Resistencia a drogas en pacientes con cáncer de mama

El cáncer de mama sigue siendo una de las principales causas de muerte en la mujer. Uno de los tratamientos que se utiliza en la actualidad para combatir esta enfermedad es la quimioterapia. Lamentablemente en muchos casos esta terapia fracasa porque las células tumorales desarrollan múltiples mecanismos de resistencia a las drogas antitumorales. Existen diversos genes/proteínas que cuando se expresan anormalmente en los tumores impiden que las drogas antitumorales cumplan su función. Entre las proteínas relacionadas con resistencia a drogas antineoplásicas figuran la proteína P170, la proteína HER-2/neu y las proteínas de golpe de calor. Nuestro grupo de trabajo estudia diversas moléculas que se expresan en los tumores de mama y que podrían predecir la sensibilidad/resistencia a la quimioterapia. El objetivo es poder orientar a los oncólogos en la selección de las terapias más efectivas para cada paciente.Breast cancer is one of the principal causes of death in women. Chemotherapy is one of the effective treatments used in breast cancer patients. However, in several cases chemotherapy fails because tumor cells may develop several mechanisms of antitumor drug resistance. There are different genes/proteins that, when abnormally expressed in the tumors, prevent the function of the antitumor drugs. Among the proteins related with chemotherapy resistance is the proteins P170, HER-2/neu, and heat shock. Our research group is studying molecules expressed in breast tumors that can be associated with sensitivity/resistance to chemotherapy.Fil: Vargas Roig, Laura Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

Comprehensive transcriptomic analysis of heat shock proteins in the molecular subtypes of human breast cancer

Background: Heat Shock Proteins (HSPs), a family of genes with key roles in proteostasis, have been extensively associated with cancer behaviour. However, the HSP family is quite large and many of its members have not been investigated in breast cancer (BRCA), particularly in relation with the current molecular BRCA classification. In this work, we performed a comprehensive transcriptomic study of the HSP gene family in BRCA patients from both The Cancer Genome Atlas (TCGA) and the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) cohorts discriminating the BRCA intrinsic molecular subtypes. Methods: We examined gene expression levels of 1097 BRCA tissue samples retrieved from TCGA and 1981 samples of METABRIC, focusing mainly on the HSP family (95 genes). Data were stratified according to the PAM50 gene expression (Luminal A, Luminal B, HER2, Basal, and Normal-like). Transcriptomic analyses include several statistical approaches: differential gene expression, hierarchical clustering and survival analysis. Results: Of the 20,531 analysed genes we found that in BRCA almost 30% presented deregulated expression (19% upregulated and 10% downregulated), while of the HSP family 25% appeared deregulated (14% upregulated and 11% downregulated) (|fold change| > 2 comparing BRCA with normal breast tissues). The study revealed the existence of shared HSP genes deregulated in all subtypes of BRCA while other HSPs were deregulated in specific subtypes. Many members of the Chaperonin subfamily were found upregulated while three members (BBS10, BBS12 and CCTB6) were found downregulated. HSPC subfamily had moderate increments of transcripts levels. Various genes of the HSP70 subfamily were upregulated; meanwhile, HSPA12A and HSPA12B appeared strongly downregulated. The strongest downregulation was observed in several HSPB members except for HSPB1. DNAJ members showed heterogeneous expression pattern. We found that 23 HSP genes correlated with overall survival and three HSP-based transcriptional profiles with impact on disease outcome were recognized. Conclusions: We identified shared and specific HSP genes deregulated in BRCA subtypes. This study allowed the recognition of HSP genes not previously associated with BRCA and/or any cancer type, and the identification of three clinically relevant clusters based on HSPs expression patterns with influence on overall survival.Fil: Zoppino, Felipe Carlos Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Guerrero Gimenez, Martin Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Castro, Gisela Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

The involvement of heat shock proteins and related molecules in the resistance to therapies in breast and gynecologic cancer

