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Melphalan-flufenamide is cytotoxic and potentiates treatment with chemotherapy and the Src inhibitor dasatinib in urothelial carcinoma.

ABSTRACT: BACKGROUND:Chemotherapy options in advanced urothelial carcinoma (UC) remain limited. Here we evaluated the peptide-based alkylating agent melphalan-flufenamide (mel-flufen) for UC. METHODS:UC cell lines J82, RT4, TCCsup and 5637 were treated with mel-flufen, alone or combined with cisplatin, gemcitabine, dasatinib or bestatin. Cell viability (MTT assay), intracellular drug accumulation (liquid chromatography) apoptosis induction (apoptotic cell nuclei morphology, western blot analysis of PARP-1/caspase-9 cleavage and Bak/Bax activation) were evaluated. Kinome alterations were characterized by PathScan array and phospho-Src validated by western blotting. Aminopeptidase N (ANPEP) expression was evaluated in UC clinical specimens in relation to patient outcome. RESULTS:In J82, RT4, TCCsup and 5637 UC cells, mel-flufen amplified the intracellular loading of melphalan in part via aminopeptidase N (ANPEP), resulting in increased cytotoxicity compared to melphalan alone. Mel-flufen induced apoptosis seen as activation of Bak/Bax, cleavage of caspase-9/PARP-1 and induction of apoptotic cell nuclei morphology. Combining mel-flufen with cisplatin or gemcitabine in J82 cells resulted in additive cytotoxic effects and for gemcitabine also increased apoptosis induction. Profiling of mel-flufen-induced kinome alterations in J82 cells revealed that mel-flufen alone did not inhibit Src phosphorylation. Accordingly, the Src inhibitor dasatinib sensitized for mel-flufen cytotoxicity. Immunohistochemical analysis of the putative mel-flufen biomarker ANPEP demonstrated prominent expression levels in tumours from 82 of 83 cystectomy patients. Significantly longer median overall survival was found in patients with high ANPEP expression (P = 0.02). CONCLUSION:Mel-flufen alone or in combination with cisplatin, gemcitabine or Src inhibition holds promise as a novel treatment for UC.

SUBMITTER: Viktorsson K

PROVIDER: S-EPMC5423156 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature

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Melphalan-flufenamide is cytotoxic and potentiates treatment with chemotherapy and the Src inhibitor dasatinib in urothelial carcinoma.

Viktorsson Kristina K Shah Carl-Henrik CH Juntti Therese T Hååg Petra P Zielinska-Chomej Katarzyna K Sierakowiak Adam A Holmsten Karin K Tu Jessica J Spira Jack J Kanter Lena L Lewensohn Rolf R Ullén Anders A

Molecular oncology 20160102 5

<h4>Background</h4>Chemotherapy options in advanced urothelial carcinoma (UC) remain limited. Here we evaluated the peptide-based alkylating agent melphalan-flufenamide (mel-flufen) for UC.<h4>Methods</h4>UC cell lines J82, RT4, TCCsup and 5637 were treated with mel-flufen, alone or combined with cisplatin, gemcitabine, dasatinib or bestatin. Cell viability (MTT assay), intracellular drug accumulation (liquid chromatography) apoptosis induction (apoptotic cell nuclei morphology, western blot ana ...[more]

PMID: 26827254

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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.

Dataset Information

Melphalan-flufenamide is cytotoxic and potentiates treatment with chemotherapy and the Src inhibitor dasatinib in urothelial carcinoma.

Publications

Melphalan-flufenamide is cytotoxic and potentiates treatment with chemotherapy and the Src inhibitor dasatinib in urothelial carcinoma.

Similar Datasets

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