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PO-156 Differential effect of integrin αV knockdown and cilengitide in sensitisation of triple-negative breast cancer and melanoma cell lines to microtubule poisons
  1. N Stojanović1,
  2. A Dekanić1,
  3. M Paradžik1,
  4. D Majhen1,
  5. J Ruščić1,
  6. C Supina1,
  7. M Christmann2,
  8. MT Tomicic2,
  9. A Ambriović-Ristov1
  1. 1Ruđer Bošković Institute, Division of Molecular Biology, Zagreb, Croatia
  2. 2University Medical Center Mainz, Department of Toxicology, Mainz, Germany

Abstract

Introduction Integrin signalling regulates proliferation, migration, metastasis and cell death and has been suggested as a possible target in antitumor therapy. The aim of this study was to investigate the potential of different integrin α subunits knockdown in increasing sensitivity of triple-negative breast cancer (TNBC) and melanoma cell lines to cisplatin, microtubule poisons paclitaxel (PTX) and vincristine (VCR), and mitigating metastatic potential.

Material and methods The knockdown of integrin subunits αv, β3, β5, α3 and α4 was achieved by transfection of siRNA. The sensitivity of cell lines to anticancer drugs was determined using MTT assay. For monitoring cell migration and invasion, migration or matrigel-coated invasion Transwell Cell Culture Inserts were used. Flow cytometry, western blot, and confocal microscopy were done using standard protocols.

Results and discussions We analysed the proof of concept in melanoma cell line MDA-MB-435S that expresses integrins αvβ3 and αvβ5 but not αvβ1. Following transfection with integrin-specific siRNAs integrin subunit αv knockdown was selected since it increased sensitivity to PTX and VCR, and decreased metastatic potential. We observed an integrin switching effect upon β3, β5 and α4 knockdown that led to differing changes of sensitivity to antitumor drugs and cell migration. Using a panel of TNBC (MDA-MB-231,–468 and −436) and melanoma (RPMI-7951, MeWo and A375) cell lines we showed that knockdown of integrin αv increased sensitivity to PTX and VCR and inhibited metastatic potential in MDA-MB-231, MDA-MB-468, RPMI-7951 and MeWo, while no effect was observed in MDA-MB-436 and A375 cells. To assess the importance of integrins αvβ3/β5 in sensitisation to PTX we exposed all cell lines to combination of αvβ3/β5 inhibitor cilengitide and PTX and observed increased sensitivity to PTX in MDA-MB-231, MDA-MB-436, RPMI-7951 and A375 but surprisingly decreased sensitivity in MeWo and MDA-MB-468 cell lines, implying either involvement of other integrins αv or integrin switching effect. The correlation between loss of phosphorylated focal adhesion kinase pFAK(Y397) and increased sensitivity to PTX was observed in melanoma cell lines, however, sensitisation was not achieved using pFAK(Y397) inhibitor.

Conclusion Our results highlight that knocking down integrin αv, rather than inhibiting αvβ3/β5 using cilengitide, could be more effective for increasing sensitivity to microtubule poisons and decreasing metastatic potential in TNBC and melanoma.

This is an Open Access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/

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