Abstract 1778: 5’-nucleotidases are involved in the biology of human lung cancer cell lines

Abstract

Abstract Cytosolic-5’-nucleotidase II (cN-II) and ecto-5′-nucleotidase (CD73) are enzymes involved in the nucleotide metabolism by dephosphorylating intracellular and extracellular purine nucleotide monophosphates, respectively. Both enzymes have been shown to be involved in cancer by modifying anticancer drug activity, cancer cell biology and immune modulation. To increase the knowledge of cN-II and CD73 roles in cancer cell biology, we developed lung cancer cell models (NCI-H292) with a complete knockout of either or both enzymes using the CRISPR-Cas9 technique. These cell models are used to study cell proliferation and migration, cancer drug sensitivity, cellular behaviour and apoptosis using CFSE staining, IncuCyte relative confluence and wound healing assay, MTT assay, xCELLigence RTCA and caspase-3/7 apoptotic assay. SCID CB17 mice are also used to study the in vivo tumor growth using these cell models. Our results show that there is no significant difference in proliferation between different cell models exposed or not to different concentrations of either adenosine or AMP under normoxic and hypoxic conditions. On the other hand, using xCELLigence RTCA technique, we have observed that CD73-deficient cells have higher cell index as compared to CD73 expressing cells under basal conditions during the first 72 hours of culture. However, adenosine globally decreases the cell index of all NCI-H292 cells, unlike AMP, which influences the cell index of cN-II-deficient cells but not CD73-deficient cells. Under hypoxic condition, cN-II-deficient cells showed higher cell index than the other phenotypes in untreated conditions, whereas cells lacking CD73 showed higher sensitivity to adenosine or AMP. As compared to other phenotypes, CD73 deficient cells are also more sensitive towards vincristine but less sensitive towards mitomycin and fludarabine. These cells are also more prone to adapt to hypoxic conditions and exhibit slower migration rate than their corresponding control cells. However, the effect of adenosine and AMP on migration rate is higher on cN-II-deficient cells. Mice experiments showed no difference in in vivo tumor growth using these cell models. Overall, we show that both CD73 and cN-II are important players in the cell biology of lung cancer cells and in their response to purines. Upcoming experiments should help us understand the molecular mechanisms underlying these observed differences. Citation Format: Muhammad Zawwad Raza, Octavia Cadassou, Emeline Cros-Perrial, Alain Puisieux, Charles Dumontet, Lars Petter Jordheim. 5’-nucleotidases are involved in the biology of human lung cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1778.