A Small Interfering RNA Screen of Genes Involved in DNA Repair Identifies Tumor-Specific Radiosensitization by POLQ Knockdown

Abstract

The effectiveness of radiotherapy treatment could be significantly improved if tumour cells could be rendered more sensitive to ionizing radiation without altering the sensitivity of normal tissues. However many of the key, therapeutically exploitable mechanisms that determine intrinsic tumour radiosensitivity are largely unknown. We have conducted a siRNA screen of 200 genes involved in DNA damage repair aimed at identifying genes whose knockdown increased tumour radiosensitivity. Parallel siRNA screens were conducted in irradiated and unirradiated tumour cells (SQ20B) and irradiated normal tissue cells (MRC5). Using γH2AX foci at 24 hours after ionising radiation we identified several genes such as BRCA2, Lig IV and XRCC5, whose knockdown is known to cause increased cell radiosensitivity thereby validating the primary screening endpoint. In addition we identified POLQ (DNA polymerase theta) as a potential tumour-specific target. Subsequent investigations demonstrated that POLQ knockdown resulted in radiosensitisation of a panel of tumour cell lines from different primary sites, whilst having little or no effect on normal tissue cell lines. These findings raise the possibility that POLQ inhibition might be used clinically to cause tumour specific radiosensitisation.

A Polymerase Theta-dependent repair pathway suppresses extensive genomic instability at endogenous G4 DNA sites

“…We anticipate that ectopic activation of TMEJ may be of high clinical significance, also considering the recent finding that Polymerase Theta upregulation is associated with poor survival in cancer 35,36 ; if not properly controlled, POL y's ability to tie DNA ends together can have very undesirable effects, as it provides cells with the means to proliferate in the presence of increased replication stress 37 . Blocking this activity may thus constitute a potent strategy towards preventing cancerous growth.…”

Depletion of RUVBL2 in Human Cells Confers Moderate Sensitivity to Anticancer Agents

“…After treatment with various DNA-damaging agents, RUVBL1 is required for the dephosphorylation of phosphorylated histone H2AX, a marker of DNA double-strand breaks (DSBs), through the histone acetyltransferase activity of TIP60 [8]. Likewise, depletion of RUVBL2 increases the persistence of phosphorylated histone H2AX upon treatment with X-rays [10]. Apart from the pathway involving the TIP60 complex, RUVBLs regulate the abundance and activity of phosphatidylinositol 3-kinases such as ataxia telangiectasia mutated (ATM), ATM-and Rad3-related (ATR) kinase, and DNA-dependent protein kinase (DNA-PK) that sense and activate the DNA damage signal [11].…”

The therapeutic ratio is preserved for radiotherapy or cisplatin treatment in BRCA2-mutated prostate cancers

“…Although the data are varied, some studies have suggested that HR-defective cell lines and xenografts can be more sensitive to ionizing radiation (IR), cisplatin, mitomycin C (MMC), etoposides, melphalan and poly(ADP-ribose) polymerase inhibitors (PARPi). 6,7 These cell lines, however, have also been reported to be variably sensitive or even resistant to taxane-based chemotherapy (e.g., docetaxel). [8][9][10][11][12][13][14][15][16][17] These data suggest that patients with a BRCA2 mutation with tumours defective in HR may be targeted by agents, such as RT, cisplatin, anthracyclines or PARPi.…”