Vitamin D opposes multilineage cell differentiation induced by Notch inhibition and BMP4 pathway activation in human colon organoids.

Understanding the mechanisms involved in colonic epithelial differentiation is key to unraveling the alterations causing inflammatory conditions and cancer. Organoid cultures provide an unique tool to address these questions but studies are scarce. We report a differentiation system toward enterocytes and goblet cells, the two major colonic epithelial cell lineages, using colon organoids generated from healthy tissue of colorectal cancer patients. Culture of these organoids in medium lacking stemness agents resulted in a modest ultrastructural differentiation phenotype with low-level expression of enterocyte (KLF4, KRT20, CA1, FABP2) and goblet cell (TFF2, TFF3, AGR2) lineage markers. BMP pathway activation through depletion of Noggin and addition of BMP4 resulted in enterocyte-biased differentiation. Contrarily, blockade of the Notch pathway using the γ-secretase inhibitor dibenzazepine (DBZ) favored goblet cell differentiation. Combination treatment with BMP4 and DBZ caused a balanced strong induction of both lineages. In contrast, colon tumor organoids responded poorly to BMP4 showing only weak signals of cell differentiation, and were unresponsive to DBZ. We also investigated the effects of 1α,25-dihydroxyvitamin D3 (calcitriol) on differentiation. Calcitriol attenuated the effects of BMP4 and DBZ on colon normal organoids, with reduced expression of differentiation genes and phenotype. Consistently, in normal organoids, calcitriol inhibited early signaling by BMP4 as assessed by reduction of the level of phospho-SMAD1/5/8. Our results show that BMP and Notch signaling play key roles in human colon stem cell differentiation to the enterocytic and goblet cell lineages and that calcitriol modulates these processes favoring stemness features.

Clinico-Pathological Features, Outcomes and Impacts of COVID-19 Pandemic on Patients with Early-Onset Colorectal Cancer: A Single-Institution Experience.

BACKGROUND: The rising incidence of colorectal cancer (CRC) among young patients is alarming. We aim to characterize the clinico-pathological features and outcomes of patients with early-onset CRC (EOCRC), as well as the impacts of COVID-19 pandemic. METHODS: We included all patients with pathologically confirmed diagnoses of CRC at Hospital Universitario La Paz from October 2016 to December 2021. The EOCRC cut-off age was 50 years old. RESULTS: A total of 1475 patients diagnosed with CRC were included, eighty (5.4%) of whom had EOCRC. Significant differences were found between EOCRC and later-onset patients regarding T, N stage and metastatic presentation at diagnosis; perineural invasion; tumor budding; high-grade tumors; and signet ring cell histology, with all issues having higher prevalence in the early-onset group. More EOCRC patients had the RAS/ BRAF wild type. Chemotherapy was administered more frequently to patients with EOCRC. In the metastatic setting, the EOCRC group presented a significantly longer median OS. Regarding the COVID-19 pandemic, more patients with COVID-19 were diagnosed with metastatic disease (61%) in the year after the lockdown (14 March 2020) than in the pre-pandemic EOCRC group (29%). CONCLUSIONS: EOCRC is diagnosed at a more advanced stage and with worse survival features in localized patients. More patients with EOCRC were diagnosed with metastatic disease in the year after the COVID-19 pandemic lockdown. The long-term consequences of COVID-19 are yet to be determined.

From molecular basis to clinical insights: a challenging future for the vitamin D endocrine system in colorectal cancer.

Colorectal cancer (CRC) is one of the most life-threatening neoplasias in terms of incidence and mortality worldwide. Vitamin D deficiency has been associated with an increased risk of CRC. 1α,25-Dihydroxyvitamin D3 [1,25(OH)2 D3 ], the most active vitamin D metabolite, is a pleiotropic hormone that, through its binding to a transcription factor of the nuclear receptor superfamily, is a major regulator of the human genome. 1,25(OH)2 D3 acts on colon carcinoma and stromal cells and displays tumor protective actions. Here, we review the variety of molecular mechanisms underlying the effects of 1,25(OH)2 D3 in CRC, which affect multiple processes that are dysregulated during tumor initiation and progression. Additionally, we discuss the epidemiological data that associate vitamin D deficiency and CRC, and the most relevant randomized controlled trials of vitamin D3 supplementation conducted in both healthy individuals and CRC patients.

