Cathepsin B is involved in the apoptosis intrinsic pathway induced by Bacillus Calmette-Guérin in transitional cancer cell lines

Bacillus Calmette-Guérin (BCG) is the most effective treatment for superficial and in situ transitional bladder cancer. Although the complete mechanisms for its effect are not fully understood yet, both immunological and direct effects on tumor cells have been proposed. It has been proposed that apoptotic tumor cells could be better inducers of immunity than necrotic ones. Thus, apoptosis of bladder cancer cells could contribute to a global response to BCG. Lysosomal hydrolase cathepsin B (CB) is involved in the apoptotic process and has a key role in breast cancer cell programmed death through the activation of a pro-apoptotic protein BID. Truncated BID participates in the mitochondrial apoptotic pathway that involves the activation of pro-caspase 9. The possibility that CB can be involved in apoptosis of TCC line has not been explored yet. Therefore, we analyzed the participation of CB in BCG-induced apoptosis of human and murine TCC lines. Apoptosis was evaluated by a morphologic assay and CB activity by a substrate-specific colorimetric method. Expression of CB, BID and pro-caspase 9 was determined by Western blotting. BCG induced apoptosis of murine (MBT2, MB49) and human (T24) TCC lines. An increase in both CB activity and protein was also observed. The apoptosis of T24 and MB49 cell lines was mediated by activation of pro-caspase 9 and BID, both proteins are involved in mitochondrial apoptosis. Apoptosis and activation of pro-caspase 9 and BID were inhibited by CA-074Me (CA), a cell permeable CB inhibitor. Thus, CB is involved in BCG-induced apoptosis of TCC lines, using at least in part the mitochondrial pathway.Fil: Sandes, Eduardo Omar. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Lodillinsky, Catalina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Cwirenbaum, Ruth Ana. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Argüelles, Claudia Lidia. Dirección Nacional de Instituto de Investigación.Administración Nacional de Laboratorios e Institutos de Salud "Dr. Carlos G. Malbrán"; ArgentinaFil: Casabé, Alberto. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Eijan, Ana Maria. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Expression of bladder cancer‑associated glycans in murine tumor cell lines

The characterization of murine cell lines is of great importance in order to identify preclinical models that could resemble human diseases. Aberrant glycosylation includes the loss, excessive or novel expression of glycans and the appearance of truncated structures. MB49 and MB49-I are currently the only two murine cell lines available for the development of preclinical bladder cancer models. The glycans Lewis X (LeX), Sialyl lewis X (SLeX) and Sialyl Tn (STn) have previously been associated with aggressiveness, dissemination and poor prognosis in human bladder cancer, additionally N-glycolyl GM3 (NGcGM3) is a neo-antigen expressed in many types of tumors; however, to the best of our knowledge, its expression has not previously been assessed in this type of cancer. Taking into account the relevance of glycans in tumor biology and considering that they can act as targets of therapies and biomarkers, the present study evaluated the expression of LeX, SLeX, STn and NGcGM3 in MB49 and MB49-I cells, in different growth conditions such as mono-layer cultures, three-dimensional multicellular spheroids and mouse heterotopic and orthotopic tumors. The expression of LeX was not detected in either cell line, whereas SLeX was expressed in monolayers, spheroids and orthotopic tumors of both cell lines. STn was only identified in MB49 monolayers and spheroids. There are no reports concerning the expression of NGcGM3 in human or murine bladder cancer. In our hands, MB49 and MB49-I expressed this ganglioside in all the growth conditions evaluated. The assessment of its expression in cancer cell lines and patient tumors is of great importance, considering the relevance of this ganglioside in tumor biology. The data obtained by the present study demonstrates that glycan expression may be substantially altered depending on the growth conditions, highlighting the importance of the characterization of murine cancer models. To the best of our knowledge, the present study is the first to examine the expression of cancer-associated glycans, in the two murine cell lines available for the development of preclinical studies in bladder cancer.Fil: Alberto, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; ArgentinaFil: Cuello, Héctor Adrián. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; ArgentinaFil: Gulino, Cynthia Antonella. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; ArgentinaFil: Pifano, Marina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Belgorosky, Denise. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; ArgentinaFil: Gabri, Mariano. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; ArgentinaFil: Eijan, Ana Maria. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Segatori, Valeria Inés. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentin

Synthesis of new para-aminobenzoic acid derivatives, in vitro biological evaluation and preclinical validation of DAB-2-28 as a therapeutic option for the treatment of bladder cancer

