Abstract:

Despite NK cells being originally identified because of their ability to kill tumor cells in vitro, only limited information is available on NK cells infiltration of malignant tumors, especially in humans. NK cells infiltrating human colorectal carcinomas (CRCs) were analyzed to identify their potential protective role in an antitumor immune response. The expression and function of relevant molecules were analyzed from different sources, comparing tumor-associated NK cells (TANKs) with autologous peripheral blood NK cells (PB-NKs) from CRC patients-the latter in comparison with PB-NKs from normal donors. TANKs displayed a profound alteration of their phenotype with a drastic reduction of NK cell receptor expression. Co-culture experiments showed that CRC cells produce modulation in NK phenotype and functionality. Moreover, PB-NKs from CRC patients also exhibited an altered phenotype and profound defects in the ability to activate degranulation and IFN-γ production. For the first time, TANK and PB-NK cells from CRC patients have been characterized. It is shown that they are not capable of producing relevant cytokines and degranulate. Taken together, our results suggest that NK cells from CRC patients present alterations of phenotype and function therefore supporting the progression of cancer.

Author affiliation: Rocca, Yamila Sol. Fundacion Cancer. Centro de Investigaciones Oncologicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina

Author affiliation: Roberti, Maria Paula. Fundacion Cancer. Centro de Investigaciones Oncologicas; Argentina

Author affiliation: Arriaga, Juan Martín. Fundacion Cancer. Centro de Investigaciones Oncologicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina

Author affiliation: Amat, Mora. Instituto MÉdico Especializado Alexander Fleming;

Author affiliation: Bruno, Luisina. Instituto MÉdico Especializado Alexander Fleming;

Author affiliation: Pampena, María Betina. Fundacion Cancer. Centro de Investigaciones Oncologicas; Argentina

Author affiliation: Huertas, Eduardo. Instituto Médico Especializado Alexander Fleming;

Author affiliation: Sánchez Loria, Fernando. Instituto Médico Especializado Alexander Fleming;

Author affiliation: Pairola, Alejandro. Instituto Médico Especializado Alexander Fleming;

Author affiliation: Bianchini, Michele. Fundacion Cancer. Centro de Investigaciones Oncologicas; Argentina

Author affiliation: Mordoh, Jose. Fundación Instituto Leloir; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina

Author affiliation: Levy, Estrella Mariel. Fundacion Cancer. Centro de Investigaciones Oncologicas; Argentina

Abstract:

Metallothioneins are a family of small, cysteine-rich proteins with many functions. Immunohistochemical evaluation of all metallothionein 1 + 2 isoforms in colorectal tumors has demonstrated an important down-regulation compared with normal tissue, although its prognostic significance is unclear. Moreover, the contribution of individual isoforms to overall metallothionein down-regulation is not known. To address these important issues, we analyzed the messenger RNA expression levels of all functional metallothionein 1 + 2 isoforms by quantitative reverse transcription polymerase chain reaction in 22 pairs of normal and tumor-microdissected epithelia and correlated these to the overall immunohistochemical protein expression. Our results showed that 5 isoforms (MT1G, 1E, 1F, 1H, and 1M) were lost during the transition from normal mucosa to tumor, whereas MT1X and MT2A were less down-regulated, and their expression was correlated with overall protein positivity. Second, we showed that MT1G hypermethylation occurred in cell lines and in 29% of tumor samples, whereas histone deacetylase inhibitors are able to induce most isoforms. Furthermore, we analyzed by immunohistochemistry 107 normal mucosae, 25 adenomas, 81 carcinomas, and 19 lymph node metastases to evaluate metallothionein expression during different stages of cancer development and to assess its relationship to patient survival. A lower immunohistochemical expression was associated with poorer survival, although it was not an independent predictor. Overall, this study identifies for the first time the relevant metallothionein isoforms for colorectal cancer progression, supports the concept that their loss is associated with worse prognosis, and suggests 2 mechanisms for epigenetic repression of metallothionein expression in colorectal tumors.

Author affiliation: Arriaga, Juan Martín. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentina. Fundación Cáncer. Centro de Investigaciones Oncológicas; Argentina

Author affiliation: Levy, Estrella Mariel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fundación Cáncer. Centro de Investigaciones Oncológicas; Argentina

Author affiliation: Bravo, Alicia Inés. Provincia de Buenos Aires. Ministerio de Salud. Hospital Interzonal de Agudos "Eva Perón"; Argentina

Author affiliation: Bayo, Sergio Morales. Municipio de Vicente López. Hospital Municipal "Prof. Dr Houssay"; Argentina

Author affiliation: Amat, Mora. Instituto Médico Especializado "Alexander Fleming"; Argentina

Author affiliation: Aris, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentina

Author affiliation: Hannois, Adrián. Provincia de Buenos Aires. Ministerio de Salud. Hospital Interzonal de Agudos "Eva Perón"; Argentina

Author affiliation: Bruno, Luisina Inés. Instituto Médico Especializado "Alexander Fleming"; Argentina

Author affiliation: Roberti, Maria Paula. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fundación Cáncer. Centro de Investigaciones Oncológicas; Argentina

Author affiliation: Sánchez Loria, Fernando. Instituto Médico Especializado "Alexander Fleming"; Argentina

Author affiliation: Pairola, Alejandro. Instituto Médico Especializado "Alexander Fleming"; Argentina

Author affiliation: Huertas, Eduardo. Instituto Médico Especializado "Alexander Fleming"; Argentina

Author affiliation: Mordoh, Jose. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentina. Instituto Médico Especializado "Alexander Fleming"; Argentina. Fundación Cáncer. Centro de Investigaciones Oncológicas; Argentina

Author affiliation: Bianchini, Michele. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Fundación Cáncer. Centro de Investigaciones Oncológicas; Argentina