Polo-like Kinase 1 Inhibition as a Therapeutic Approach to Selectively Target BRCA1-Deficient Cancer Cells by Synthetic Lethality Induction

Abstract

Purpose: BRCA1 and BRCA2 deficiencies are widespread drivers of human cancers that await the development of targeted therapies. We aimed to identify novel synthetic lethal relationships with therapeutic potential using BRCA-deficient isogenic backgrounds.Experimental Design:We developed a phenotypic screening technology to simultaneously search for synthetic lethal (SL) interactions in BRCA1-and BRCA2-deficient contexts. For validation, we developed chimeric spheroids and a dualtumor xenograft model that allowed the confirmation of SL induction with the concomitant evaluation of undesired cytotoxicity on BRCA-proficient cells. To extend our results using clinical data, we performed retrospective analysis on The Cancer Genome Atlas (TCGA) breast cancer database.Results: The screening of a kinase inhibitors library revealed that Polo-like kinase 1 (PLK1) inhibition triggers strong SL induction in BRCA1-deficient cells. Mechanistically, we found no connection between the SL induced by PLK1 inhibition and PARP inhibitors. Instead, we uncovered that BRCA1 downregulation and PLK1 inhibition lead to aberrant mitotic phenotypes with altered centrosomal duplication and cytokinesis, which severely reduced the clonogenic potential of these cells. The penetrance of PLK1/BRCA1 SL interaction was validated using several isogenic and nonisogenic cellular models, chimeric spheroids, and mice xenografts. Moreover, bioinformatic analysis revealed high-PLK1 expression in BRCA1-deficient tumors, a phenotype that was consistently recapitulated by inducing BRCA1 deficiency in multiple cell lines as well as in BRCA1-mutant cells.Conclusions: We uncovered an unforeseen addiction of BRCA1-deficient cancer cells to PLK1 expression, which provides a new means to exploit the therapeutic potential of PLK1 inhibitors in clinical trials, by generating stratification schemes that consider this molecular trait in patient cohorts.

Persistent double strand break accumulation does not precede cell death in an Olaparib-sensitive BRCA-deficient colorectal cancer cell model

“…Control for mycoplasma contamination was performed periodically with a PCR-based method with internal loading control. To generate cell lines expressing fluorescent proteins, all the fluorescent proteins were expressed from the same backbone from Clontech as de-scribed in Carbajosa et al (2019). Briefly, transfection of vectors encoding fluorescent proteins (piRFP-C1, pECFP-C1, pmCherry-C1) was performed using JetPrime (Polyplus-transfection) according to manufacturer's instructions.…”

“…Multiple rounds of cell sorting (3-5) were performed (FACS Aria II, BD Bioscience) to select pools that express CFP, iRFP, and mCherry fluorescent proteins. Each "colored" cell line was sub-sequentially used to generate shSCR, shBRCA1, and shBRCA2 expressing pools (Carbajosa et al, 2019). The transduced cells that showed the higher downregulation of BRCA1 and BRCA2 by qPCR and WB, yet keeping similar proliferation rates to the shSCR-transduced cell lines, were selected for survival assays.…”

“…In order to achieve an efficient knockdown of BRCA proteins, we set up an shRNA-based downregulation of BRCA1 or BRCA2. Many cell lines did not recover after an acute crisis, and other cell lines regained BRCA expression after a few passages (Carbajosa et al, 2019). We then explored colorectal cancer cells that are not characterized by a high frequency of BRCA loss.…”

Advances in synthetic lethality for cancer therapy: cellular mechanism and clinical translation

“…Multiple studies have used bioinformatic screening to identify synthetic lethal pathway that can theoretically be used to monitor disease progression [82]. The predictions are produced by scoring gene pair interaction by using expression profiles data, screening results for shRNA, and copy number data [83].…”

Development of synthetic lethality in cancer: molecular and cellular classification

“…Although numerous small-molecule and antibody-based drugs for oncogenes or tumor-suppressor genes have proven to be effective for several tumors with certain gene mutations, 31 not all oncogenes or tumor-suppressor genes could be targeted and resistance is common, 7 In such cases, identifying and exploiting a second or several other functional genes that interact with the primary oncogene or tumor-suppressor gene provides an alternative method for cancer treatment. Therefore, SL is increasingly being explored recently, in an effort to identify new anticancer therapeutic targets through large-scale SL screening in model organisms and human cell lines such as NSCLC (NCI-H1355, NCI-H1299, NCI-H1155), hepatocellular carcinoma (HCC1954, HCC1937, HCC1806), and breast cancer (MDA-MB-468, MDA-MB-436, MDA-MB-415) via clustered regularly interspaced short palindromic repeats (CRISPR), 32 tumor genomic sequence database, RNA interference (RNAi) technology, 33 , 34 etc. The most remarkable finding in SL is the hypersensitivity of BRCA1/2-mutant tumor cells to poly-(ADP-ribose) polymerase (PARP) inhibitors.…”