Chronic inflammation plays a crucial role in bladder cancer (BCa) development and progression. To offer a unique treatment opportunity for this type of cancer, a hydrazide derivative namely, DAB-1, was recently identified in our laboratory as a potential drug to target cancer-related inflammation. In preclinical models of murine BCa, this particular compound exhibited remarkable anticancer activities. Structurally, DAB-1 is made from para-aminobenzoic acid and bears two different components, a maleimide and a hydrazide moieties, which are critical for its anti-inflammatory activity and its anticancer properties. In order to improve its biological potential, the hydrazide moiety was further modified to provide 3 ​s-generation molecules named, DAB-2-28, DAB-2-31A, and DAB-2-31B, and two third-generation molecules named, DAB-3-27 and DAB-3-33. Data from in vitro studies revealed that, among the different DAB molecules under study, DAB-2-28 has less cytotoxic activity with greater efficiency than DAB-1 to inhibit the production of nitric oxide (NO) induced by the combination of IFNγ with TNFα, as well as the activation of pro-tumoral and pro-inflammatory signaling pathways IL6/STAT3 and TNFα/NFκB. Moreover, while DAB-2-28 exhibited similar anti-inflammatory activity in vivo to DAB-1 in a model of carrageenan-induced acute inflammation, it efficiently inhibited the expression of the enzymes iNOS and COX-2 induced by the combined activation of IFNγ with LPS in peritoneal macrophages. Notably, analysis of the growth kinetics of MB49-I tumors implanted subcutaneously in C57Bl/6 mice showed that DAB-2-28 was more efficient to inhibit tumor development. In conclusion, this study provided preclinical proof-of-principle for DAB-2-28 molecule in the treatment of BCa.Fil: Oufqir, Yassine. Université du Québec a Montreal; CanadáFil: Fortin, Laurie. Université du Québec a Montreal; CanadáFil: Girouard, Julie. Université du Québec a Montreal; CanadáFil: Cloutier, Francis. Université du Québec a Montreal; CanadáFil: Cloutier, Maude. Université du Québec a Montreal; CanadáFil: Leclerc, Marie France. Université du Québec a Montreal; CanadáFil: Belgorosky, Denise. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Eijan, Ana Maria. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Bérubé, Gervais. Université du Québec a Montreal; CanadáFil: Reyes-Moreno, Carlos. Université du Québec a Montreal; Canad

Human papillomavirus (HPV) DNA in penile carcinomas in Argentina: Analysis of primary tumors and lymph nodes

Among sexually transmitted diseases, infection by human papillomavirus (HPV) has become one of the most important. On the other hand, though epidemiological data show that some HPV types are closely associated with cervical cancer, few reports have been found with reference to penile carcinoma because of its rare occurrence. The aim of this study was to investigate the relationship between HPV infection and penile cancer in Argentina. A retrospective study was carried out on 38 white men with penile squamous-cell carcinoma. Sixty-five archival fixed biopsies taken from 34 primary penile tumors, 25 nodal metastases, 1 skin “satellite” metastasis and 5 histologically normal lymph nodes were used as specimens. HPV detection and typing were carried out by the polymerase chain reaction (PCR) using generic primers, combined with single-stranded conformational polymorphism (SSCP) analysis. HPV DNA was found in 71% patients, corresponding 81% of them to “high risk” types, with predominance of HPV 18. Both primary tumors and metastases showed concordance of HPV occurrence and type in both lesions. In 3 patients, HPV 16 was detected not only in primary tumors and metastases, but also in histologically normal lymph nodes. Our data indicate that most penile carcinomas in Argentine patients are etiologically related to HPV, especially to “high risk” genital types. The agreement in HPV detection between primary tumors and metastases suggests a potential viral role in tumor progression. HPV detection in otherwise histologically normal lymph nodes might be useful as early marker of a metastatic process.Fil: Picconi, María A.. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud “Dr. C.G. Malbrán”; ArgentinaFil: Eijan, Ana Maria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires; ArgentinaFil: Distéfano, Angélica L.. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud “Dr. C.G. Malbrán”; ArgentinaFil: Pueyo, Silvia. Provincia de Buenos Aires. Ministerio de Salud. Hospital Materno Infantil de San Isidro. Servicio de Dermatología; ArgentinaFil: Alonio, Lidia Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud “Dr. C.G. Malbrán”; ArgentinaFil: Gorostidi, Susana. Universidad de Buenos Aires; ArgentinaFil: Teyssié, Angélica R.. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud “Dr. C.G. Malbrán”; ArgentinaFil: Casabone, Bernardo Adalberto. Universidad de Buenos Aires; Argentin

