Genome-wide CRISPR screens reveal cyclin C as synthetic survival target of BRCA2

Abstract

Poly (ADP-ribose) polymerase inhibitor (PARPi)-based therapies initially reduce tumor burden but eventually lead to acquired resistance in cancer patients with BRCA1 or BRCA2 mutation. To understand the potential PARPi resistance mechanisms, we performed whole-genome CRISPR screens to discover genetic alterations that change the gene essentiality in cells with inducible depletion of BRCA2. We identified that several RNA Polymerase II transcription Mediator complex components, especially Cyclin C (CCNC) as synthetic survival targets upon BRCA2 loss. Total mRNA sequencing demonstrated that loss of CCNC could activate the transforming growth factor (TGF)-beta signaling pathway and extracellular matrix (ECM)-receptor interaction pathway, however the inhibition of these pathways could not reverse cell survival in BRCA2 depleted CCNC-knockout cells, indicating that the activation of these pathways is not required for the resistance. Moreover, we showed that the improved survival is not due to restoration of homologous recombination repair although decreased DNA damage signaling was observed. Interestingly, loss of CCNC could restore replication fork stability in BRCA2 deficient cells, which may contribute to PARPi resistance. Taken together, our data reveal CCNC as a critical genetic determinant upon BRCA2 loss of function, which may help the development of novel therapeutic strategies that overcome PARPi resistance.

A CRISPR Path to Finding Vulnerabilities and Solving Drug Resistance: Targeting the Diverse Cancer Landscape and Its Ecosystem

“…[91] Moreover, using complementary genome-wide knockout and activation screens, Clements, et al, [93] revealed additional novel genetic determinants of PARP-i resistance, namely loss of ubiquitin ligase HUWE1, or the histone acetyltransferase KAT5, this time in the context of BRCA2-deficiency. These hits, with distinct roles in the regulation of end resection at DSBs and subsequent repair, [93] are joined by the TP53 induced glycolysis and apoptosis regulator (TIGAR) [94] and Cyclin C, [95] as further putative biomarkers and modifiers of treatment response to PARP-i, as a result of other recent CRISPR screening initiatives. Collectively, diverse alterations that tumors acquire while on PARP-i therapy may thus play important roles not only in signaling the emergence of treatment resistance but will likely also have important implications for the selection of subsequent rounds of therapy.…”

PICKLES v3: the updated database of pooled in vitro CRISPR knockout library essentiality screens

“…While the technology behind CRISPR-mediated genetic and epigenetic perturbation has evolved rapidly, by far the largest data sources remain the genome-scale CRISPR/Cas9 knockout screens in hundreds of cancer cell lines. Under the various banners of The Cancer Dependency Map (DepMap) (1), Project Achilles (2), and Project Score (3), as well as additional independent projects (4–10), roughly a thousand cancer and other human cell lines have been screened using whole-genome CRISPR/Cas9 knockout libraries.…”

“…In this PICKLES update we include knockout fitness data from three CRISPR libraries (Table 1), used in Project Achilles (Avana) and Project Score (Score) as well as several smaller independent screening projects that screened multiple cell lines (4–10) using the TKOv3 library (15) (Figure 1). We acquired raw read count data and processed each dataset with the BAGEL2 (13) pipeline, where essential genes have positive logBF scores.…”

PICKLES v3: the updated database of pooled in vitro CRISPR knockout library essentiality screens

“…While the technology behind CRISPR-mediated genetic and epigenetic perturbation has evolved rapidly, by far the largest data sources remain the genome-scale CRISPR/Cas9 knockout screens in hundreds of cancer cell lines. Under the various banners of The Cancer Dependency Map (DepMap) ( 1 ), Project Achilles ( 2 ) and Project Score ( 3 ), as well as additional independent projects ( 4–10 ), roughly a thousand cancer and other human cell lines have been screened using whole-genome CRISPR/Cas9 knockout libraries.…”

“…In this PICKLES update, we include knockout fitness data from three CRISPR libraries (Table 1 ), used in Project Achilles (Avana) and Project Score (Score) as well as several smaller independent screening projects that screened multiple cell lines ( 4–10 ) using the TKOv3 library ( 15 ) (Figure 1 ). We acquired raw read count data and processed each dataset with the BAGEL2 ( 13 ) pipeline, where essential genes have positive logBF scores.…”