H2 antihistamines: May be useful for combination therapies in cancer?

Cancer morbimortality is still a great concern despite advances in research and therapies. Histamine and its receptors' ligands can modulate different biological responses according to the cell type and the receptor subtype involved. Besides the wide variety of histamine functions in normal tissues, diverse roles in the acquisition of hallmarks of cancer such as sustained proliferative signaling, resistance to cell death, angiogenesis, metastasis, altered immunity and modified microenvironment have been described. This review summarizes the present knowledge of the various roles of histamine H2 receptor (H2R) ligands in neoplasias. A bioinformatic analysis of human tumors showed dissimilar results in the expression of the H2R gene according to tumor type when comparing malignant versus normal tissues. As well, the relationship between patients' survival parameters and H2R gene expression levels also varied, signaling important divergences in the role of H2R in neoplastic progression in different cancer types. Revised experimental evidence showed multiple effects of H2R antihistamines on several of the cited hallmarks of cancer. Interventional and retrospective clinical studies evaluated different H2R antihistamines in cancer patients with two main adjuvant uses: improving antitumor efficacy (which includes regulation of immune response) and preventing toxic adverse effects produced by chemo or radiotherapy. While there is a long path to go, research on H2R antihistamines may provide new opportunities for developing more refined combination therapeutic strategies for certain cancer types to improve patients' survival and health-related quality of life.

Non-tumorigenic epithelial breast cells and ionizing radiation cooperate in the enhancement of mesenchymal traits in tumorigenic breast cancer cells.

AIMS: Radioresistance and recurrences are crucial hindrances in cancer radiotherapy. Fractionated irradiation can elicit a mesenchymal phenotype in irradiated surviving cells and a deep connection exists between epithelial mesenchymal transition, radioresistance, and metastasis. The aim of this study was to analyze the effect of the secretoma of irradiated non-tumorigenic mammary epithelial cells on surviving irradiated breast tumor cells regarding the gain of mesenchymal traits and migratory ability. MAIN METHODS: MDA-MB-231 and MCF-7 breast cancer cells, irradiated or not, were incubated with conditioned media from MCF-10A non-tumorigenic epithelial breast cells, irradiated or not. After five days, we evaluated the expression and localization of epithelial and mesenchymal markers (by western blot and indirect immunofluorescence), cell migration (using transwells) and metalloproteinases activity (by zymography). We also assessed TGF-ß1 content in conditioned media by immunoblot, and the effect of A83-01 (a selective inhibitor of TGF-ß receptor I) and PP2 (a Src-family tyrosine kinase inhibitor) on nuclear Slug and cell migration. KEY FINDINGS: Conditioned media from MCF-10A cells caused phenotypic changes in breast tumor cells with attainment or enhancement of mesenchymal traits mediated at least in part by the activation of the TGF-ß type I receptor and a signaling pathway involving Src activation/phosphorylation. The effects were more pronounced mostly in irradiated tumor cells treated with conditioned media from irradiated MCF-10A. SIGNIFICANCE: Our results suggest that non-tumorigenic epithelial mammary cells included in the irradiation field could affect the response to irradiation of post-surgery residual cancer cells enhancing EMT progression and thus modifying radiotherapy efficacy.

Histamine H4 receptor agonists induce epithelial-mesenchymal transition events and enhance mammosphere formation via Src and TGF-ß signaling in breast cancer cells.

Epithelial-mesenchymal transition (EMT) contributes to cell invasion and metastasis during the progression of epithelial cancers. Though preclinical evidence suggests a role for histamine H4 receptor (H4R) in breast cancer growth, its function in the EMT is less known. In this study we proposed to investigate the effects of H4R ligands on EMT and mammosphere formation as a surrogate assay for cancer stem cells in breast cancer cells with different invasive phenotype. We also investigated the participation of Src and TGF-ß signaling in these events. Breast cancer cells were treated with the H4R agonists Clobenpropit, VUF8430 and JNJ28610244 and the H4R antagonist JNJ7777120. Immunodetection studies showed cytoplasmic E-cadherin, cytoplasmic and nuclear beta-catenin, nuclear Slug and an increase in vimentin and α-smooth muscle actin expression. There was also an enhancement in cell migration and invasion assessed by transwell units. All these effects were prevented by JNJ7777120. Moreover, H4R agonists induced an increase in phospho-Src levels detected by Western blot. Results revealed the involvement of phospho-Src in EMT events. Upon treatment with H4R agonists there was an increase in phospho-ERK1/2 and TGF-ß1 levels by Western blot, in Smad2/3 positive nuclei by indirect immunofluorescence, and in tumor spheres formation by the mammosphere assay. Notably, the selective TGF-ß1 kinase/activin receptor-like kinase inhibitor A83-01 blocked these effects. Moreover, cells derived from mammospheres exhibited higher Slug expression and enhanced migratory behavior. Collectively, findings support the interaction between H4R and TGF-ß receptor signaling in the enhancement of EMT features and mammosphere formation and point out intracellular TGF-ß1 as a potential mediator of these events.