Flavonoid silybin improves the response to radiotherapy in invasive bladder cancer

Conservative treatment for invasive bladder cancer (BC) involves a complete transurethral tumor resection combined with chemotherapy (CT) and radiotherapy (RT). The major obstacles of chemo-radiotherapy are the addition of the toxicities of RT and CT, and the recurrence due to RT and CT resistances. The flavonoid Silybin (Sb) inhibits pathways involved in cell survival and resistance mechanisms, therefore the purpose of this paper was to study in vitro and in vivo, the ability of Sb to improve the response to RT, in two murine BC cell lines, with different levels of invasiveness, placing emphasis on radio-sensitivity, and pathways involved in radio-resistance and survival. In vitro, Sb radio-sensitized murine invasive cells through the inhibition of RT-induced NF-κB and PI3K pathways, and the increase of oxidative stress, while non-invasive cells did not show to be sensitized. In vivo, Sb improved RT-response and overall survival in invasive murine tumors. As Sb is already being tested in clinical trials for other urological cancers and it improves RT-response in invasive BC, these results could have translational relevance, supporting further research.Fil: Prack Mc Cormick, Bárbara Patricia. Universidad Nacional de Lomas de Zamora. Facultad de Cs.agrarias. Laboratorio de Suelos; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Langle, Yanina Verónica. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; ArgentinaFil: Belgorosky, Denise. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; ArgentinaFil: Vanzulli, Silvia. Academia Nacional de Medicina de Buenos Aires; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; ArgentinaFil: Balarino, Natalia Patricia. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Sandes, Eduardo Omar. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; ArgentinaFil: Eijan, Ana Maria. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Effect of nitric oxide inhibition in Bacillus Calmette-Guerin bladder cancer treatment

Background: Bacillus Calmette-Guerin (BCG) is the standard treatment for patients with high-risk non-muscle invasive bladder cancer (BC). Despite its success, about 30–50% of patients are refractory. It was reported that inducible nitric oxide synthase (iNOS) tumor expression is presented in 50% of human BC, associated with bad prognosis and BCG failure. Objective: to evaluate in human bladder tumors the association between iNOS expression and the tumor microenvironment focusing on the immunosuppressive protein S100A9. Also, investigate in a preclinical murine MB49-BC model the tumor immunoresponse induced by BCG in combination with the nitric oxide production inhibitor L-NAME. Results: In human bladder tumors, we detected a positive association between iNOS and S100A9 tumor expression, suggesting a relationship between both immunomodulatory proteins. We also found a positive correlation between iNOS tumor expression and the presence of S100A9+ tumor-infiltrating cells, suggesting an immunosuppressive tumor microenvironment induced by the nitric oxide production. Using the subcutaneous murine BC model, we show that similarly to the human pathology, MB49 tumors constitutively expressed iNOS and S100A9 protein. MB49 tumor-bearing mice presented an immunosuppressive systemic profile characterized by fewer cytotoxic cells (CD8+ and NK) and higher suppressor cells (Treg and myeloid-derived suppressor cells -MDSC-) compared to normal mice. BCG treatment reduced tumor growth, increasing local CD8+-infiltrating cells and induced a systemic increase in CD8+ and a reduction in Treg. BCG combined with L-NAME, significantly reduced tumor growth compared to BCG alone, diminishing iNOS and S100A9 tumor expression and increasing CD8+-infiltrating cells in tumor microenvironment. This local response was accompanied by the systemic increase in CD8+ and NK cells, and the reduction in Treg and MDSC, even more than BCG alone. Similar results were obtained using the orthotopic BC model, where an increase in specific cytotoxicity against MB49 tumor cells was detected. Conclusion: The present study provides preclinical information where NO inhibition in iNOS-expressing bladder tumors could contribute to improve BCG antitumor immune response. The association between iNOS and S100A9 in human BC supports the hypothesis that iNOS expression is a negative prognostic factor and a promising therapeutic target.Fil: Langle, Yanina Verónica. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Balarino, Natalia Patricia. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Belgorosky, Denise. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Cresta Morgado, Pablo Damián. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Sandes, Eduardo Omar. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Marino, Lina. Universidad de Buenos Aires. Facultad de Medicina; ArgentinaFil: Rojas Bilbao, Erica. Universidad de Buenos Aires. Facultad de Medicina; ArgentinaFil: Zambrano, Macarena. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Lodillinsky, Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Eijan, Ana Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentin

