Role of Humanin in the response of cancer cells to chemotherapy

ZUCCATO, CAMILA; MORENO AYALA, MARIELA A.; GOTTARDO, M. FLORENCIA; PIDRE, MATIAS L.; ASAD, ANTONELA S.; NICOLA CANDIA, ALEJANDRO J.; ROMANOWSKI, VICTOR; SEILICOVICH, ADRIANA; CANDOLFI, M

Congreso; Reunión conjunta de sociedades de biociencias; LXII Reunión Anual de la Sociedad Argentina de Investigación Clínica (SAIC).; 2017

Humanin (HN) is a mitochondrial-derived peptide with potent cytoprotective action in many cell types. Although HN can protect normal cells against toxic effects of chemotherapy, the role of this peptide in tumor pathogenesis is not well understood. We have previously observed that recombinant HN inhibits the response of triple negative breast cancer (TNBC) cells to cytotoxic stimuli, facilitating tumor progression and chemoresistance in vivo. Here we aimed to evaluate whether chemotherapy modulates the expression of HN in cancer cell types that are characterized by their chemoresistance: TNBC and glioblastoma (GBM) cells. We evaluated the expression of HN in murine 4T1 TNBC cells and in human U251 GBM cells by flow cytometry following serum deprivation and chemotherapy with Doxorubicin (DOXO), Cisplatin (CP) or Temozolomide (TMZ). Expression of HN in 4T1 TNBC cells was upregulated by DOXO (500 nM) and CP (1 ìM, p<0.05), but not by serum deprivation. In U251 GBM cells, the expression of HN was increased by CP (5 ìM) and TMZ (15 ìM), and at a lesser extent by serum deprivation (p<0.05). We next evaluated the role of endogenous HN in the apoptotic response of TNBC cells, using a plasmid encoding a short hairpin RNA to block HN (pUC.shHN). To readily assess transduction efficiency, the plasmid also encodes the red fluorescent protein dTomato as a reporter gene. Transfection of TNBC 4T1 tumor cells with pUC.shHN increased the percentage of apoptotic cells when compared to cells transfected with control plasmid, as assessed by flow cytometry after staining with propidium iodide (p<0.05). When transfected 4T1 cells were incubated with different concentrations of DOXO, pUC.shHN increased their chemosensitivity, reducing their clonogenic capacity when incubated with Doxo (p<0.05). Our findings indicate that blockade of HN expression could constitute a therapeutic strategy to improve the efficacy of chemotherapy in chemoresistant cancer cells.