Histamine prevents radiation-induced mesenchymal changes in breast cancer cells.

Radiotherapy is a prime option for treatment of solid tumors including breast cancer though side effects are usually present. Experimental evidence shows an increase in invasiveness of several neoplastic cell types through conventional tumor irradiation. The induction of epithelial to mesenchymal transition is proposed as an underlying cause of metastasis triggered by gamma irradiation. Experiments were conducted to investigate the role of histamine on the ionizing radiation-induced epithelial to mesenchymal transition events in breast cancer cells with different invasive phenotype. We also evaluated the potential involvement of Src phosphorylation in the migratory capability of irradiated cells upon histamine treatment. MCF-7 and MDA-MB-231 mammary tumor cells were exposed to a single dose of 2Gy of gamma radiation and five days after irradiation mesenchymal-like phenotypic changes were observed by optical microscope. The expression and subcellular localization of E-cadherin, ß-catenin, vimentin and Slug were determined by immunoblot and indirect immunofluorescence. There was a decrease in the epithelial marker E-cadherin expression and an increase in the mesenchymal marker vimentin after irradiation. E-cadherin and ß-catenin were mainly localized in cytoplasm. Slug positive nuclei, matrix metalloproteinase-2 activity and cell migration and invasion were significantly increased. In addition, a significant enhancement in Src phosphorylation/activation could be determined by immunoblot in irradiated cells. MCF-7 and MDA-MB-231 cells also received 1 or 20µM histamine during 24h previous to be irradiated. Notably, pre-treatment of breast cancer cells with 20µM histamine prevented the mesenchymal changes induced by ionizing radiation and also reduced the migratory behavior of irradiated cells decreasing phospho-Src levels. Collectively, our results suggest that histamine may block events related to epithelial to mesenchymal transition in irradiated mammary cancer cells and open a perspective for the potential use of histamine to improve radiotherapy efficacy.

Fibroblasts induce epithelial to mesenchymal transition in breast tumor cells which is prevented by fibroblasts treatment with histamine in high concentration.

Epithelial to mesenchymal transition (EMT) of cancer cells is an essential process in cancer progression. Cancer cells that undergone EMT loose cell-cell contacts, acquire mesenchymal properties and develop migratory and invasive abilities. In previous studies we have demonstrated that histamine may modify the invasive phenotype of pancreatic and mammary tumor cells. In this work we proposed to investigate whether histamine may also influence the interaction between tumor cells and normal fibroblasts. The potential activation of normal CCD-1059Sk fibroblasts by histamine and EMT phenotypic changes induced in MCF-7 and MDA-MB-231 breast tumor cells by the conditioned media (CM) derived from fibroblasts were evaluated. Initially, we determined the presence of H1, H2 and H4 histamine receptors and matrix metalloproteinase 2 (MMP2) mRNA in CCD-1059Sk fibroblasts. MMP2 gelatinolytic activity, cell migration and alpha-smooth muscle actin expression were increased in fibroblasts by low doses (<1µM) and decreased by high doses (20µM) of histamine. MCF-7 cells cultured with CM from fibroblasts exhibited spindle-shaped morphology, cell spreading and cytoplasmic expression of ß-catenin but there was no change in MMP2 activity and cell migration. MDA-MB-231 cells cultured with CM from fibroblasts showed a more elongated phenotype, cell spreading, cytoplasmic ß-catenin, increased MMP2 activity and endogenous TGF-ß1 expression, and enhanced cell migration and invasion. Notably, all these features were reversed when mammary tumor cells were cultured with CM from fibroblasts treated with 20µM histamine. In conclusion, high doses of histamine may prevent the activation of fibroblasts and also avert the EMT related changes induced in epithelial tumor cells by fibroblasts CM.

PANC-1 cells proliferative response to ionizing radiation is related to GSK-3ß phosphorylation.

