Genomic Basis for Resistance to BCL-2 Inhibition in Hematologic Malignancies
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Liu, Vivian M.
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CitationLiu, Vivian M. 2019. Genomic Basis for Resistance to BCL-2 Inhibition in Hematologic Malignancies. Doctoral dissertation, Harvard Medical School.
AbstractB-cell chronic lymphocytic leukemia (CLL) is the most common adult leukemia in Western countries. The standard frontline therapy is a combination immune-chemotherapy, but relapse remains the rule for this disease. The treatment landscape of CLL and other B-cell malignancies is changing dramatically. Recently the FDA has approved the use of a BCL-2 inhibitor, venetoclax, to treat a subset of patients with relapsed CLL. Although this drug had a high response rate in clinical trials, clinical progression has been noted due to the development of drug resistance. Therefore, understanding the resistance mechanisms to this drug is a priority to glean maximum benefit from its use and to implement more personalized treatment strategies.
To study the mechanisms of venetoclax resistance in malignant B-cells, we used a genome-scale CRISPR-Cas9 screen of B-cell lines to determine the specific genes that are necessary for sensitivity to the drug. Our screen resulted in a total of 11 genes that are required for sensitivity to BCL-2 inhibition with venetoclax. Four of these genes are well-known pro-apoptotic genes in the apoptosis pathway (PMAIP1, BAX, BAK1, and BCL2L11). The other seven genes have relatively unknown function with respect to BCL-2 and apoptosis, and they have known functions of being transcription factors, transcriptional modulators, and ubiquitin modifiers. Isogenic cell lines with these genes knocked-out have confirmed resistance to venetoclax with increased IC50 values and increased rate of proliferation under drug treatment.
Transcriptomic and proteomic analysis of a lymphoma cell line with induced resistance to venetoclax demonstrated upregulation of MCL-1 and downregulation of ID2. Comparison transcriptomes of a resistant line and isogenic cell lines revealed the ID3 knockout as the most similar to the resistant cell line. Further analysis revealed a resistance circuit involving ID3 repression and increased AMPK and PKA signaling as mechanisms of resistance in cell lines. Targeting AMPK and the mitochondrial electron transport chain (mETC) resulted in sensitization of cells to venetoclax. Whole exome sequencing of our resistant cell line along with pre- and post-venetoclax samples from 6 patients revealed a common amplification of chromosome 1q23 in both the resistant cell line and 3 of the 6 patients. This region contained the genes MCL1 and PRKAB2. Immunohistochemistry studies revealed increased expression of MCL1 and AMPK signaling components in patient samples. These results provide greater insight into mechanisms of resistance to venetoclax and suggest MCL-1 inhibitors, AMPK inhibitors, and mETC modulators as possibilities for combination therapies to combat resistance.
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