adrenomedullin and Colorectal-Neoplasms

Hypertension is a common early adverse event of anti-angiogenic treatment of cancer and may associate with treatment response. However, blood pressure measurement as a surrogate response biomarker has methodological limitations, and predictive biomarkers of angiogenesis inhibitors are lacking. In disease associated with hypertension, vasoactive peptides have been linked to cardiovascular pressure load. Here, we have explored potential associations between circulating levels of vasoactive peptides and tumor response during bevacizumab-containing treatment of colorectal cancer.. Metastatic colorectal cancer (mCRC) patients with available best objective response (ORR) and time to tumor progression (TTP) data were included from a randomized clinical trial investigating maintenance therapy after first line chemotherapy plus bevacizumab. Midregional-pro-adrenomedullin (MR-proADM), midregional-pro-atrial-natriuretic-peptide (MR-proANP), and C-terminal-prepro-vasopressin (Copeptin) vasoactive peptide concentrations were measured in plasma at baseline and after 6 weeks of chemotherapy and bevacizumab treatment (n = 97). We determined associations among clinical outcome (ORR and TTP), peptide levels, and hypertension (NCI-CTCAE 4.0 criteria), using Spearman's test, multiple linear regression, and Mann-Whitney's test.. Increasing levels of vasoactive peptides from baseline and after six weeks of treatment were associated with improved treatment outcome (MR-proADM: ORR, p = .0003; TTP, p = .05; MR-proANP: ORR, p = .05; TTP, p = .03; Copeptin: ORR, p = .10; TTP, p = .02). Patients with increasing levels of all three peptides (n = 28) versus increasing levels of one or two peptides (n = 59) showed a median TTP of 284 and 225 d, respectively (p = .02).. Our results suggest that increasing systemic levels of vasoactive peptides associate with improved tumor response and TTP in mCRC patients treated with a bevacizumab-containing regimen. These findings support the proposed link between the tumor vasculature and the cardiovascular system of the host. This should motivate further studies that investigate the potential role of vasoactive peptides as a novel class of dynamic biomarkers in the treatment of cancer.

Topics: Adrenomedullin; Adult; Aged; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Antineoplastic Agents; Atrial Natriuretic Factor; Biomarkers, Tumor; Colorectal Neoplasms; Female; Follow-Up Studies; Glycopeptides; Humans; Lymphatic Metastasis; Male; Middle Aged; Prognosis; Protein Precursors; Survival Rate

The KRAS oncogene influences angiogenesis, metastasis and chemoresistance in colorectal cancers (CRCs), and these processes are all enhanced in hypoxic conditions. To define functional activities of mutant KRAS in a hypoxic microenvironment, we first performed cDNA microarray experiments in isogenic DKs5 and DKO3 colon cancer cell lines that differ only by their expression of mutant KRAS (K-ras(D13)). Adrenomedullin (ADM) was identified as one of the most significantly upregulated genes in DKs5 cells that express the KRAS oncogene in hypoxia (3.2-fold, p = 1.47 × 10(-5)). Ectopic expression of mutant KRAS (K-ras(V12)) in Caco-2 cells (K-ras(WT)) induced ADM, whereas selective knockdown of mutant KRAS alleles (K-ras(D13) or K-ras(V12)) in HCT116, DLD1 and SW480 colon cancer cells suppressed the expression of ADM in hypoxia. Knockdown of ADM in colon tumor xenografts blocked angiogenesis and stimulated apoptosis, resulting in tumor suppression. Furthermore, ADM also regulated colon cancer cell invasion in vitro. Among 56 patients with CRC, significantly higher expression levels of ADM were observed in samples harboring a KRAS mutation. Collectively, ADM is a new target of oncogenic KRAS in the setting of hypoxia. This observation suggests that therapeutic targets may differ depending upon the specific tumor microenvironment.

