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Vitamin D supplementation prior to surgery for colorectal cancer: a randomised pilot study.

ABSTRACT: Interventions: Colorectal cancer patients scheduled to undergo routine elective surgery at Dunedin Hospital will be randomised to receive either a single oral 5mg dose of vitamin D3 (cholecalciferol), or identical placebo to be taken 7-21 days prior to surgery. A dose of 5mg of vitamin D3 will consistently achieve therapeutic serum levels >80nmol/L, peaking at 1 week and gradually falling thereafter. The dose is safe and designed to rapidly achieve desirable levels without delaying surgery. All patients enrolled will recieve all other treatment and care as they would have otherwise done so. Primary outcome(s): A significant increase in the expression of genes containing the Vitamin D responsive element (VDRE), in tumour tissue, in patients receiving active study medication compared to placebo. Previously published data on the effects of vitamin D on gene expression in a colorectal cancer cell line will be used to identify genes that are responsive to vitamin D. Affymetrix microarray data from these experiments are available from the NCBI GEO database under the accession number GSE444. In addition, genes representing vitamin D targets in normal colon tissues will be identified by searching publically available colon array databases for expressed genes which contain the VDRE. The VDRE element is present within the promoter region of genes activated by vitamin D and therefore acts as a convenient tag to identify vitamin D responsive genes. RNA will be isolated from CRC samples from 50 patients, half of whom will have received vitamin D. RNA expression profiles of the 50 tumour RNA samples will then be generated in the Otago Genomics Facility using Affymetrix HG-U133+2.0 GeneChips. Evidence of activation of vitamin D pathways in the tumours from treated and untreated patients will be determined using the vitamin D responsive genes identified above. This will be accomplished by generating a vitamin D meta-gene to represent the coordinated expression of these genes across all tumours, so that tumours exhibiting a molecular response to vitamin D will have high levels of the meta-gene. This meta-gene will then be tested for association with treatment status, as well as clinicopathological (e.g. tumour stage, histology etc), molecular (e.g. proliferation) and immune reponse variables.[Tissue samples will be collected on the day of surgery. These will be batched and analysed in the laboratory within 6 months of the final patients surgery.];A significant difference between treatment groups with respect to macrophage activation against tumour tissue and their ability to prime T cells. Macrophages isolated from the blood and tumour tissue of CRC patients will be infected with bacterial pathogens in vitro and their ability to destroy these agents measured by quantifying nitric oxide production using the Greiss reaction. Their ability to prime the adaptive immune response will also be measured by analysing activation of T cells recovered from patient tissues using flow cytometry of surface activation markers. Results will be compared between treatment and placebo groups.[Tissue samples will be collected on the day of surgery. These will be batched and analysed in the laboratory within 6 months of the final patients surgery.] Study Design: Purpose: Treatment; Allocation: Randomised controlled trial; Masking: Blinded (masking used);Assignment: Parallel;Type of endpoint: Bio-availability

DISEASE(S): Colorectal Cancer,Cancer-bowel-back Passage (rectum) Or Large Bowel (colon)

