Radiosensitization by the histone deacetylase inhibitor vorinostat under hypoxia and with capecitabine in experimental colorectal carcinoma

Background The histone deacetylase inhibitor vorinostat is a candidate radiosensitizer in locally advanced rectal cancer (LARC). Radiosensitivity is critically influenced by hypoxia; hence, it is important to evaluate the efficacy of potential radiosensitizers under variable tissue oxygenation. Since fluoropyrimidine-based chemoradiotherapy (CRT) is the only clinically validated regimen in LARC, efficacy in combination with this established regimen should be assessed in preclinical models before a candidate drug enters clinical trials. Methods Radiosensitization by vorinostat under hypoxia was studied in four colorectal carcinoma cell lines and in one colorectal carcinoma xenograft model by analysis of clonogenic survival and tumor growth delay, respectively. Radiosensitizing effects of vorinostat in combination with capecitabine were assessed by evaluation of tumor growth delay in two colorectal carcinoma xenografts models. Results Under hypoxia, radiosensitization by vorinostat was demonstrated in vitro in terms of decreased clonogenicity and in vivo as inhibition of tumor growth. Adding vorinostat to capecitabine-based CRT increased radiosensitivity of xenografts in terms of inhibited tumor growth. Conclusions Vorinostat sensitized colorectal carcinoma cells to radiation under hypoxia in vitro and in vivo and improved therapeutic efficacy in combination with capecitabine-based CRT in vivo. The results encourage implementation of vorinostat into CRT in LARC trials

Enriched biological processes in patients' peripheral blood mononuclear cells during 24 hours of vorinostat treatment.

a<p>Gene Ontology (GO) terms in bold: present in both comparisons.</p>b<p>Verified by reverse transcriptase quantitative polymerase chain reaction analysis.</p>c<p>T0 represents baseline peripheral blood mononuclear cells (PBMC) samples; T2 and T24 represent PBMC samples collected two and 24 hours, respectively, after the patients had received the daily dose of vorinostat.</p

Validation of vorinostat-regulated expression of selected genes.

<p>Study patients' peripheral blood mononuclear cells (PBMC) were sampled at baseline (T0) and on-treatment two (T2) and 24 (T24) hours after administration of the daily dose of the study medication vorinostat, and expression of <i>MYC</i>, <i>GADD45B</i>, <i>MSH6</i>, <i>BARD1</i>, and <i>DDIT3</i> was analyzed by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). Correspondingly, mice bearing HCT116 or SW620 xenografts were injected intraperitoneally with vehicle (control, C) or vorinostat, and xenografts were harvested three (T3) and 12 (T12) hours after injection for RT-qPCR analysis of <i>MYC</i> expression. Relative gene expression (log₂-fold change) for each comparison is given as mean ± SEM of the PBMC sample values (<i>n</i> = 14) and as median and range of the values from control (<i>n</i> = 8 for HCT116; <i>n</i> = 4 for SW620) and vorinostat-treated (<i>n</i> = 4 for HCT116; <i>n</i> = 2 for SW620) xenografts. The compared gene expression levels were significantly different within the PBMC (<i>P</i><0.01) and HCT116 (<i>P</i><0.05) sample groups, while the differences were non-significant for the SW620 tumors.</p

Venn diagram illustrating differentially expressed genes.

<p>Study patients' peripheral blood mononuclear cells were sampled at baseline (T0) and on-treatment two and 24 hours after administration of the daily dose of the study medication vorinostat (T2 and T24, respectively). Gene expression was analyzed by Illumina Human WG-6 v3 Expression BeadChip arrays. The array data from all patient samples at each time point (T0, T2, and T24) were pooled for the analysis. Probes with false discovery rate-adjusted <i>P</i>-values less than 0.05 were considered differentially expressed and subjected to Venn analysis, comparing by pairs T2 <i>versus</i> T0, T24 <i>versus</i> T2, and T24 <i>versus</i> T0. The figures represent numbers of probes in common for the various conditions.</p

Study patients.

a<p>Peripheral blood mononuclear cells.</p

Enriched biological pathways in patients' peripheral blood mononuclear cells during 24 hours of vorinostat treatment.

a<p>Genes in bold: verified by reverse transcriptase quantitative polymerase chain reaction analysis.</p