Enhancing synthetic lethality of PARP-inhibitor and cisplatin in BRCA-proficient tumour cells with hyperthermia

Background: Poly-(ADP-ribose)-polymerase1 (PARP1) is involved in repair of DNA single strand breaks. PARP1-inhibitors (PARP1-i) cause an accumulation of DNA double strand breaks, which are generally repaired by homologous recombination (HR). Therefore, cancer cells harboring HR deficiencies are exceptionally sensitive to PARP1-i. For patients with HR-proficient tumors, HR can be temporarily inhibited by hyperthermia, thereby inducing synthetic lethal conditions in every tumor type. Since cisplatin is successfully used combined with hyperthermia (thermochemotherapy), we investigated the effectiveness of combining PARP1-i with thermochemotherapy. Results: The in vitro data demonstrate a decreased in cell survival after addition of PARP1-i to thermochemotherapy, which can be explained by increased DNA damage induction and less DSB repair. These in vitro findings are in line with in vivo model, in which a decreased tumor growth is observed upon addition of PARP1-i. Materials and Methods: Survival of three HR-proficient cell lines after cisplatin, hyperthermia and/or PARP1-i was studied. Cell cycle analyses, quantification of γ-H2AX foci and apoptotic assays were performed to understand these survival data. The effects of treatments were further evaluated by monitoring tumor responses in an in vivo rat model. Conclusions: Our results in HR-proficient cell lines suggest that PARP1-i combined with thermochemotherapy can be a promising clinical approach for all tumors independent of HR status

Enhancing radiosensitisation of BRCA2-proficient and BRCA2-deficient cell lines with hyperthermia and PARP1-i

Poly(ADP-ribose)polymerase1 (PARP1) is an important enzyme in regulating DNA replication. Inhibition of PARP1 can lead to collapsed DNA forks which subsequently causes genomic instability, making DNA more susceptible in developing fatal DNA double strand breaks. PARP1-induced DNA damage is generally repaired by homologous recombination (HR), in which BRCA2 proteins are essential. Therefore, BRCA2-deficient tumour cells are susceptible to treatment with PARP1-inhibitors (PARP1-i). Recently, BRCA2 was shown to be down-regulated by hyperthermia (HT) temporarily, and this consequently inactivated HR for several hours. In this study, we investigated whether HT exclusively interferes with HR by analysing thermal radiosensitisation of BRCA2-proficient and deficient cells. After elucidating the equitoxicity of PARP1-i on BRCA2-proficient and deficient cells, we studied the cell survival, apoptosis, DNA damage (γ-H2AX foci and comet assay) and cell cycle distribution after different treatments. PARP1-i sensitivity strongly depends on the BRCA2 status. BRCA2-proficient and deficient cells are radiosensitised by HT, indicating that HT does not exclusively act by inhibition of HR. In all cell lines, the addition of HT to radiotherapy and PARP1-i resulted in the lowest cell survival, the highest levels of DNA damage and apoptotic levels compared to duo-modality treatments. Concluding, HT not only inhibits HR, but also has the capability of radiosensitising BRCA2-deficient cells. Thus, in case of BRCA2-mutation carriers, combining HT with PARP1-i may boost the treatment efficacy. This combination therapy would be effective for all patients with PARP1-i regardless of their BRCA statu

Hyperthermia Selectively Targets Human Papillomavirus in Cervical Tumors via p53-Dependent Apoptosis

Human papillomavirus (HPV) is associated with cervical cancer, the third most common cancer in women. The high-risk HPV types 16 and 18 are found in over 70% of cervical cancers and produce the oncoprotein, early protein 6 (E6), which binds to p53 and mediates its ubiquitination and degradation. Targeting E6 has been shown to be a promising treatment option to eliminate HPV-positive tumor cells. In addition, combined hyperthermia with radiation is a very effective treatment strategy for cervical cancer. In this study, we examined the effect of hyperthermia on HPV-positive cells using cervical cancer cell lines infected with HPV 16 and 18, in vivo tumor models, and ex vivo-treated patient biopsies. Strikingly, we demonstrate that a clinically relevant hyperthermia temperature of 42 °C for 1 hour resulted in E6 degradation, thereby preventing the formation of the E6-p53 complex and enabling p53-dependent apoptosis and G2-phase arrest. Moreover, hyperthermia combined with p53 depletion restored both the cell-cycle distribution and apoptosis to control levels. Collectively, our findings provide new insights into the treatment of HPV-positive cervical cancer and suggest that hyperthermia therapy could improve patient outcome