. Moreover, CDK4 protein expression was significantly downregulated by PNT2258 in WSU-FSCCL and WSU-WM cell lines.
Discussion: DNAi may work not only through knocking down the intended gene but also by knocking down other genes. PNT2258 affects CDK4 expression and promoter activity. Results of the present study suggest a broader mechanism of action for DNAi targeting both intended (BCL-2) and unintended (CDK4) genes.
Keywords: non-Hodgkin’s lymphoma, BCL-2, PNT2258, BLAST, CDK4
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