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Molecular disruption of RAD50 sensitizes human tumor cells to cisplatin-based chemotherapy.

ABSTRACT: Platinum-based drugs that induce DNA damage are commonly used first-line chemotherapy agents for testicular, bladder, head and neck, lung, esophageal, stomach, and ovarian cancers. The inherent resistance of tumors to DNA damage often limits the therapeutic efficacy of these agents, such as cisplatin. An enhanced DNA repair and telomere maintenance response by the Mre11/Rad50/Nbs1 (MRN) complex is critical in driving this chemoresistance. We hypothesized therefore that the targeted impairment of native cellular MRN function could sensitize tumor cells to cisplatin. To test this, we designed what we believe to be a novel dominant-negative adenoviral vector containing a mutant RAD50 gene that significantly downregulated MRN expression and markedly disrupted MRN function in human squamous cell carcinoma cells. A combination of cisplatin and mutant RAD50 therapy produced significant tumor cytotoxicity in vitro, with a corresponding increase in DNA damage and telomere shortening. In cisplatin-resistant human squamous cell cancer xenografts in nude mice, this combination therapy caused dramatic tumor regression with increased apoptosis. Our findings suggest the use of targeted RAD50 disruption as what we believe to be a novel chemosensitizing approach for cancer therapy in the context of chemoresistance. This strategy is potentially applicable to several types of malignant tumors that demonstrate chemoresistance and may positively impact the treatment of these patients.

SUBMITTER: Abuzeid WM

PROVIDER: S-EPMC2701852 | biostudies-other | 2009 Jul

REPOSITORIES: biostudies-other

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Molecular disruption of RAD50 sensitizes human tumor cells to cisplatin-based chemotherapy.

Abuzeid Waleed M WM Jiang Xiaoling X Shi Guoli G Wang Hui H Paulson David D Araki Koji K Jungreis David D Carney James J O'Malley Bert W BW Li Daqing D

The Journal of clinical investigation 20090701 7

Platinum-based drugs that induce DNA damage are commonly used first-line chemotherapy agents for testicular, bladder, head and neck, lung, esophageal, stomach, and ovarian cancers. The inherent resistance of tumors to DNA damage often limits the therapeutic efficacy of these agents, such as cisplatin. An enhanced DNA repair and telomere maintenance response by the Mre11/Rad50/Nbs1 (MRN) complex is critical in driving this chemoresistance. We hypothesized therefore that the targeted impairment of ...[more]

PMID: 19487811

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Project description:Simple Summary Muscle-invasive urothelial carcinoma of the bladder (UC) is treated with chemotherapies based on the DNA-damaging drug cisplatin, which only works temporarily due to the development of drug resistance. In this study, we show that it may be possible to overcome such resistances by treating the cancer cells with specific epigenetic drugs. We investigated the “epidrug” PLX51107 that inhibits the chromatin regulator BRD4 (Bromodomain Containing 4). PLX51107 inhibited cell growth, caused DNA damage, and blocked DNA repair response in UC cells. Concomitant application of PLX51107 with cisplatin or the drug talazoparib, interfering with DNA repair, caused cell death very efficiently. PLX51107 thus sensitizes UC cells to other drugs and may allow therapy with novel effective anti-tumor drugs like talazoparib that normally only work in a small proportion of patients with specific gene mutations. These results may help to improve current standard therapy and to develop new treatment options urgently required for UC patients. Abstract Muscle-invasive urothelial carcinoma (UC) is treated with cisplatin-based chemotherapy, which is only moderately efficient, mostly due to development of resistance. New therapy approaches are therefore urgently needed. Epigenetic alterations due to frequent mutations in epigenetic regulators contribute to development of the disease and to treatment resistance, and provide targets for novel drug combination therapies. Here, we determined the cytotoxic impact of the second-generation bromodomain protein inhibitor (BETi) PLX51107 on UC cell lines (UCC) and normal HBLAK control cells. PLX51107 inhibited proliferation, induced apoptosis, and acted synergistically with the histone deacetylase inhibitor romidepsin. While PLX51107 caused significant DNA damage, DNA damage signaling and DNA repair were impeded, a state defined as BRCAness. Accordingly, the drug strongly synergized with cisplatin more efficiently than romidepsin, and with the PARP inhibitor talazoparib to inhibit proliferation and induce cell death in UCC. Thus, a BETi can be used to “episensitize” UC cells to cytotoxic chemotherapy and inhibitors of DNA repair by inducing BRCAness in non BRCA1/2 mutated cancers. In clinical applications, the synergy between PLX51107 and other drugs should permit significant dosage reductions to minimize effects on normal tissues.

Project description:BACKGROUND: Worldwide, cervical cancer is the second most common causes of cancer in women and represents an important mortality rate. Cisplatin (CIS) is a very important antitumoral agent and can lead tumor cells toward two important cellular states: apoptosis and senescence. In some types of cancers pentoxifylline (PTX) sensitizes these cells to the toxic action of chemotherapeutics drugs such as adriamycin, inducing apoptosis. In the present work, we studied in vitro whether PTX alone or in combination with CIS induces apoptosis and/or senescence in cervix cancer HeLa and SiHa cell lines infected with HPV types 16 and 18, respectively, as well as in immortalized keratinocytyes HaCaT cells. METHODS: HeLa (HPV 18+), SiHa (HPV 16+) cervix cancer cells and non-tumorigenic immortalized HaCaT cells (control) were treated with PTX, CIS or both. The cellular toxicity and survival fraction of PTX and CIS were determinate by WST-1 and clonogenic assays respectively. Apoptosis, caspase activation and phosphorylation of ERK1/2, p38, p65 (NF-?B), Bcl-2 and Bcl-XL anti-apoptotic proteins were determinated by flow cytometry. Senescence by microscopy. Phosphorylation of I?B? and I?B total were measured by ELISA. Pro-apoptotic, anti-apoptotic and senescence genes, as well as HPV-E6/7 mRNA expression, were detected by RT-PCR. RESULTS: Our results show that after 24 hours of incubation PTX per se is toxic for cancer cells affecting cell viability and inducing apoptosis. The toxicity in HaCaT cells was minimal. CIS induces apoptosis in HeLa and SiHa cells and its effect was significantly increases when the cells were treated with PTX + CIS. In all studies there was a direct correlation with levels of caspases (-3, -6, -7, -9 and -8) activity and apoptosis. CIS induces important levels of senescence and phosphorylation of ERK1/2, p38, p65/RELA, and I?B?, and decreased the expression of anti-apoptotic protein Bcl-XL. Surprisingly these levels were significantly reduced by PTX in tumor cells, and at the same time, increases the expression of pro-apoptotic genes. CONCLUSION: PTX sensitizes cervical cancer cells to CIS-induced apoptosis and decreases the CIS-induced senescence in these cells via inhibition of NF-?B signaling pathway; diminishes expression of antiapoptotic proteins and the activation of caspases.

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Molecular disruption of RAD50 sensitizes human tumor cells to cisplatin-based chemotherapy.

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Molecular disruption of RAD50 sensitizes human tumor cells to cisplatin-based chemotherapy.

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