Wnt and Vitamin D at the Crossroads in Solid Cancer.

Abnormal activation of the Wnt/ß-catenin pathway is common in many types of solid cancers. Likewise, a large proportion of cancer patients have vitamin D deficiency. In line with these observations, Wnt/ß-catenin signaling and 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3), the active vitamin D metabolite, usually have opposite effects on cancer cell proliferation and phenotype. In recent years, an increasing number of studies performed in a variety of cancer types have revealed a complex crosstalk between Wnt/ß-catenin signaling and 1,25(OH)2D3. Here we review the mechanisms by which 1,25(OH)2D3 inhibits Wnt/ß-catenin signaling and, conversely, how the activated Wnt/ß-catenin pathway may abrogate vitamin D action. The available data suggest that interaction between Wnt/ß-catenin signaling and the vitamin D system is at the crossroads in solid cancers and may have therapeutic applications.

Vitamin D and Wnt3A have additive and partially overlapping modulatory effects on gene expression and phenotype in human colon fibroblasts.

The Wnt/ß-catenin signalling pathway is essential for intestinal epithelium homeostasis, but its aberrant activation is a hallmark of colorectal cancer (CRC). Several studies indicate that the bioactive vitamin D metabolite 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) inhibits proliferation and promotes epithelial differentiation of colon carcinoma cells in part through antagonism of the Wnt/ß-catenin pathway. It is now accepted that stromal fibroblasts are crucial in healthy and pathologic intestine: pericryptal myofibroblasts are constituents of the stem cell niche and cancer-associated fibroblasts (CAFs) contribute to CRC progression. However, studies on the combined action of 1,25(OH)2D3 and Wnt factors in colon fibroblasts are lacking. Here we show by global transcriptomic studies that 1,25(OH)2D3 and Wnt3A have profound, additive, partially overlapping effects on the gene expression profile of CCD-18Co human colon myofibroblasts. Moreover, 1,25(OH)2D3 and Wnt3A inhibit CCD-18Co cell proliferation and migration, while 1,25(OH)2D3 reduces, but Wnt3A increases, their capacity to contract collagen gels (a marker of fibroblast activation). These data were largely confirmed in patient-derived primary colon normal fibroblasts and CAFs, and in fibroblasts from other origins. Our results indicate that 1,25(OH)2D3 and Wnt3A are strong regulators of colon fibroblast biology and contribute to a better knowledge of intestinal homeostasis and stromal fibroblast action in CRC.

The human PKP2/plakophilin-2 gene is induced by Wnt/ß-catenin in normal and colon cancer-associated fibroblasts.

Colorectal cancer results from the malignant transformation of colonic epithelial cells. Stromal fibroblasts are the main component of the tumour microenvironment, and play an important role in the progression of this and other neoplasias. Wnt/ß-catenin signalling is essential for colon homeostasis, but aberrant, constitutive activation of this pathway is a hallmark of colorectal cancer. Here we present the first transcriptomic study on the effect of a Wnt factor on human colonic myofibroblasts. Wnt3A regulates the expression of 1,136 genes, of which 662 are upregulated and 474 are downregulated in CCD-18Co cells. A set of genes encoding inhibitors of the Wnt/ß-catenin pathway stand out among those induced by Wnt3A, which suggests that there is a feedback inhibitory mechanism. We also show that the PKP2 gene encoding the desmosomal protein Plakophilin-2 is a novel direct transcriptional target of Wnt/ß-catenin in normal and colon cancer-associated fibroblasts. PKP2 is induced by ß-catenin/TCF through three binding sites in the gene promoter and one additional binding site located in an enhancer 20 kb upstream from the transcription start site. Moreover, Plakophilin-2 antagonizes Wnt/ß-catenin transcriptional activity in HEK-293T cells, which suggests that it may act as an intracellular inhibitor of the Wnt/ß-catenin pathway. Our results demonstrate that stromal fibroblasts respond to canonical Wnt signalling and that Plakophilin-2 plays a role in the feedback control of this effect suggesting that the response to Wnt factors in the stroma may modulate Wnt activity in the tumour cells.

Nuclear DICKKOPF-1 as a biomarker of chemoresistance and poor clinical outcome in colorectal cancer.