Topics: Adrenomedullin; Animals; Cell Hypoxia; Cell Line, Tumor; Colorectal Neoplasms; Female; Humans; Immunohistochemistry; Mice, Nude; Oligonucleotide Array Sequence Analysis; Proto-Oncogene Proteins; Proto-Oncogene Proteins p21(ras); ras Proteins; Tumor Microenvironment; Xenograft Model Antitumor Assays

Adrenomedullin (AM) is a multifunctional peptide vasodilator that transduces its effects through calcitonin receptor-like receptor/receptor activity-modifying protein-2 and -3 (CLR/RAMP2 and CLR/RAMP3). In this study, real-time quantitative reverse transcription demonstrated a significant expression of AM mRNA in tumor samples from colorectal cancer (CRC) patients in clinical stage II, III, and IV when compared with normal colorectal tissue. AM, CLR, RAMP2, and RAMP3 proteins were immunohistochemically localized in the carcinomatous epithelial compartment of CRC tissue. Tissue microarray analysis revealed a clear increase of AM, CLR, RAMP2, and RAMP3 staining in lymph node and distant metastasis when compared with primary tumors. The human colon carcinoma cells HT-29 expressed and secreted AM into the culture medium with a significant increase under hypoxia. Treatment of HT-29 cells with synthetic AM stimulated cell proliferation and invasion in vitro. Incubation with anti-AM antibody (αAM), anti-AM receptors antibodies (αAMR), or AM antagonist AM22-52 inhibited significantly basal levels of proliferation of HT-29 cells, suggesting that AM may function as an autocrine growth factor for CRC cells. Treatment with αAM significantly suppressed the growth of HT-29 tumor xenografts in vivo. Histological examination of αAM-treated tumors showed evidence of disruption of tumor vascularity with decreased microvessel density, depletion of endothelial cells and pericytes, and increased tumor cell apoptosis. These findings highlight the potential importance of AM and its receptors in the progression of CRC and support the conclusion that αAM treatment inhibits tumor growth by suppression of angiogenesis and tumor growth, suggesting that AM may be a useful therapeutic target.

Topics: Adrenomedullin; Adult; Aged; Aged, 80 and over; Animals; Calcitonin Receptor-Like Protein; Cell Line, Tumor; Cell Proliferation; Colorectal Neoplasms; Female; HT29 Cells; Humans; Lymphatic Metastasis; Male; Mice; Mice, Nude; Middle Aged; Neoplasm Invasiveness; Neoplasm Staging; Neoplasm Transplantation; Neovascularization, Pathologic; Receptor Activity-Modifying Protein 2; Receptor Activity-Modifying Protein 3; Receptors, Adrenomedullin; Receptors, Calcitonin; RNA, Messenger

Colorectal cancer (CRC) is the most severe complication in inflammatory bowel disease (IBD). In the present study we investigated different mechanistic links between chronic colonic inflammation and its progression to adenocarcinoma. Along these lines, given that adrenomedullin (AM) has been implicated in carcinogenesis, we also analyzed changes in its colonic expression.. Mice were exposed to 5, 10, and 15 cycles of dextran sulfate sodium (DSS); each cycle consisted of 0.7% DSS for 1 week followed by distilled water for 10 days. After each period, macroscopic and histological studies, as well as characterization of inflammatory and tumor biomarkers, were carried out.. The disease activity index (DAI) showed that the disease was present from the third cycle and it gradually increased during the course of DSS treatment. Macroscopic tumors were only seen after 15 cycles, and microscopic study showed that inflammation, dysplasia, and adenocarcinomas correlated with DSS cycles. β-Catenin and proliferating cell nuclear antigen expressions progressively increased in animals treated with the different cycles of DSS. TNF-α and IFN-γ showed the highest production at the tenth cycle. COX-2, mPGES-1, and iNOS levels were also appreciably higher at the fifth and tenth cycles. Moreover, we observed a progressive enhancement in AM expression and changes in its intracellular location during the progression of the disease.. Our results show an early induction of proinflammatory factors, which may contribute to the development of colon cancer, as well as demonstrate, for the first time, the expression of AM in IBD-derived CRC.