S100A9 expression associated with nitric oxide is a good marker of prognosis in patients with bladder cancer: its inhibition as a potential therapeutic target

El óxido nítrico (NO) es producido por las enzimas NO sintasas (NOS). Anteriormente describimos que la isoforma inducible (iNOS) se expresa en el 50% de los cánceres de vejiga (CaV) humanos, asociada a recurrencia e invasión. La producción de NO puede alterar la expresión de proteínas relacionadas con la progresión tumoral. Utilizando el modelo de CaV murino MB49 que genera tumores no músculo invasores (NMI) y expresa iNOS identificamos que el tratamiento con el inhibidor de la producción de NO, L-NAME, redujo la expresión de la proteína S100A9, vinculada a la generación de una respuesta inmune supresora mediante el reclutamiento de células supresoras derivadas del linaje mieloide (MDSC). Objetivos: Evaluar 1) S100A9 como marcador pronóstico en pacientes con CaV. 2) La inhibición de NO como blanco terapéutico utilizando el modelo murino, enfocándonos en la expresión de S100A9, las MDSC y el crecimiento tumoral solo o bajo tratamiento con BCG. Identificamos que S100A9 se expresa en células tumorales y células inmunes que infiltran los tumores (células acompañantes). Detectamos una correlación positiva entre iNOS y S100A9 en las células tumorales, sugiriendo que su expresión estaría vinculada. Los tumores invasores presentan mayor número de células acompañantes positivas para S100A9, en comparación con los tumores NMI (p <0,05). Este infiltrado es a predominio de monocitos/macrófagos (CD14) en los tumores NMI (p <0,01), mientras que en los invasores hay un nivel similar de células CD14 y CD15 (granulocitos). En el modelo MB49, demostramos que L-NAME redujo el crecimiento tumoral ortotópico (46% de los ratones presentaron remisión completa) y la expresión de S100A9 en los tumores. Observamos que el número de MDSC en ganglio y bazo estaba aumentado y que los valores solo se normalizaban bajo tratamiento combinado de BCG+L-NAME. Conclusión: Estos hallazgos respaldan la hipótesis de que la inhibición del NO es un buen blanco terapéutico reduciendo en parte la generación de una respuesta inmune supresora. La expresión de S100A9 en células inmunológicas es un buen marcador de progresión tumoral, asociado a la producción de NO.Fil: Langle, Yanina Verónica. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Sandes, Eduardo Omar. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Belgorosky, Denise. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Balarino, Natalia Patricia. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Prack Mc Cormick, Bárbara Patricia. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Marino, Lina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Rojas Bilbao, Érica. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Malagrino, Héctor. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Pasik, Leonardo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Casabé, Alberto. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Eijan, Ana Maria. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Hypoxia-induced nitric oxide release by luminal cells stimulates proliferation and uPA secretion of myoepithelial cells in a bicellular murine mammary tumor

Introduction: LM38 murine mammary adenocarcinoma model is formed by LM38-LP (myoepithelial and luminal), LM38-HP (luminal) and LM38-D2 (myoepithelial) cell lines. In a previous work, we had shown that LM38-HP and LM38-D2 cell lines are less malignant than the bicellular LM38-LP cell line. Purpose: To study the role of nitric oxide (NO) as one of the mediators of functional interactions between malignant luminal and myoepithelial cells. Methods and results: Using immunohistochemistry, in vivo iNOS expression was only detected in the luminal cells of bicellular LM38-LP and most cells of LM38-HP tumors. In cobalt-induced pseudohypoxia, LM38-LP and LM38-HP cell lines significantly increased HIF-1α and iNOS expression (Western blotting) and therefore NO production (Griess method). This increase was inhibited by the iNOS inhibitor 1400 W. On the other side, NO was not detectable in LM38-D2 cells either in basal or in pseudohypoxia. In addition, pseudohypoxia increased urokinase-type plasminogen activator (uPA) secretion by LM38-LP and LM38-HP cells and migration in the LM38-LP cell line, without modulating these properties in LM38-D2 cells (radial caseinolysis). The NO donor DETA/NONOate (500 μM) was able to increase uPA secretion and in vitro growth of LM38-D2. In agreement, 1400 W prevented in vivo growth of the myoepithelial LM38-D2 cells. Conclusions: Hypoxia leads to an enhanced NO production by the luminal component, through HIF-1α and iNOS, which can stimulate myoepithelial cell proliferation and uPA secretion. In these new conditions, myoepithelial cells might act as an invasive forefront generating gaps that could help luminal cells to escape from the primary tumor.Fil: Krasnapolski, Martin Alejandro. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología ; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Lodillinsky, Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología ; Argentina. Institut Curie Research Center; FranciaFil: Bal, Elisa Dora. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología ; ArgentinaFil: Eijan, Ana Maria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología ; Argentin

