Mariangela Correa

Transient inflammatory response induced by apoptotic cells is an important mediator of melanoma cell engraftment and growth.

Two murine melanoma cell lines, Tm1 and Tm5, were derived from a nontumorigenic lineage of pigmented murine melanocytes, melan‐a. Both Tm1 and Tm5 are invariably tumorigenic in syngeneic mice when inoculated s.c. in doses higher than 104 cells; 103 or fewer cells rarely give rise to tumors. We demonstrate that subtumorigenic inocula of Tm1 or Tm5 cells (103) as well as of a known murine melanoma cell line (B16F10) develop as vigorously growing tumor grafts only when coinoculated with apoptotic, but not necrotic cells. The presence of apoptotic cells correlates with a transient inflammatory infiltrate, composed mainly of neutrophils and macrophages. Kinin B1 receptor–deficient mice, which have impaired transmigration of neutrophils to inflamed tissues, had significant growth inhibition of subtumorigenic doses of melanoma cells coinjected with apoptotic cells. Using the same model, tumor take in athymic mice was similar to that seen in wild‐type mice, suggesting that a T cell–dependent inflammatory response is not necessary to promote the survival and growth of subtumorigenic doses of melanoma cells. Taken together, our results describe how tumor engraftment and growth can be profoundly affected by microenvironmental alterations in response to the presence of apoptotic cells. Disrupting the delicate balance between apoptotic cells and leukocyte infiltration may provide potentially important insights for understanding and interfering with tumor cell viability during treatment with either γ‐radiation or apoptosis‐inducing drugs.

Leukotriene B4 creates a favorable microenvironment for murine melanoma growth.

Chronic inflammation has long been associated with neoplastic progression. Our group had recently shown that the addition of a large number of apoptotic tumor cells to the tumor microenvironment induces a potent acute inflammatory reaction capable of promoting melanoma growth; however, primarily necrotizing cells do not cause such a reaction. Here, we show that potent inflammatory agents, such as lipopolysaccharide (LPS) and carrageenan, also promote growth of subtumorigenic doses of melanoma cells, having no effect on melanoma proliferation in vitro. Inhibition of 5-lipoxygenase (5-LOX) seems to have a pivotal role in this model because caffeic acid and MK886, a FLAP (5-LOX–activating protein) inhibitor, partially hindered tumor growth induced by apoptotic cells or LPS. Other enzymes of the arachidonic acid pathway, cyclooxygenase-1 and cyclooxygenase-2, seem to have no participation in this tumor promoter effect, as the inhibitor of both enzymes (indomethacin) did not alter melanoma growth. Leukotriene B4 (LTB4), the main product of the 5-LOX pathway, was able to induce growth of subtumorigenic inocula of melanoma cells, and a LTB4 receptor antagonist inhibited acute inflammation-associated tumor growth. Addition to the tumor inflammatory microenvironment of eicosapentaenoic acid, an ω3-polyunsaturated fatty acid with anti-inflammatory properties, or leukotriene B5, an eicosapentaenoic acid–derived leukotriene, significantly inhibited tumor development. These results give new insights to the mechanisms through which inflammation may contribute to tumor progression and suggest that LOX has an important role in tumor progression associated with an inflammatory state in the presence of apoptosis, which may be a consideration for apoptosis-inducing treatments, such as chemotherapy and radiotherapy. (Mol Cancer Res 2009;7(9):1417–24)

Effects of transforming growth factor-β in the development of inflammatory pseudotumour-like lesions in a murine model

Alterations in transforming growth factor (TGF)‐β signalling have been frequently implicated in human cancer, and an important mechanism underlying its pro‐oncogenic nature is suppression of the host antitumour immune response. Considering the immunosuppressive effect of TGF‐β, we asked whether human tumour cells, known to secrete TGF‐β in culture, would survive and grow when implanted into the peritoneal cavity of immunocompetent mice. Therefore, we developed a xenogeneic model where mice were intraperitoneally (i.p.) injected with a TGF‐β‐secreting human colorectal adenocarcinoma cell line, LISP‐A10. Although animals did not develop macroscopic tumours, the recovery and isolation of human tumour cells was achieved when an inflammatory environment was locally induced by the administration of complete Freunds adjuvant (CFA). This procedure significantly increased TGF‐β concentrations in the peritoneal fluid and was accompanied by impaired activation of the host‐specific immune response against LISP‐A10 cells. Furthermore, inflammatory lesions resembling human inflammatory pseudotumours (IPTs) were observed on the surface of i.p. organs. These lesions could be induced by either injection of LISP‐A10 cells, cells‐conditioned medium or recombinant TGF‐β but only after administration of CFA. In addition, host cyclooxygenase‐2 and kinin receptors played an important role in the induction of TGF‐β‐mediated IPT‐like lesions in our experimental model.