The HSP response is implicated in conferring to breast and gynecologic malignancies different sensitivities to anticancer therapies including chemotherapy, endocrine therapy and immunotherapy (weare in the need of more studies about radiotherapy). The heat shock proteins are mainly implicated in cell death mechanisms, in cell differentiation including epithelial-mesenchymal transition, in tumordormancy, in angiogenesis, metastasis formation, and in the escape of immunosurveillance. Considering the ample functions where the HSPs are implicated and that the HSP response is quite complex it is not surprising that the HSP response affects the anticancer therapies. Several of the HSPs have different predominant roles according to the molecular partners with which they interact, thus it is difficult to dissect the molecular mechanisms to find the sensitivity to the therapies. In this review we present the implications of some the major HSPs (HSP27, HSP70 and HSP90) with drug resistance and present some of the main partners that are also implicated in drug resistance like p53, PTEN and MDR. We have given priority to the incorporation of clinical data where the HSPs have been studied using standard chemotherapies and new therapeutic strategies. It is clear that in order to have a significant understanding of the degree of drug resistance/sensitivity presented by a particular patient we need to examine the molecular status of several key molecular markers involved in the drug resistance pathways and that in this context the study of the HSP response should be incorporated. One of the other major problems in this field is that an inhibitor of one particular HSP will not be enough to achieve a significant anticancer response. Now that we know the complexity of this field we need to design strategies aiming to inhibit several molecular HSP pathways simultaneously without significantly affecting the normal cells, this is the principal challenge for the near future.Fil: Cuello Carrión, Fernando Darío. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Fanelli, Mariel Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Castro, Gisela Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Cayado Gutiérrez, Niubys de Los Milagros. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

In MMTV-Her-2/neu transgenic mammary tumors the absence of caveolin-1−/− alters PTEN and NHERF1 but not β-catenin expression

In a recent study, we have shown that in mammary tumors from mice lacking the Cav-1 gene, there are alterations in specific heat shock proteins as well as in tumor development. With this in mind, we have now investigated other proteins in the same mammary mouse tumor model (Her-2/neu expressing mammary tumors from Cav-1 wild type and Cav-1 null mice), to further comprehend the complex tumor-stroma mechanisms involved in regulating stress responses during tumor development. In this tumor model the cancer cells always lacked of Cav-1, so the KO influenced the Cav-1 in the stroma. By immunohistochemistry, we have found a striking co-expression of β-catenin and Her-2/neu in the tumor cells. The absence of Cav-1 in the tumor stroma had no effect on expression or localization of β-catenin and Her-2/neu. Both proteins appeared co-localized at the cell surface during tumor development and progression. Since Her-2/neu activation induces MTA1, we next evaluated MTA1 in the mouse tumors. Although this protein was found in numerous nuclei, the absence of Cav-1 did not alter its expression level. In contrast, significantly more PTEN protein was noted in the tumors lacking Cav-1 in the stroma, with the protein localized mainly in the nuclei. P-Akt levels were relatively low in tumors from both Cav-1 WT and Cav-1 KO mice. There was also an increase in nuclear NHERF1 expression levels in the tumors arising from Cav-1 KO mice. The data obtained in the MMTV-neu model are consistent with a role for Cav-1 in adjacent breast cancer stromal cells in modulating the expression and localization of important proteins implicated in tumor cell behavior.Fil: Cuello Carrión, Fernando Darío. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Cayado Gutiérrez, Niubys de Los Milagros. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Natoli, Anthony L. . Peter MacCallum Cancer Centre. Metastasis Research Laboratory; AustraliaFil: Restall, Christina. Peter MacCallum Cancer Centre. Metastasis Research Laboratory; AustraliaFil: Anderson, Robin L.. Peter MacCallum Cancer Centre. Metastasis Research Laboratory; AustraliaFil: Nadin, Silvina Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Alvarez Olmedo, Daiana Gisela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Castro, Gisela N.. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Gago, Francisco E.. Facultad de Ciencias Médicas. Universidad Nacional de Cuyo; ArgentinaFil: Fanelli, Mariel Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

Heat shock proteins: Stress proteins with Janus-like properties in cancer

Heat shock proteins (HSPs) were first identified as stress proteins that confer resistance to physical stresses such as elevated temperatures in all cellular organisms. HSPs are rapidly elevated after stress and confer a temperature resistant phenotype. Temperature resistance is dependent on the ability of HSPs to function as molecular chaperones and prevent aggregation and on the capacity of Hsp27 and Hsp70 to act as wide spectrum inhibitors of the cell death pathways. HSP expression becomes deregulated in cancer leading to elevated expression. Elevated HSP expression promotes cancer by inhibiting programmed cell death (Hsp27, Hsp70) and by promoting autonomous growth (Hsp90) and leads to resistance to chemotherapy and hyperthermia. Tumor HSPs have another property that can be exploited in therapy. They are immunogenic and can be used to form the basis of anticancer vaccines. Elevation in HSP levels may thus have competing effects in tumor growth, being required for tumor cell survival but conferring a hazard for cancer cells due to their immunogenic properties. This dichotomy is also reflected by the approaches used to target HSP in therapy. Pharmacological approaches are being employed to inhibit activity or expression of tumor HSP. Immunological approaches aim at increasing HSP levels in cells and tissues with the aim of increasing tumor antigen presentation to the immune system.Fil: Calderwood, Stuart K.. Harvard Medical School; Estados UnidosFil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentina. Fundación Argentina para la Investigación del Cáncer; Argentin