Differential nitric oxide release and sensitivity to injury in different murine mammary tumor cell lines

The purpose of this study was to determine whether nitric oxide (NO) production by different mammary tumor cell lines correlated with their sensitivity to NO mediated injury. Three mammary tumor cell lines LM2, LM3 and LMM3 syngeneic to BALB/c mice were cultured in vitro with IFNgamma + LPS. Different levels of NO production among the three lines were detected in culture supernatants. The only tumor cell line which did not produce NO (LM2) showed the highest sensitivity to SNP-derived NO cytotoxicity (87%), while LM3 and LMM3 which both produced higher levels of NO than LM2, showed lower cytotoxicity by SNP (39% and 22% respectively). Spleen cells (SC) from M2 tumor bearing mice (TBM) were able to lyse LM2 cells by NO-dependent mechanisms. SC from M3-TBM exerted cytotoxicity against LM3 cells mainly by NO-independent mechanisms. Thus, we postulate an inverse correlation between NO production and NO mediated cytotoxicity in the three mammary tumor cell lines. It is possible that tumor cells producing NO develop mechanisms to resist NO injury.Fil: Eijan, Ana Maria. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Davel, L. E.. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Rueda, H. A.. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Rozenberg, G.. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaFil: Sacerdote de Lustig, Eugenia. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Jasnis, Maria Adela. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología "Ángel H. Roffo"; ArgentinaUnidad documental simpl

Microdevices for cancer stem cell culture as a predictive chemotherapeutic response platform

Abstract: Microfluidic platforms for clinical use are a promising translational strategy for cancer research specially for drug screening. Identifying cancer stem cells (CSC) using sphere culture techniques in microfluidic devices (MDs) showed to be better reproducing physiological responses than other in vitro models and allow the optimization of samples and reagents. We evaluated individual sphere proliferation and stemness toward chemotherapeutic treatment (CT) with doxorubicin and cisplatin in bladder cancer cell lines (MB49-I and J82) cultured in MDs used as CSC treatment response platform. Our results confirm the usefulness of this device to evaluate the CT effect in sphere-forming efficiency, size, and growth rate from individual spheres within MDs and robust information comparable to conventional culture plates was obtained. The expression of pluripotency genetic markers (Oct4, Sox2, Nanog, and CD44) could be analyzed by qPCR and immunofluorescence in spheres growing directly in MDs. MDs are a suitable platform for sphere isolation from tumor samples and can provide information about CT response. Microfluidic-based CSC studies could provide information about treatment response of cancer patients from small samples and can be a promising tool for CSC-targeted specific treatment with potential in precision medicine. Key messages: We have designed a microfluidic platform for CSC enriched culture by tumor sphere formation.Using MDs, we could quantify and determine sphere response after CT using murine and human cell lines as a proof of concept.MDs can be used as a tumor-derived sphere isolation platform to test the effect of antitumoral compounds in sphere proliferation.Fil: Agüero, Eduardo Imanol. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; ArgentinaFil: Belgorosky, Denise. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; ArgentinaFil: García Silva, Julio Israel. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Booth, Ross. Roche Sequencing Solutions; Estados UnidosFil: Lerner, Betiana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Florida International University; Estados Unidos. Collaborative Research Institute Intelligent Oncology; Alemania. Universidad Tecnológica Nacional; Argentina. Universidad de Buenos Aires. Facultad de Ingeniería. Instituto de Ingeniería Biomédica; ArgentinaFil: Perez, Maximiliano Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Florida International University; Estados Unidos. Collaborative Research Institute Intelligent Oncology; Alemania. Universidad Tecnológica Nacional; Argentina. Universidad de Buenos Aires. Facultad de Ingeniería. Instituto de Ingeniería Biomédica; ArgentinaFil: Eijan, Ana Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Oncología; Argentin