Gene expression profiling of homologous recombination repair pathway indicates susceptibility for olaparib treatment in malignant pleural mesothelioma in vitro

Abstract

Background: Malignant pleural mesothelioma (MPM) is a tumour arising from pleural cavities with poor prognosis. Multimodality treatment with pemetrexed combined with cisplatin shows unsatisfying response-rates of 40%. The reasons for the rather poor efficacy of chemotherapeutic treatment are largely unknown. However, it is conceivable that DNA repair mechanisms lead to an impaired therapy response. We hypothesize a major role of homologous recombination (HR) for genome stability and survival of this tumour. Therefore, we analysed genes compiled under the term "BRCAness". An inhibition of this pathway with olaparib might abrogate this effect and induce apoptosis.

Methods: We investigated the response of three MPM cell lines and lung fibroblasts serving as a control to treatment with pemetrexed, cisplatin and olaparib. Furthermore, we aimed to find possible correlations between response and gene expression patterns associated with BRCAness phenotype. Therefore, 91 clinical MPM samples were digitally screened for gene expression patterns of HR members.

Results: A BRCAness-dependent increase of apoptosis and senescence during olaparib-based treatment of BRCA-associated-protein 1 (BAP1)-mutated cell lines was observed. The gene expression pattern identified could be found in approx. 10% of patient samples. Against this background, patients could be grouped according to their defects in the HR system. Gene expression levels of Aurora Kinase A (AURKA), RAD50 as well as DNA damage-binding protein 2 (DDB2) could be identified as prognostic markers in MPM.

Conclusions: Defects in HR compiled under the term BRCAness are a common event in MPM. The present data can lead to a better understanding of the underlaying cellular mechanisms and leave the door wide open for new therapeutic approaches for this severe disease with infaust prognosis. Response to Poly (ADP-ribose)-Polymerase (PARP)-Inhibition could be demonstrated in the BAP1-mutated NCI-H2452 cells, especially when combined with cisplatin. Thus, this combination therapy might be effective for up to 2/3 of patients, promising to enhance patients' clinical management and outcome.

Keywords: BRCAness - BAP1; Malignant pleural mesothelioma - overall survival; Olaparib; PARP1.

Fig. 1

Enhanced base excision repair, due to defective HRR that is caused by BRCAness phenotype, increases the reliance on PARP1. It is suggested that loss-of-function mutation of BAP1 also results in BRCAness phenotype. Inhibition of PARP1 prevents the alternative repair pathway and thus could lead to apoptosis of the cell

Fig. 2

Senescence and apoptosis rate of cell lines after 48 h of incubation. a: The effect of both senescence and apoptosis is comparative in MRC-5. b: In MSTO-211H, the sharp increase of apoptotic effect of 181% of pemetrexed alone and in combination with cisplatin is illustrated, while senescence showed an effect of 100%. The well treated with 10 μM olaparib showed a senescence effect of 80%, while no apoptotic effect was detected. c: NCI H2052 cells showed apoptotic effects only in wells treated with cisplatin or in combination with pemetrexed or olaparib and senescence of 30–50%. Wells treated with 1 μM and 10 μM olaparib showed 40% of senescence, while no apoptotic effect was detected. d: NCI H2452 cells showed 70% of apoptosis and only 20% of senescence in wells treated with 0.5 μM, 1 μM or 10 μM olaparib combined with cisplatin. 10 μM Olaparib alone showed 15% higher senescence than apoptosis

Fig. 3

Comparison of significant differences in gene expression level between cell lines with respect to response to treatment with olaparib. Red dotted lines were placed and may represent thresholds between gene expression patterns leading to response to olaparib treatment or not. a: BAP1 was rarely expressed in the cell line that showed response to olaparib treatment (NCI H2452), while it was expressed in other cell lines that showed no response (160 to 250 counts). Threshold was set at 140 counts. b: PARP1 was expressed in NCI H2452 with 350 counts, but other cell lines showed significantly higher expression patterns with 640–785 counts. Threshold was set at 600 counts. c/d: BRCA2 and CHEK2 are expressed in the cell line that showed response to olaparib (NCI H2452), while no expression was detected in cell lines that showed no response. Thresholds were set at 100 counts for BRCA2 and CHEK2. e/f: AURKA and RPA1 are more expressed in cell lines that showed no response, than in NCI H2452. Thresholds were set at 400 counts for AURKA and 540 counts for RPA1

Fig. 4

Overall correlation pattern between samples. Unsupervised clustering revealed two distinct groups. The group in the yellow box includes all samples with altered HRR. This box also includes expression patterns of olaparib-responsive NCI-H2052 as well as BAP1-mutated NCI-H2452 cells

Fig. 5

Overall and progression-free survival dependent on gene expression of AURKA, RAD50 and DDB2. Low expression of AURKA and high expression of RAD50 and DDB2 resulted in prolonged overall survival (A-C). Low expression of AURKA (D) also resulted in prolonged progression-free survival with p < 0.0030. Low expression is highlighted in red, high expression is highlighted in green. The median was used to set a cut-off between high and low gene expression