The reality of scientific research in Latin America; an insider’s perspective

There is tremendous disparity in scientific productivity among nations, particularly in Latin America. At first sight, this could be linked to the relative economic health of the different countries of the region, but even large and relatively rich Latin American countries do not produce a good level of science. Although Latin America has increased the number of its scientists and research institutions in recent years, the gap between developed countries and Latin American countries is startling. The prime importance of science and technology to the development of a nation remains unacknowledged. The major factors contributing to low scientific productivity are the limited access to grant opportunities, inadequate budgets, substandard levels of laboratory infrastructure and equipment, the high cost and limited supply of reagents, and inadequate salaries and personal insecurity of scientists. The political and economic instability in several Latin America countries results in a lack of long-term goals that are essential to the development of science. In Latin America, science is not an engine of the economy. Most equipment and supplies are imported, and national industries are not given the incentives to produce these goods at home. It is a pity that Latin American society has become accustomed to expect new science and technological developments to come from developed countries rather than from their own scientists. In this article, we present a critical view of the Latin American investigator’s daily life, particularly in the area of biomedicine. Too many bright young minds continue to leave Latin America for developed countries, where they are very successful. However, we still have many enthusiastic young graduates who want to make a career in science and contribute to society. Governments need to improve the status of science for the sake of these young graduates who represent the intellectual and economic future of their countries.Fil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Delgado, Gabriela. Universidad Nacional de Colombia; Colombi

Participation of heat shock proteins in DNA repair mechanisms in cancer

Heat shock proteins (HSPs) are well known as molecular chaperones, playing important roles in cellular metabolism, escorting other proteins during aggregation, disaggregation, folding, and unfolding. They have been classified in families according to their molecular weight, i.e. HSPA (HSP70) and HSPH (HSP110), HSP90/HSPC, HSPD1 (HSP60), DNAJ (HSP40), and HSPB (small heat shock proteins including HSP27). HSPs are produced under normal conditions (constitutive) and in response to various stressful conditions/agents such as heat (inducible form). Several HSPs have been involved in cytoprotection, having antiapoptotic roles, and in addition some have been involved in drug resistance to antineoplastic drugs. More recently, it has been also studied the relationship of the HSPs with DNA repair proteins. DNA is constantly subject to numerous insults from endogenous sources (cellular metabolism) and exogenous sources (environmental agents), if this damage is not corrected can lead to genome instability and cancer. Fortunately, our cells count with several DNA-repair pathways to correct the DNA damage and to prevent its consequences. Although the participation of HSPs in DNA repair has received little attention, they are now receiving more interest as possible targets for cancer therapy. Here, we review the participation of HSPs in DNA repair pathways and their implications in cancer therapy and drug sensitivity. Some of the HSPs can travel to the nucleus and it is clear that although the HSPs are not capable of repairing the DNA damages by themselves, they efficiently contribute to the different mechanisms of DNA repair as part of their molecular chaperone capabilities, interacting with DNA repair proteins producing their stimulation and reactivation.Fil: Nadin, Silvina Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentin

A silver staining method for single-cell gel assay

The single-cell gel assay (comet assay) is a very useful microelectrophoretic technique for evaluation of DNA damage and repair in individual cells. Usually, the comets are visualized and evaluated with fluorescent DNA stains. This staining requires specific equipment (e.g., a high-quality fluorescence microscope), the slides must be analyzed immediately, and they cannot be stored for long periods of time. Here we describe, using human lymphocytes, some modifications of the silver staining for comets that significantly increase the sensitivity/reproducibility of the assay. This silver staining was compared with fluorescence staining and commercial silver stains.The single-cell gel assay (comet assay) is a very useful microelectrophoretic technique for evaluation of DNA damage and repair in individual cells. Usually, the comets are visualized and evaluated with fluorescent DNA stains. This staining requires specific equipment (e.g., a high-quality fluorescence microscope), the slides must be analyzed immediately, and they cannot be stored for long periods of time. Here we describe, using human lymphocytes, some modifications of the silver staining for comets that significantly increase the sensitivity/reproducibility of the assay. This silver staining was compared with fluorescence staining and commercial silver stains.Fil: Nadin, Silvina Beatriz. Fundación Argentina para la Investigación del Cáncer; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Laboratorio de Reproducción y Lactancia (i); ArgentinaFil: Nadin, Silvina Beatriz. Fundación Argentina para la Investigación del Cáncer; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Laboratorio de Reproducción y Lactancia (i); ArgentinaFil: Vargas Roig, Laura Maria. Fundación Argentina para la Investigación del Cáncer; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Laboratorio de Reproducción y Lactancia (i); ArgentinaFil: Vargas Roig, Laura Maria. Fundación Argentina para la Investigación del Cáncer; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Laboratorio de Reproducción y Lactancia (i); ArgentinaFil: Ciocca, Daniel Ramon. Fundación Argentina para la Investigación del Cáncer; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Laboratorio de Reproducción y Lactancia (i); ArgentinaFil: Ciocca, Daniel Ramon. Fundación Argentina para la Investigación del Cáncer; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Laboratorio de Reproducción y Lactancia (i); Argentin

