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Immunological hallmarks of cis-DDP-resistant Lewis lung carcinoma cells

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Abstract

Purpose

Tumor cell resistance to platinum-based chemotherapeutic agents is one of the major hurdles to successful cancer treatment with these drugs, and is associated with alterations in tumor cell immune evasion and immunomodulatory properties. Immunocyte targeting is considered as a relevant approach to fight drug-resistant cancer. In this study, immunological hallmarks of cis-DDP-resistant Lewis lung carcinoma cells (LLC/R9) were investigated.

Methods

Immunological features of LLC/R9 cells cultured in vitro in normoxic and hypoxic conditions as well as of those that were grown in vivo were examined. The expression of immunologically relevant genes was evaluated by RT-PCR. Tumor cell susceptibility to the macrophage contact tumoricidal activity and NK-mediated cytolysis was investigated in MTT test. TNF-α-mediated tumor cell apoptosis as well as macrophage phagocytosis, oxidative metabolism, and CD206 expression after the treatment with conditioned media from normoxic and hypoxic tumor cells were studied by flow cytometry. Flow cytometry was also used to characterize dendritic cell maturity.

Results

When growing in vitro, LLC/R9 were characterized by slightly increased immunosuppressive cytokine gene expression. Transition to in vivo growth was associated with the enhancement of transcription of these genes in tumor cells. LLC/R9 cells had lowered sensitivity to contact-dependent macrophage-mediated cytotoxicity and to the TNFα-mediated apoptosis in vitro. Conditioned media from hypoxic LLC/R9 cells stimulated reactive oxygen species generation and CD206 expression in non-sensitized macrophages. Acquisition of drug resistance by LLC/R9 cells was associated with their increased sensitivity to NK-cell-mediated cytolysis. Meanwhile, the treatment of LLCR/9-bearing animals with generated ex vivo and loaded with LLC/R9 cell-lysate dendritic cells (DCs) resulted in profoundly enhanced tumor metastasizing.

Conclusion

Decreased sensitivity to macrophage cytolysis, polarizing effect on DCs maturation along with increased susceptibility to NK-cell cytotoxic action promote extensive local growth of chemoresistant LLC/R9 tumors in vivo, but hamper their metastasizing.

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Correspondence to Larysa Skivka.

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Animal protocol was reviewed and approved by the IEPOR NASU and Taras Shevchenko National University animal welfare committee according to the Animal Welfare Act guidelines. Study was conducted in compliance with the standards of the Convention on Bioethics of the Council of Europe ‘Europe Convention for the Protection of Vertebrate Animals used for experimental and other scientific purposes’ (1997), the general ethical principles of animal experiments, approved by the First National Congress on Bioethics in Ukraine (September 2001) and national law (LAW OF UKRAINE no. 3447-IV) issued by the Cabinet of Ministers of Ukraine (2006).

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Fedorchuk, O., Susak, Y., Rudyk, M. et al. Immunological hallmarks of cis-DDP-resistant Lewis lung carcinoma cells. Cancer Chemother Pharmacol 81, 373–385 (2018). https://doi.org/10.1007/s00280-017-3503-6

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