Sporadic colorectal cancer (CRC) insurgence and progression depend on the activation of Wnt/ß-catenin signaling. Dickkopf (DKK)-1 is an extracellular inhibitor of Wnt/ß-catenin signaling that also has undefined ß-catenin-independent actions. Here we report for the first time that a proportion of DKK-1 locates within the nucleus of healthy small intestine and colon mucosa, and of CRC cells at specific chromatin sites of active transcription. Moreover, we show that DKK-1 regulates several cancer-related genes including the cancer stem cell marker aldehyde dehydrogenase 1A1 (ALDH1A1) and Ral-binding protein 1-associated Eps domain-containing 2 (REPS2), which are involved in detoxification of chemotherapeutic agents. Nuclear DKK-1 expression is lost along CRC progression; however, it remains high in a subset (15%) of CRC patients (n = 699) and associates with decreased progression-free survival (PFS) after chemotherapy administration and overall survival (OS) [adjusted HR, 1.65; 95% confidence interval (CI), 1.23-2.21; P = 0.002)]. Overexpression of ALDH1A1 and REPS2 associates with nuclear DKK-1 expression in tumors and correlates with decreased OS (P = 0.001 and 0.014) and PFS. In summary, our findings demonstrate a novel location of DKK-1 within the cell nucleus and support a role of nuclear DKK-1 as a predictive biomarker of chemoresistance in colorectal cancer.

WNT-3A regulates an Axin1/NRF2 complex that regulates antioxidant metabolism in hepatocytes.

AIMS: Nuclear factor (erythroid-derived 2)-like 2 (NRF2) is a master regulator of oxidant and xenobiotic metabolism, but it is unknown how it is regulated to provide basal expression of this defense system. Here, we studied the putative connection between NRF2 and the canonical WNT pathway, which modulates hepatocyte metabolism. RESULTS: WNT-3A increased the levels of NRF2 and its transcriptional signature in mouse hepatocytes and HEK293T cells. The use of short interfering RNAs in hepatocytes and mouse embryonic fibroblasts which are deficient in the redox sensor Kelch-like ECH-associated protein 1 (KEAP1) indicated that WNT-3A activates NRF2 in a ß-Catenin- and KEAP1-independent manner. WNT-3A stabilized NRF2 by preventing its GSK-3-dependent phosphorylation and subsequent SCF/ß-TrCP-dependent ubiquitination and proteasomal degradation. Axin1 and NRF2 were physically associated in a protein complex that was regulated by WNT-3A, involving the central region of Axin1 and the Neh4/Neh5 domains of NRF2. Axin1 knockdown increased NRF2 protein levels, while Axin1 stabilization with Tankyrase inhibitors blocked WNT/NRF2 signaling. The relevance of this novel pathway was assessed in mice with a conditional deletion of Axin1 in the liver, which showed upregulation of the NRF2 signature in hepatocytes and disruption of liver zonation of antioxidant metabolism. INNOVATION: NRF2 takes part in a protein complex with Axin1 that is regulated by the canonical WNT pathway. This new WNT-NRF2 axis controls the antioxidant metabolism of hepatocytes. CONCLUSION: These results uncover the participation of NRF2 in a WNT-regulated signalosome that participates in basal maintenance of hepatic antioxidant metabolism.

Interaction of vitamin D with membrane-based signaling pathways.

Many studies in different biological systems have revealed that 1α,25-dihydroxyvitamin D3 (1α,25(OH)2D3) modulates signaling pathways triggered at the plasma membrane by agents such as Wnt, transforming growth factor (TGF)-ß, epidermal growth factor (EGF), and others. In addition, 1α,25(OH)2D3 may affect gene expression by paracrine mechanisms that involve the regulation of cytokine or growth factor secretion by neighboring cells. Moreover, post-transcriptional and post-translational effects of 1α,25(OH)2D3 add to or overlap with its classical modulation of gene transcription rate. Together, these findings show that vitamin D receptor (VDR) cannot be considered only as a nuclear-acting, ligand-modulated transcription factor that binds to and controls the transcription of target genes. Instead, available data support the view that much of the complex biological activity of 1α,25(OH)2D3 resides in its capacity to interact with membrane-based signaling pathways and to modulate the expression and secretion of paracrine factors. Therefore, we propose that future research in the vitamin D field should focus on the interplay between 1α,25(OH)2D3 and agents that act at the plasma membrane, and on the analysis of intercellular communication. Global analyses such as RNA-Seq, transcriptomic arrays, and genome-wide ChIP are expected to dissect the interactions at the gene and molecular levels.