Metastatic melanoma positively influences pregnancy outcome in a mouse model: could a deadly tumor support embryo life?

The incidence of melanoma is increasing worldwide. It is one of the leading cancers in pregnancy and the most common malignancy to metastasize to placenta and fetus. There are no publications about experimental models of melanoma and pregnancy. We propose a new experimental murine model to study the effects of melanoma on pregnancy and its metastatic process. We tested several doses of melanoma cells until we arrived at the optimal dose, which produced tumor growth and allowed animal survival to the end of pregnancy. Two control groups were used: control (C) and stress control (SC) and three different routes of inoculation: intravenous (IV), intraperitoneal (IP) and subcutaneous (SC). All the fetuses and placentas were examined macroscopically and microscopically. The results suggest that melanoma is a risk factor for intrauterine growth restriction but does not affect placental weight. When inoculated by the SC route, the tumor grew only in the site of implantation. The IP route produced peritoneal tumoral growth and also ovarian and uterine metastases in 60% of the cases. The IV route produced pulmonary tumors. No placental or fetal metastases were obtained, regardless of the inoculation route. The injection of melanoma cells by any route did not increase the rate of fetal resorptions. Surprisingly, animals in the IV groups had no resorptions and a significantly higher number of fetuses. This finding may indicate that tumoral factors released in the host organism to favor tumor survival may also have a pro-gestational action and consequently improve the reproductive performance of these animals.

Abstract 115: Correlation between homocysteine concentration in serum and response to anti-hormonal treatment in breast tumor patients

Background: Breast tumors are today the most incident tumor in women. One of the widely used markers for breast cancer is the estrogen receptor (ER), which is member of the nuclear receptor superfamily of transcription factors. Almost 70% of the breast cancer patients are ER positive and can be beneficiated with hormonal therapy. Patients with ER+ tumors are candidates for anti-hormonal therapy, including tamoxifen and anastrozol. Many studies proposed that elevated levels of total homocysteine (tHcy) in the blood can be a risk factor for cancer. It is also known that estrogen status is associated with tHcy concentration. One important epigenetic mechanism which is a well-described for carcinogenesis, and seems to be related to homocysteine pathways, is the DNA methylation. In tumors, it is well documented a decrease in total DNA methylation and a hipermethylation of specific gene promoters. There are no studies showing the impact of anti-hormonal treatment in the tHcy concentration and in the DNA methylation pattern of the breast tumor patients. Objectives: In this study we aimed to verify which is the impact of the anti-hormonal treatment in the tHcy concentration and its correlation with the clinical data. Samples and Methods: To investigate the effect of anti-hormonal therapy on homocysteine metabolism, 50 breast tumor patients were randomly assigned to therapy with anastrozol or tamoxifen or placebo. Sera from the patients were obtained before and after the treatment with anti-hormonal drugs. Homocysteine levels were determined by HPLC. Total DNA methylation was evaluated in the tumor biopsies before the treatment by restriction enzyme method. Pearson9s correlation test and t test were performed for statistical analysis. Results: We observed that the tHcy before the anti-hormonal treatment increase in the patients with a high agressive tumor (p=0,0401), and with positive lifonodes (p=0,0379). After the treatment with tamoxifen, the tHcy level decreases (p=0,0263). We could observe that the decrease was significant in the patients with positive linfonodes (p=0,0400). No significant correlations were observed when the patients were treated with anastrozol or placebo and also no correlation were observed in these patients when we correlated the tHcy variation with the clinical data set. We observed a total DNA hypomethylation in the tumors before treatment when compared with adjacent tissue (p=0,0157). Conclusions and perspectives: We could observe that the higher tHcy levels are related to aggressive tumors. Our results also show that the treatment with tamoxifen, mainly in aggressive tumors, decreases the tHcy levels showing that this treatment can have a role in the tHcy level. Other clinical data and new studies should be done to evaluate the tamoxifen impact in tHcy and also its relation with the tumor DNA methylation. Financial support: Fapesp, CAPES, CNPq. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 115. doi:10.1158/1538-7445.AM2011-115