Radiotherapy may be used to treat pancreatic cancer and relieve pain. We have previously reported that histamine modulates pancreatic adenocarcinoma PANC-1 cell proliferation. This work was aimed to evaluate whether histamine improves radiosensitivity of PANC-1 cells in relation to phosphorylation/inhibition of glycogen synthase kinase-3ß (GSK-3ß). Immediately after γ irradiation, intracellular hydrogen peroxide was markedly decreased together with a rapid increase in catalase activity. Although histamine diminished catalase activity in nonirradiated cells, it only partially hindered the increase observed in irradiated cells and could not modify radiosensitivity. In control cells, a high expression of total and a very low expression of phosphorylated/inactive GSK-3ß were found. An increment in reactive oxygen species levels produced an augmentation in GSK-3ß phosphorylation and suppressed cell proliferation. In both control and histamine-treated irradiated cells, the rise in catalase activity lowered reactive oxygen species levels and only a small increase in phosphorylated GSK-3ß was detected. Alternatively, 3-aminotriazole, an irreversible inhibitor of catalase, reduced the survival fraction in irradiated control cells along with an increment in phosphorylated GSK-3ß. These results suggest that upon irradiation, early catalase activation may be responsible for keeping GSK-3ß active conceding cells a survival advantage toward cytotoxic effects of ionizing radiation.

Aminoguanidine impedes human pancreatic tumor growth and metastasis development in nude mice.

AIM: To study the action of aminoguanidine on pancreatic cancer xenografts in relation to cell proliferation, apoptosis, redox status and vascularization. METHODS: Xenografts of PANC-1 cells were developed in nude mice. The animals were separated into two groups: control and aminoguanidine treated. Tumor growth, survival and appearance of metastases were determined in vivo in both groups. Tumors were excised and ex vivo histochemical studies were performed. Cell growth was assessed by Ki-67 expression. Apoptosis was studied by intratumoral expression of B cell lymphoma-2 protein (Bcl-2) family proteins and Terminal deoxynucleotidyl transferase biotin-dUTP Nick End Labeling (Tunel). Redox status was evaluated by the expression of endothelial nitric oxide synthase (eNOS), catalase, copper-zinc superoxide dismutase (CuZnSOD), manganese superoxide dismutase (MnSOD) and glutathione peroxidase (GPx). Finally, vascularization was determined by Massons trichromic staining, and by VEGF and CD34 expression. RESULTS: Tumor volumes after 32 d of treatment by aminoguanidine (AG) were significantly lower than in control mice (P < 0.01). Median survival of AG mice was significantly greater than control animals (P < 0.01). The appearance of both homolateral and contralateral palpable metastases was significantly delayed in AG group. Apoptotic cells, intratumoral vascularization (trichromic stain) and the expression of Ki-67, Bax, eNOS, CD34, VEGF, catalase, CuZnSOD and MnSOD were diminished in AG treated mice (P < 0.01), while the expression of Bcl-2 and GPx did not change. CONCLUSION: The antitumoral action of aminoguanidine is associated with decreased cell proliferation, reduced angiogenesis, and reduced expression of antioxidant enzymes.

Mechanisms underlying the radioprotective effect of histamine on small intestine.

PURPOSE: To examine the protective effects of histamine on intestinal damage produced by gamma-radiation. MATERIALS AND METHODS: 56 mice were divided into 4 groups. Histamine and Histamine-10 Gy groups received a daily subcutaneous histamine injection (0.1 mg/kg) starting 20 hours before irradiation and continued until the end of the experimental period; the untreated group received saline. Histamine-10 Gy and untreated-10 Gy groups were irradiated with a single dose on whole-body using Cesium-137 source (7 Gy/min) and were sacrificed 3 days after irradiation. Small intestine was removed, fixed and stained with hematoxylin and eosin. The number of intestinal crypts per circumference, and other histological characteristics of intestinal cells were evaluated. We further determined by immunohistochemistry the expression of proliferating cell nuclear antigen (PCNA), Bax, Bcl-2 (pro- and anti-apoptotic protein, respectively), antioxidant enzymes (Superoxide dismutase (SOD), Catalase and Glutathione peroxidase), histamine content and apoptosis by terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate biotin nick end labeling (TUNEL) assay. Cells in the S phase of the cell cycle were identified by immunohistochemical detection of 5-bromo-2'-deoxyuridine (BrdU) incorporation. RESULTS: Histamine treatment reduced mucosal atrophy, edema and preserved villi, crypts and nuclear and cytoplasmic characteristics of small intestine after radiation exposure. Additionally, histamine treatment increased PCNA expression and the BrdU-positive cell number, histamine content, decreased the number of apoptotic cells and significantly increased Catalase and copper-zinc-containing SOD of irradiated mice. CONCLUSIONS: Histamine prevents radiation-induced toxicity by increasing proliferation of damaged intestinal mucosa and suppressing apoptosis that was associated with an increase in SOD and Catalase levels. This effect might be of clinical value in patients undergoing radiotherapy.