Vitamin D Is a Multilevel Repressor of Wnt/b-Catenin Signaling in Cancer Cells.

The Wnt/b-catenin signaling pathway is abnormally activated in most colorectal cancers and in a proportion of other neoplasias. This activation initiates or contributes to carcinogenesis by regulating the expression of a large number of genes in tumor cells. The active vitamin D metabolite 1a,25-dihydroxyvitamin D3 (1,25(OH)2D3) inhibits Wnt/b-catenin signaling by several mechanisms at different points along the pathway. Additionally, paracrine actions of 1,25(OH)2D3 on stromal cells may also repress this pathway in neighbouring tumor cells. Here we review the molecular basis for the various mechanisms by which 1,25(OH)2D3 antagonizes Wnt/b-catenin signaling, preferentially in human colon carcinoma cells, and the consequences of this inhibition for the phenotype and proliferation rate. The effect of the vitamin D system on Wnt/b-catenin signaling and tumor growth in animal models will also be commented in detail. Finally, we revise existing data on the relation between vitamin D receptor expression and vitamin D status and the expression of Wnt/b-catenin pathway genes and targets in cancer patients.

The use of an active learning approach to teach metabolism to students of nutrition and dietetics.

This article describes the transition from a traditional instructor-centered course, based on lectures, to a student-centered course based on active learning methodologies as part of the reform of the Spanish higher education system within the European Higher Education Area (EHEA). Specifically, we describe the use of active learning methodologies to teach metabolism to students of nutrition and dietetics during the first year of their professional training in a 4-year undergraduate degree (Bachelor of Human Nutrition and Dietetics). In the new course design, the number of didactic lectures was largely reduced and complemented with a series of activities (problems/case studies, discussion workshops, self-assessment quizzes) aimed to get students actively engaged, to encourage self-learning, and to promote sustained work throughout the length of the course. The article presents quantitative data demonstrating a clear and significant improvement in students' performance when an active approach was implemented. Importantly, the improved performance was achieved without work overload. Finally, students' responses to this new teaching methodology have been very positive and overall satisfaction high. In summary, our results strongly argue in favor of the teaching model described herein.

Epigenetic repression of ROR2 has a Wnt-mediated, pro-tumourigenic role in colon cancer.

BACKGROUND: Wnt factors control cell differentiation through semi-independent molecular cascades known as the beta-catenin-dependent (canonical) and -independent (non-canonical) Wnt signalling pathways. Genetic and epigenetic alteration of components of the canonical Wnt signalling pathway is one of the primary mechanisms underlying colon cancer. Despite increasing evidence of the role of the non-canonical pathways in tumourigenesis, however, the underlying molecular mechanisms are poorly understood. RESULTS: Here we report that the receptor tyrosine kinase-like orphan receptor 2 (ROR2), a transmembrane receptor for Wnt factors that activates non-canonical pathways, is frequently repressed by aberrant promoter hypermethylation in human colon cancer cell lines and primary tumours. By restoring ROR2 activity in colon cancer cells harbouring ROR2 promoter hypermethylation, we show that the role of ROR2 in colon cancer cells is mediated, at least in part, by canonical Wnt and that its epigenetic-dependent loss can be pro-tumourigenic. CONCLUSIONS: Our data show the importance of epigenetic alterations of ROR2 in colon cancer, highlighting the close interconnection between canonical and non-canonical Wnt signalling pathways in this type of tumour.

Vitamin D and Wnt/beta-catenin pathway in colon cancer: role and regulation of DICKKOPF genes.

Colorectal cancer is a major health problem worldwide. Aberrant activation of the Wingless-type mouse mammary tumour virus integration site family (Wnt)/beta-catenin signalling pathway due to mutation of adenomatous polyposis coli (APC), beta-catenin (CTNNB1) or AXIN genes is the most common and initial alteration in sporadic colorectal tumours. Numerous epidemiological and experimental studies have indicated a protective action of vitamin D against colorectal cancer. Previous work has demonstrated that the most active vitamin D metabolite, 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3) inhibits beta-catenin transcriptional activity by promoting vitamin D receptor (VDR) binding to beta-catenin and the induction of E-cadherin expression. Recently, 1,25(OH)2D3 has been shown to distinctly regulate two genes encoding the extracellular Wnt inhibitors DICKKOPF-1 and DICKKOPF-4 (DKK-1, DKK-4). By an indirect transcriptional mechanism, 1,25(OH)2D3 increases the expression of DKK-1 RNA and protein, which acts as a tumour suppressor in human colon cancer cells harbouring endogenous mutations in the Wnt/beta-catenin pathway. In contrast, 1,25(OH)2D3 represses DKK-4 transcription by inducing direct VDR binding to its promoter. Unexpectedly, DKK-4 is a target of the Wnt/beta-catenin pathway and is up-regulated in colorectal tumours, and it has been shown to increase cell migration and invasion and to promote a proangiogenic phenotype. Together, these results show that 1,25(OH)2D3 exerts a complex set of regulatory actions leading to the inhibition of the Wnt/beta-catenin pathway in colon cancer cells that is in line with its protective effect against this neoplasia.