PROVIDER: 2456306 | ecrin-mdr-crc |

REPOSITORIES: ECRIN MDR

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Project description:Abstract Background: Several studies on the use of erythropoietin (Epo) to treat anaemia in patients undergoing cancer treatment have shown adverse effects on tumour control and survival. Experimental studies indicate that this could be linked to an interaction with wound healing processes and not an effect on tumour cells per se. We have previously shown that erythropoietin in combination with surgical trauma stimulates tumour growth. In the present study, we investigated the effect of surgery and Epo on gene expression. Methods: Human tumours from oral squamous cell cancer were xenotransplanted to nude mice treated with Epo. The tumours were then transected in a standardised procedure to mimic surgical trauma and the change in gene expression of the tumours was investigated by microarray analysis. qRT-PCR was used to measure the levels of mRNAs of pro-apoptotic genes. The frequency of apoptosis in the tumours was assessed using immunohistochemistry for caspase-3. Results: There was little change in the expression of genes involved in tumour growth and angiogenesis but a significant down-regulation of the expression of genes involved in apoptosis. This effect on apoptosis was confirmed by a general decrease in the expression of mRNA for selected pro-apoptotic genes. Epo-treated tumours had a significantly lower frequency of apoptosis as measured by immunohistochemistry for caspase 3. Conclusions: Our results suggest that the increased tumour growth during erythropoietin treatment might be due to inhibition of apoptosis, an effect that becomes significant during tissue damage such as surgery. This further suggests that the decreased tumour survival during erythropoietin treatment might be due to inhibition of apoptosis. Key words: Erythropoietin, Head and neck cancer, surgery, apoptosis, wound healing, xenograft. We wanted to study the effect of wound healing mechanisms on remaining tumour tissue efter incomplete surgery or biopsy. We xenografted human Head and Neck squamous Cell Cancer (HNSCC) tumours on Balb/c nude mice. The mice were divided in six groups. Three of the groups received intrapertoneal doses of erythropoietin (Epo) and the other three groups were given NaCl as placebo. One treated and one non-treated group were harvested without surgical transection. In the other four groups, the tumours were transected mimicking subtotal surgery. One Epo-treated and one non-treated group was analysed after 24 hours after transection and the remaining two groups 48 hours after surgery. Groups C and D were not analyzed due to harvesting at an unsuitable time interval. Group A No surgery (=FALSE) + Epo Group B No surgery (=FALSE) + NaCl Group E Surgery (=TRUE) + Epo 24 hours after surgical transection Group F Surgery (=TRUE) + NaCl 24 hours after surgical resection Group G Surgery (=TRUE) + Epo 48 hours after surgical transection Group H Surgery (=TRUE) + Nacl 48 hours after surgical transection

Project description:AIM Vitamin D deficiency is associated with risk of several common cancers, including colorectal cancer (CRC). We investigated the effects of vitamin D on in vitro and ex vivo epithelial cell gene expression and assessed dysregulation of pathways involved in anti-tumour effects. METHODS We performed in vitro and ex vivo intervention studies using established CRC cell lines and patient-derived epithelial organoids developed from normal human colonic (resectional colorectal surgery) and rectal mucosa (rectal biopsy). RNA and protein were harvested after 16-24 hours’ calcitriol treatment. Gene expression was assessed by qRT-PCR, microarray and RNA-sequencing, and protein expression by immunoblotting and mass-spectrometry. Gene Ontology enrichment analysis was performed using GOrilla and Gene Set Enrichment Analysis. RESULTS Calcitriol increased the expression of the tumour suppressor gene CDH1 in CRC cell lines (SW480; FC=3.60, 95%CI 2.17-5.03) and patient-derived epithelial cell organoids (n=6, FC=1.64, 95%CI 1.49-1.80), with associated in vitro E-cadherin protein induction. Significant differential expression of a further 6 common genes was observed on microarray analysis of in vitro and ex vivo experiments, including genes with established links to carcinogenesis GADD45, EFTUD1 and KIAA1199. Numerous ontologies relevant to carcinogenesis were enriched (e.g. ‘regulation of Wnt signaling pathway’, ‘regulation of cell death’), with common enriched processes across in vitro and ex vivo cultures including ‘negative regulation of cell proliferation’, ‘regulation of cell migration’ and ‘regulation of cell differentiation’. CONCLUSIONS We report for the first time, common genes and pathways modulated by calcitriol treatment in both in vitro and ex vivo models, with several differentially expressed genes strongly linked to CRC tumourigenesis. Enrichment of biological pathways after calcitriol treatment provides further evidence of anti-tumour effects. These findings support epidemiological data and provide strong rationale for well-designed trials of vitamin D supplementation as a novel CRC chemopreventive and chemotherapeutic agent. see above

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Vitamin D supplementation prior to surgery for colorectal cancer: a randomised pilot study.

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