Synthetic Lethality of Combinatory Inhibition of BET and CDK7 in BET(i) Resistant Leukemia Cells
Authors: Guo Lei , Li Jia , Fang Shaohai , Lee Minjung , Zeng Hongxiang , Li Tingting , Han Wei , Sun Deqiang , Huang Yun 
Published: October 2019

Bromodomain and extra terminal proteins (BETs) act as non-oncogene addiction targets1,2. BET inhibitors (BETi) are epigenetic drugs being explored as a promising therapeutic avenue in cancer, especially acute myeloid leukemia (AML)3,4,5. Although early clinical trials have highlighted the efficacy of this first-in-class therapy, chemotherapy resistance, which is commonly emerged during drug treatment, still hurdle the prevalence of BETi6,7.

To seek for a strategy to overcome this hurdle, we carried out a combinatorial drug screening and discovered a synthetic lethality via combined inhibition of BRD4 and CDK7. Cyclin-dependent kinase 7 (CDK7), acting as a member of the general transcription factor H (TFIIH), can directly target the carboxyl-terminal domain (CTD) of the Rpb1 subunit of RNAPII for phosphorylation at serine 5 (Ser5) and serine 7 (Ser7), which is critical for transcription initiation. Recently, CDK7 has also been implicated in super-enhancer (SE) regulation8.

Dual pharmacological inhibition of BRD4 and CDK7 caused a synergistic effect on the growth and apoptosis of K562 cells. Knockdown of BRD4 and CDK7 in the same cell line showed similar results as drug treatment, thus ruling out the off-target effects of inhibitors used in this study. Furthermore, we confirmed this synergistic effect in vivo using a xenograft mouse model. After 2 weeks of treatment, mice bearing BET-resistant cells showed prolonged lifespan with less leukemic burden and reduced leukemic blast infiltration in spleen, liver, and bone marrow when treated with combinatorial regimen compared with control and single agent groups. Furthermore, RNA-seq analysis showed that genes involved in cell cycle and proliferation were significantly down-regulated, while genes associated with apoptosis were up-regulated. Myc-associated genes were prominently altered, suggesting that c-Myc is one of the top targets of BRD4 and CDK7 inhibition. Ectopically expressing c-Myc partially rescued the synthetic lethality induced by BRD4 and CDK7 treatment, a finding further confirmed that Myc is potential targets of inhibition of BRD4 and CDK7. Further molecular analysis showed that BRD4 and CDK7 inhibition altered the RNAPII activity at actively transcribed genes. Interestingly, we also observed a strong reduction of RNAPII enrichment, but not H3K27Ac enrichment, at super-enhancers (SE). Loci-specific experiment on c-Myc SE regions will further reveal detailed mechanisms on how BRD4 and CDK7 inhibition affect RNAPII activity at SE to influence gene transcription.

Congruently, by adopting a combinatorial drug screen approach, we identified the synthetic lethality of BRD4 and CDK7 inhibition in BRDi resistant leukemia cells both in vitro and in vivo. Our findings identify a new strategy that may enhance the clinical utility of BET inhibitors, especially for the treatment of BETi resistance.

Refereneces

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2. Shi, J. & Vakoc, C. R. The mechanisms behind the therapeutic activity of BET bromodomain inhibition. Mol. Cell 54, 728-736 (2014)

3. Herait, P. E. et al. BET-bromodomain inhibitor OTX015 shows clinically meaningful activity at nontoxic doses: interim results of an ongoing phase I trial in hematologic malignancies. Cancer Res.74, CT231 (2014)

4. Zuber, J. et al. RNAi screen identifies Brd4 as a therapeutic target in acute myeloid leukaemia. Nature 478, 524-528 (2011)

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6. Rathert, P. et al. Transcriptional plasticity promotes primary and acquired resistance to BET inhibition. Nature 525, 543-547 (2015)

7. Fong, C. Y. et al. BET inhibitor resistance emerges from leukaemia stem cells. Nature 525, 538-542 (2015)

8. E. Chipumuro, E. Marco, C.L. Christensen, et al. CDK7 inhibition suppresses super-enhancer-linked oncogenic transcription in MYCN-driven cancer. Cell, 159, 1126-1139 (2014)

Disclosures

No relevant conflicts of interest to declare.