The Wnt antagonist DICKKOPF-1 gene is induced by 1alpha,25-dihydroxyvitamin D3 associated to the differentiation of human colon cancer cells.

The Wnt-beta-catenin pathway is aberrantly activated in most colon cancers. DICKKOPF-1 (DKK-1) gene encodes an extracellular Wnt inhibitor that blocks the formation of signalling receptor complexes at the plasma membrane. We report that 1alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3], the most active vitamin D metabolite, increases the level of DKK-1 RNA and protein in human SW480-ADH colon cancer cells. This effect is dose dependent, slow and depends on the presence of a transcription-competent nuclear vitamin D receptor (VDR). Accordingly, 1,25(OH)2D3 activates a 2300 bp fragment of the human DKK-1 gene promoter. Chromatin immunoprecipitation assays revealed that 1,25(OH)2D3 treatment induced a pattern of histone modifications which is compatible with transcriptionally active chromatin. DKK-1 is expressed at high level in colon cancer cell lines with a differentiated phenotype such as Caco-2 or HT-29. Exogenous expression of E-cadherin into SW480-ADH cells results in a strong adhesive phenotype and a 17-fold increase in DKK-1 RNA. In contrast, an E-cadherin blocking antibody inhibits 1,25(OH)2D3-induced differentiation of SW480-ADH cells and DKK-1 gene expression. Remarkably, in vivo treatment with the vitamin D analogue EB1089 induced DKK-1 protein expression in SW480-ADH cells xenografted in immunodeficient mice, and a correlation was observed in the expression of VDR and DKK-1 RNA in a series of 32 human colorectal tumours. These data indicate that 1,25(OH)2D3 activates the transcription of the DKK-1 gene, probably in an indirect way that is associated to the promotion of a differentiated phenotype. DKK-1 gene induction constitutes a novel mechanism of inhibition of Wnt signalling and antitumour action by 1,25(OH)2D3.

The inhibition of Wnt/beta-catenin signalling by 1alpha,25-dihydroxyvitamin D3 is abrogated by Snail1 in human colon cancer cells.

The Wnt/beta-catenin signalling pathway is activated in 90% of human colon cancers by nuclear accumulation of beta-catenin protein due to its own mutation or to that of adenomatous polyposis coli. In the nucleus, beta-catenin regulates gene expression promoting cell proliferation, migration and invasiveness. 1alpha,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) inhibits beta-catenin signalling by inducing its binding to vitamin D receptor (VDR) and by promoting beta-catenin nuclear export. The transcription factor Snail1 represses VDR expression and we demonstrate here that Snail1 also abolishes the nuclear export of beta-catenin induced by 1,25(OH)(2)D(3) in SW480-ADH cells. Accordingly, Snail1 relieves the inhibition exerted by 1,25(OH)(2)D(3) on genes whose expression is driven by beta-catenin, such as c-MYC, ectodermal-neural cortex-1 (ENC-1) or ephrin receptor B2 (EPHB2). In addition, Snail1 abrogates the inhibitory effect of 1,25(OH)(2)D(3) on cell proliferation and migration. In xenografted mice, Snail1 impedes the nuclear export of beta-catenin and the inhibition of ENC-1 expression induced by EB1089, a 1,25(OH)(2)D(3) analogue. The elevation of endogenous SNAIL1 protein levels reproduces the effect of an ectopic Snail1 gene. Remarkably, the expression of exogenous VDR in cells with high levels of Snail1 normalizes the transcriptional responses to 1,25(OH)(2)D(3). However, this exogenous VDR failed to fully restore the blockage of the Wnt/beta-catenin pathway by 1,25(OH)(2)D(3). This suggests that the effects of Snail1 on this pathway are not merely due to the repression of VDR gene. We conclude that Snail1 is a positive regulator of the Wnt/beta-catenin signalling pathway in part through the abrogation of the inhibitory action of 1,25(OH)(2)D(3).