Heat shock proteins and heat shock factor 1 in carcinogenesis and tumor development: an update

Heat shock proteins (HSP) are a subset of the molecular chaperones, best known for their rapid and abundant induction by stress. HSP genes are activated at the transcriptional level by heat shock transcription factor 1 (HSF1). During the progression of many types of cancer, thisheat shock transcriptional regulon becomes co-opted by mechanisms that are currently unclear, although evidently triggered in the emerging tumor cell. Concerted activation of HSF1 and the accumulation of HSPs then participate in many of the traits that permit the malignant phenotype. Thus, cancers of many histologies exhibit activated HSF1 and increased HSP levels that may help to deter tumor suppression and evade therapy in the clinic. We review here the extensive work that has been carried out and is still in progress aimed at (1) understanding the oncogenic mechanisms by which HSP genes are switched on, (2) determining the roles of HSF1/HSP in malignant transformation and (3) discovering approaches to therapy based on disrupting the influence of the HSF1-controlled transcriptome in cancer.Fil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Arrigo, Andre Patrick. Cancer Research Center of Lyon, Apoptosis Cancer and Development; Francia. Claude Bernard University; FranciaFil: Calderwood, Stuart K.. Beth Israel Deaconess Medical Center. Department of Radiation Oncology; Estados Unido

Effects of hyperthermia on Hsp27 (HSPB1), Hsp72 (HSPA1A) and DNA repair proteins hMLH1 and hMSH2 in human colorectal cancer hMLH1-deficient and hMLH1-proficient cell lines

Purpose: The objective of the present study was to examine the consequences of a mild hyperthermia in human tumour cell lines deficient and proficient in the DNA mismatch repair system (MMR) to advance our understanding on the relationship between MMR and heat shock proteins (HSPs). Materials and methods: The human colon carcinoma cell lines HCT116 (parent cells), HCT116+ch2 (MMR-deficient), and HCT116+ch3 (MMR-proficient) were used. Cells were incubated at 41°C and 42°C for 1h and then at 37°C for 4 and 24h. The expression of Hsp27 and Hsp72 was evaluated by immunocytochemistry. Hsp27, Hsp72, hMLH1 and hMSH2 levels were assessed by western blotting in nuclear and cytoplasmic fractions. The alkaline comet assay was used to evaluate the DNA damage. Results: The mild hyperthermia significantly increased the protein expression levels of Hsp27 and Hsp72 in all cell lines, which was higher in the cytoplasm and nucleus of HCT116+ch3 cells. We also observed that heat induced translocation of hMLH1 and hMSH2 proteins from the nucleus to the cytoplasm in HCT116+ch3 cells. The comet assay revealed that HCT116 parent cells were more resistant to heat-induced DNA damage. However, the MMR-proficient and deficient cell lines repaired the DNA damage at the same rate. Conclusions: The present study demonstrates that hyperthermia induced the nuclear accumulation of Hsp27 and Hsp72 and affected the subcellular localisation of hMLH1 and hMSH2 in HCT116+ch3 cells. Our findings suggest that the MMR system is not a direct determining factor for the different heat shock response in HCT116 cells.Fil: Nadin, Silvina Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Cuello Carrión, Fernando Darío. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Sottile Fleury, Mayra Lis. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Ciocca, Daniel Ramon. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; ArgentinaFil: Vargas Roig, Laura Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto de Medicina y Biología Experimental de Cuyo; Argentina. Universidad Nacional de Cuyo. Facultad de Ciencias Médicas; Argentin