Topics: Adenocarcinoma; Adrenomedullin; Animals; beta Catenin; Biomarkers, Tumor; Blotting, Western; Cell Transformation, Neoplastic; Chronic Disease; Colitis, Ulcerative; Colorectal Neoplasms; Cytokines; Dextran Sulfate; Female; Immunoenzyme Techniques; Mice; Mice, Inbred C57BL; Tumor Necrosis Factor-alpha

Recently, we determined several potential prognostic factors in vivo using hypoxic tumor cells from hepatic metastases of colorectal cancer (CRC). Among them, expression of adrenomedullin (ADM) was an interesting target because it is highly induced by hypoxia.. To evaluate the prognostic impact of the expression of ADM, samples from a total of 373 CRC patients were analyzed by microarray (n = 222), and quantitative reverse-transcriptase polymerase chain reaction (n = 151).. ADM was a novel independent prognostic factor for CRC (p = 0.027). ADM expression correlated with hypoxia-inducible factor-1 A (p < 0.0001) and vascular endothelial growth factor (p < 0.0001) in vivo.. ADM expression is a useful marker for predicting high risk of relapse and cancer-related death in patients who have undergone curative resection for CRC.

Topics: Adrenomedullin; Aged; Cell Hypoxia; Colorectal Neoplasms; Disease-Free Survival; Female; Humans; Hypoxia-Inducible Factor 1; Male; Middle Aged; Multivariate Analysis; Reverse Transcriptase Polymerase Chain Reaction; Survival Rate; Vascular Endothelial Growth Factor A

Adrenomedullin (ADM) is a vasodilator peptide, first isolated from human pheochromocytoma. To explore the pathophysiological role of ADM in ischemic conditions, we investigated the effects of hypoxia on ADM production and ADM mRNA expression in a cultured human colorectal carcinoma cell line, DLD-1. Northern blot analysis and radioimmunoassay showed that hypoxia stimulated the accumulation of ADM mRNA in the DLD-1 cells and immunoreactive ADM (ir-ADM) in the cultured media. Exposure to hypoxia for 12 hours increased ADM mRNA levels about 6-fold and ir-ADM levels about 4-fold. Moreover, treatment of DLD-1 cells with cobalt chloride, which mimics hypoxic states, significantly increased ADM mRNA levels about 18-fold and ir-ADM levels about 4-fold. These results suggest that ADM plays an important role in the pathophysiology of ischemic states.

Topics: Adrenomedullin; Cobalt; Colorectal Neoplasms; Culture Media; Gene Expression Regulation; Humans; Hypoxia; Ischemia; Peptides; RNA, Messenger; Tumor Cells, Cultured; Vasodilator Agents

Production and secretion of endothelin-1 (ET-1) and adrenomedullin (ADM) by a cultured human colorectal adenocarcinoma cell line, DLD-1, were studied by radioimmunoassay and Northern blot analysis. Both immunoreactive (IR)-ET and IR-ADM were detected by radioimmunoassay in the culture medium of DLD-1 (IR-ET 0.86 +/- 0.05 fmol/10(5) cells/ 24 h; IR-ADM 1.20 +/- 0.09 fmol/10(5) cells/24 h; n = 5, mean +/- SEM). An analysis by reverse-phase high-performance liquid chromatography (HPLC) of the IR-ET in the culture medium showed a major immunoreactive peak in the position of ET-1. Reverse-phase HPLC of the IR-ADM in the medium showed three immunoreactive peaks, one of which eluted in the position of human ADM. Northern blot analysis showed the expression of ET-1 mRNA and ADM mRNA in the DLD-1 cells. Treatment with interferon-gamma (1-100 U/ml) for 24 h decreased the IR-ET levels in the culture medium but significantly increased IR-ADM levels. This study has shown the production and secretion of two vasoactive peptides, ET-1 and ADM, by DLD-1 colorectal adenocarcinoma cells. The secretion of IR-ET was decreased by treatment with interferon-gamma. These findings suggest possible pathophysiologic roles for ET-1 and ADM in colon mucosal epithelial cells and tumors derived from them.

Topics: Adenocarcinoma; Adrenomedullin; Blotting, Northern; Colorectal Neoplasms; Endothelin-1; Humans; Interferon-gamma; Peptides; Radioimmunoassay; Tumor Cells, Cultured