Vitamin D regulates the phenotype of human breast cancer cells.

1alpha,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), the most active vitamin D metabolite, regulates proliferation, survival, and differentiation in many cell types. 1,25(OH)(2)D(3) and several less calcemic analogs are in clinical trials against various neoplasias. We studied the effects of 1,25(OH)(2)D(3) on a panel of human breast cancer cells, which show similar vitamin D receptor (VDR) content but variable transcriptional and anti-proliferative responsiveness. In MDA-MB-453 cells, one of the responsive lines, 1,25(OH)(2)D(3) increased cell and nuclear size and induced a change from a rounded to a flattened morphology. By phase contrast, laser confocal and electron microscopy, we found that 1,25(OH)(2)D(3) changed the cytoarchitecture of actin filaments and microtubules and nuclear shape, induced filopodia and lamellipodia, and promoted cell-to-cell contacts via large cytoplasmic extensions. However, although claudin-7 and occludin content in the cells increased upon exposure to 1,25(OH)(2)D(3), these proteins were not located at the plasma membrane probably due to the absence of E-cadherin expression. Additionally, 1,25(OH)(2)D(3) induced the accumulation of alpha(v)-integrin, beta(5)-integrin, focal adhesion kinase (FAK), and paxillin in focal adhesion plaques, concomitant with the increased phosphorylation of the FAK. 1,25(OH)(2)D(3) enhanced MDA-MB-453 and MDA-MB-468 cell adhesion to plastic but decreased adhesion to laminin. The expression of the mesenchymal marker N-cadherin and of the myoepithelial marker P-cadherin was down-regulated by 1,25(OH)(2)D(3) in several breast cancer cell lines. Other myoepithelial proteins such as alpha(6)-integrin, beta(4)-integrin, and smooth muscle alpha-actin (SMA) were also repressed by 1,25(OH)(2)D(3) in MDA-MB-453 and MDA-MB-468 cells. Accordingly, mice lacking VDR (Vdr(-/-)) showed abnormally high levels of SMA and P-cadherin in their mammary gland. These findings show that 1,25(OH)(2)D(3) profoundly affects the phenotype of breast cancer cells, and suggest that it reverts the myoepithelial features associated with more aggressive forms and poor prognosis in human breast cancer.

Effects of 1alpha,25-dihydroxyvitamin D3 in human colon cancer cells.

Colorectal cancer is a major health problem worldwide. Epidemiological studies and work on experimental animals strongly suggest a protective effect of 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3) against colon neoplasia. 1,25(OH)2D3 is a pleiotropic hormone that has multiple actions in the organism. By binding to the widely expressed high affinity vitamin D receptor (VDR) it regulates the transcription rate of many genes. Other non-genomic effects of 1,25(OH)2D3 also appear to modulate the physiology of numerous cell types. Human normal and cancer colon epithelial cells express VDR and the key enzymes involved in 1,25(OH)2D3 synthesis and degradation and are, thus, responsive to the hormone. 1,25(OH)2D3 inhibits proliferation, induces differentiation and sometimes the apoptosis of human colon cancer cells. A great variety of mechanisms and signaling pathways are involved. Since VDR mediates most, if not all, 1,25(OH)2D3 actions, the control of VDR expression is a crucial aspect of 1,25(OH)2D3 biology. Here, the molecular mechanisms underlying the actions of 1,25(OH)2D3 are reviewed and the repression of the VDR gene by the transcription factor SNAIL in human colon cancer cells is discussed. Understanding these mechanisms may provide the basis for the potential use of this hormone and its non-hypercalcemic derivatives in the prevention and treatment of colon cancer.

High dose brachytherapy (real time) in patients with intermediate- or high-risk prostate cancer: technical description and preliminary experience.

