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. 2024 Feb 10;16(4):741.
doi: 10.3390/cancers16040741.

Functional Homologous Recombination (HR) Screening Shows the Majority of BRCA1/2-Mutant Breast and Ovarian Cancer Cell Lines Are HR-Proficient

Titia G Meijer  1   2   3 , John W M Martens  4 , Wendy J C Prager-van der Smissen  4 , Nicole S Verkaik  1   3 , Corine M Beaufort  4 , Stanley van Herk  3   5 , Teresa Robert-Finestra  3   6 , Remco M Hoogenboezem  5 , Kirsten Ruigrok-Ritstier  4 , Maarten W Paul  1   3 , Joost Gribnau  3   6 , Eric M J Bindels  5 , Roland Kanaar  1   3 , Agnes Jager  4 , Dik C van Gent  1   3 , Antoinette Hollestelle  4
Affiliations

Functional Homologous Recombination (HR) Screening Shows the Majority of BRCA1/2-Mutant Breast and Ovarian Cancer Cell Lines Are HR-Proficient

Titia G Meijer et al. Cancers (Basel). .

Abstract

Tumors with a pathogenic BRCA1/2 mutation are homologous recombination (HR)-deficient (HRD) and consequently sensitive to platinum-based chemotherapy and Poly-[ADP-Ribose]-Polymerase inhibitors (PARPi). We hypothesized that functional HR status better reflects real-time HR status than BRCA1/2 mutation status. Therefore, we determined the functional HR status of 53 breast cancer (BC) and 38 ovarian cancer (OC) cell lines by measuring the formation of RAD51 foci after irradiation. Discrepancies between functional HR and BRCA1/2 mutation status were investigated using exome sequencing, methylation and gene expression data from 50 HR-related genes. A pathogenic BRCA1/2 mutation was found in 10/53 (18.9%) of BC and 7/38 (18.4%) of OC cell lines. Among BRCA1/2-mutant cell lines, 14/17 (82.4%) were HR-proficient (HRP), while 1/74 (1.4%) wild-type cell lines was HRD. For most (80%) cell lines, we explained the discrepancy between functional HR and BRCA1/2 mutation status. Importantly, 12/14 (85.7%) BRCA1/2-mutant HRP cell lines were explained by mechanisms directly acting on BRCA1/2. Finally, functional HR status was strongly associated with COSMIC single base substitution signature 3, but not BRCA1/2 mutation status. Thus, the majority of BRCA1/2-mutant cell lines do not represent a suitable model for HRD. Moreover, exclusively determining BRCA1/2 mutation status may not suffice for platinum-based chemotherapy or PARPi patient selection.

Keywords: BRCA1/2; breast cancer; cell lines; functional status; homologous recombination repair; mutation status; mutational signature; ovarian cancer.

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Conflict of interest statement

The authors declare no conflict of interest. The funder played no role in the conduct of the study, collection of data, management of the study, analysis of data, interpretation of data or preparation of the manuscript.

Figures

Figure 1
Figure 1
Functional HRD status of 53 breast cancer cell lines. (A) Quantification of the fraction of EdU-positive cells with ≥5 RAD51 foci. Each bar represents the mean value of three separate quantifications of the same cell line. Error bars indicate standard error of the mean (SEM). (B) Overview of the molecular and cellular characteristics of the cell lines. HR, homologous recombination; DDR, DNA damage response; SBS, single base substitution.
Figure 2
Figure 2
Functional HRD status of 38 ovarian cancer cell lines. (A) Quantification of the fraction of EdU-positive cells with ≥5 RAD51 foci. Each bar represents the mean value of three separate quantifications of the same cell line. Error bars indicate SEM. (B) Overview of the molecular and cellular characteristics of the cell lines. HR, homologous recombination; DDR, DNA damage response; SBS, single base substitution.
Figure 3
Figure 3
Promoter methylation levels of BRCA1, BRCA2, ATM and RAD51B. Graphs present square root-transformed methylation levels of 53 BC cell lines in decreasing order. (A) BRCA1 CpG19767, (B) BRCA1 CpG19770, (C) BRCA2 CpG11721, (D) ATM CpG8407 and (E) RAD51B CpG13662. Color of bars illustrates HR status: blue, HRP; orange, HRI and red, HRD. * indicates BRCA1/2 wild-type HRD cell line HCC2218.
Figure 4
Figure 4
Gene expression levels of RAD51B, PARG and SHLD2. Graphs present log2-transformed gene expression (GE) levels for 69 BC and OC cell lines in increasing order. (A) RAD51B, * indicates BRCA1/2 wild-type HRP cell line SUM190PT. (B) PARG and (C) SHLD2, * indicates three unexplained BRCA1-deficient HRP cell lines. Color of bars illustrates HR status: blue, HRP and red, HRD.
Figure 5
Figure 5
Correlation of SBS signature 3 with BRCA1/2 mutation and HR status. (A–C) Boxplots illustrate the difference in median percentage relative contribution of single base substitution signature 3 among the different groups.
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