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Synergistic antitumour activity of HDAC inhibitor SAHA and EGFR inhibitor gefitinib in head and neck cancer: a key role for ΔNp63α.

ABSTRACT:

Background

Epidermal growth factor receptor (EGFR) overexpression is associated with the development of head and neck cancer (HNC) and represents one of the main therapeutic targets for this disease. The use of EGFR inhibitors has limited efficacy due to their primary and acquired resistance, partially because of increased epithelial to mesenchymal transition (EMT). The HDAC inhibitor SAHA has been shown to revert EMT in different tumours, including HNC. In this study, we investigated the cooperative role of SAHA and the EGFR tyrosine kinase inhibitor gefitinib in both HPV-positive and HPV-negative HNC cell lines.

Methods

A panel of 12 HPV-positive and HPV-negative HNC cell lines were screened for cell viability upon treatment with SAHA, gefitinib and the combination of the two. Epithelial/mesenchymal marker expression, as well as activation of signalling pathway, were assessed upon SAHA treatment. ΔNp63α silencing with shRNA lentiviral particles was used to determine its role in cell proliferation, migration and TGFβ pathway activation.

Results

We found that both SAHA and gefitinib have antitumour activity in both HPV-positive and HPV-negative HNC cell lines and that their combination has a synergistic effect in inhibiting cell growth. SAHA treatment reverts EMT and inhibits the expression of the transcription factor ΔNp63α. Suppression of ΔNp63α reduces EGFR protein levels and decreases cell proliferation and TGFβ-dependent migration in both HPV-positive and HPV-negative HNC cell lines.

Conclusions

Our results, by giving a clear molecular mechanism at the basis of the antitumour activity of SAHA in HNC cell lines, provide a rationale for the clinical evaluation of SAHA in combination with gefitinib in both HPV-positive and HPV-negative HNC patients. Further knowledge is key to devising additional lines of combinatorial treatment strategies for this disease.

SUBMITTER: Citro S

PROVIDER: S-EPMC6461861 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND:The poly ADP ribose polymerase (PARP) inhibitor olaparib has been approved for treating prostate cancer (PCa) with BRCA mutations, and veliparib, another PARP inhibitor, is being tested in clinical trials. However, veliparib only showed a moderate anticancer effect, and combination therapy is required for PCa patients. Histone deacetylase (HDAC) inhibitors have been tested to improve the anticancer efficacy of PARP inhibitors for PCa cells, but the exact mechanisms are still elusive. METHODS:Several types of PCa cells and prostate epithelial cell line RWPE-1 were treated with veliparib or SAHA alone or in combination. Cell viability or clonogenicity was tested with violet crystal assay; cell apoptosis was detected with Annexin V-FITC/PI staining and flow cytometry, and the cleaved PARP was tested with western blot; DNA damage was evaluated by staining the cells with ?H2AX antibody, and the DNA damage foci were observed with a fluorescent microscopy, and the level of ?H2AX was tested with western blot; the protein levels of UHRF1 and BRCA1 were measured with western blot or cell immunofluorescent staining, and the interaction of UHRF1 and BRCA1 proteins was detected with co-immunoprecipitation when cells were treated with drugs. The antitumor effect of combinational therapy was validated in DU145 xenograft models. RESULTS:PCa cells showed different sensitivity to veliparib or SAHA. Co-administration of both drugs synergistically decreased cell viability and clonogenicity, and synergistically induced cell apoptosis and DNA damage, while had no detectable toxicity to normal prostate epithelial cells. Mechanistically, veliparib or SAHA alone reduced BRCA1 or UHRF1 protein levels, co-treatment with veliparib and SAHA synergistically reduced BRCA1 protein levels by targeting the UHRF1/BRCA1 protein complex, the depletion of UHRF1 resulted in the degradation of BRCA1 protein, while the elevation of UHRF1 impaired co-treatment-reduced BRCA1 protein levels. Co-administration of both drugs synergistically decreased the growth of xenografts. CONCLUSIONS:Our studies revealed that the synergistic lethality of HDAC and PARP inhibitors resulted from promoting DNA damage and inhibiting HR DNA damage repair pathways, in particular targeting the UHRF1/BRCA1 protein complex. The synergistic lethality of veliparib and SAHA shows great potential for future PCa clinical trials.