INTRODUCTION: It has been well documented that the outcome of prostate cancer treatment depends on the dose administered. Hence, techniques have been developed that allow high-dose administration without increasing the complications, e.g. external radiotherapy combined with high-dose radiation (HDR) brachytherapy. In this article we analyse the technique and protocol of real-time HDR brachytherapy together with the preliminary results that support its use. Materials and methods. Between June 1998 and December 2004, 100 patients with adenoma of the prostate were treated with 46 Gy of external irradiation to the pelvis and 2 HDR brachytherapy fractions (each of 1150 cGy) at the end of weeks 1 and 3 of a 5-week radiotherapy course. The 1997 American Joint Commission on Cancer (AJCC) system was used to establish disease stage. Patients with intermediate-risk (PSA 10-20 ng/ml or Gleason = 7 or T2c) and high-risk (two intermediate risk factors or PSA > 20 ng/ml or Gleason > 7 or > T2c) without metastases were eligible for the brachytherapy. Biochemical failure was defined according to the American Society for Therapeutic Radiology and Oncology (ASTRO) consensus panel statement. SPSS statistical package was used to quantify survival (Kaplan-Meier method). Toxicity was scored according to RTOG guidelines. RESULTS: The mean age of patients was 67 years (range 49-78). Clinical stage was T2a in 22% of the patients, 26% T2b and 52% T3. Initial PSA was = 10 ng/ml in 22% of the patients and > 10 ng/ml in 78%. Median follow-up was 28 months (range: 12-79). The 5-year overall survival and actuarial biochemical control were 99% and 87% respectively. No chronic severe complications were noted. CONCLUSIONS: The good results of local control, disease-free survival and few complications that the external radiotherapy combined with HDR brachytherapy have shown suggest that the method should be considered as first-choice in the treatment of prostate tumours of high- and intermediate-risk.

High dose brachytherapy (real time) in patients with intermediate- or high-risk prostate cancer: technical description and preliminary experience

Introduction. It has been well documented that the outcome of prostate cancer treatment depends on the dose administered. Hence, techniques have been developed that allow high-dose administration without increasing the complications, e.g. external radiotherapy combined with high-dose radiation (HDR) brachytherapy. In this article we analyse the technique and protocol of real-time HDR brachytherapy together with the preliminary results that support its use. Materials and methods. Between June 1998 and December 2004, 100 patients with adenoma of the prostate were treated with 46 Gy of external irradiation to the pelvis and 2 HDR brachytherapy fractions (each of 1150 cGy) at the end of weeks 1 and 3 of a 5week radiotherapy course. The 1997 American Joint Commission on Cancer (AJCC) system was used to establish disease stage. Patients with intermediate-risk (PSA 10-20 ng/ml or Gleason = 7 or T2c) and high-risk (two intermediate risk factors or PSA > 20 ng/ml or Gleason > 7 or > T2c) without metastases were eligible for the brachytherapy. Biochemical failure was defined according to the American Society for Therapeutic Radiology and Oncology (ASTRO) consensus panel statement. SPSS statistical package was used to quantify survival (Kaplan-Meier method). Toxicity was scored according to RTOG guidelines. Results. The mean age of patients was 67 years (range 49-78). Clinical stage was T2a in 22% of the patients, 26% T2b and 52% T3. Initial PSA was = 10 ng/ml in 22% of the patients and > 10 ng/ml in 78%. Median follow-up was 28 months (range: 12-79). The 5-year overall survival and actuarial biochemical control were 99% and 87% respectively. No chronic severe complications were noted. Conclusions. The good results of local control, di-sease-free survival and few complications that the external radiotherapy combined with HDR brachytherapy have shown suggest that the method should be considered as first-choice in the treatment of prostate tumours of high- and intermediate-risk

Vitamin D and cancer: an update of in vitro and in vivo data.

1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3, Calcitriol) is a pleiotropic hormone with anti-proliferative, pro-apoptotic and pro-differentiation effects on numerous cell types, which suggest anti-cancer activity in addition to its classical regulatory action on calcium and phosphate metabolism. 1,25(OH)2D3 exerts its actions mainly via its high affinity receptor VDR through a complex network of genomic (transcriptional and post-transcriptional) and also non-genomic mechanisms, which are partially coincident in the different cells and tissues studied. Epidemiological and experimental in vitro and in vivo data support a cancer preventive role of 1,25(OH)2D3. The anti-cancer activity of 1,25(OH)2D3 and multiple analogs with reduced calcemic properties, which are thus less toxic, is under investigation in a long list of cultured cell types and in several in vivo models of wild-type and genetically-modified animals. Some vitamin D compounds have reached clinical trials, but results are still scarce.