34P - Anti-tumor activity of recombinant human CXCR2-Fc protein

Date 05 November 2016
Event ESMO Symposium on Immuno-Oncology 2016
Session Lunch and general poster viewing
Presenter Ksenia Glukhova
Citation Annals of Oncology (2016) 27 (suppl_8): viii4-viii17. 10.1093/annonc/mdw527
Authors K.A. Glukhova1, Y.A. Trizna1, A.S. Glukhov1, V.V. Marchenkov2, G.V. Afanasieva1, A.A. Ivanov3, O.P. Popova3, T.I. Danilova3, I.P. Beletsky1, O.V. Prusakova1
  • 1Laboratory Of Cell Engineering, Institute of Theoretical and Experimental Biophysics-Russian Academy of Sciences, 142290 - Pushchino/RU
  • 2Laboratory Of Protein Physics, Institute of Protein Research of RAS, Pushchino/RU
  • 3Department Of Molecular Medicine, IM Sechenov First Moscow State Medical University, Moscow/RU

Abstract

Aim/Background

Interleukin-8 (IL-8), a proinflammatory CXC chemokine, via its cognate receptors, CXCR1 and CXCR2, is able to stimulate angiogenesis, enhance the proliferation and survival of endothelial and tumor cells as well as tumor cell migration (metastasis). Accordingly, the development of therapeutic agents, targeting IL-8 signalling, seems an urgent task. The anti-cancer potential of various small molecule antagonists and humanized monoclonal antibodies to IL-8 is currently actively investigated. In this study we present the results of investigation of the anti-tumor activity of recombinant soluble protein in which the N-terminal domain of CXCR2 is juxtaposed on the Fc-fragment of human IgG1.

Methods

Recombinant human CXCR2-Fc protein produced by HEK 293 was isolated and purified by affinity chromatography with protein A-Sepharose followed by two-step ion-changed chromatography. The physical and chemical properties of recombinant protein were studied using gel-filtration chromatography and SDS-PAGE-electrophoresis. The IL-8 binding activity of CXCR2-Fc protein was analyzed in ELISA-tests with recombinant IL-8. The cytotoxic activity of CXCR2-Fc protein was estimated in experiments with human breast adenocarcinoma MDA-MD-231, highly expressing IL-8, and the IL-8-expressing primary tumor cell cultures from breast cancer patients by MTT-test and Annexin V staining. The content of IL-8 in the supernatant of primary breast tumor cell cultures was determined using the sandwich ELISA.

Results

The electrophoretic and chromatographic analyses show that recombinant human CXCR2-Fc protein is present mostly as dimers effectively binding IL-8. The elevated level of the secreted IL-8 (up to 70 ng/ml) is found in supernatants of some primary tumor cell cultures from breast cancer patients. The MTT assay and Annexin V staining show that CXCR2-Fc protein is capable of inducing the death of MDA-MD-231 cells as well as some primary tumor cells expressing IL-8.

Conclusions

The results obtained indicate that recombinant human CXCR2-Fc protein may be regarded as the promising anti-cancer agent. This work was supported by a grant from the Ministry of Education and Science of the Russian Federation (# 14.604.21.0025, ID RFMEFI60414X0025).

Clinical trial identification


Legal entity responsible for the study

Institute of Theoretical and Experimental Biophysics of RAS

Funding

Ministry of Education and Science of the Russian Federation (grant # 14.604.21.0025, ID RFMEFI60414X0025)

Disclosure

All authors have declared no conflicts of interest.