Tubulin-Targeted Therapy in Melanoma Increases Cell Invasive Potential by Activation of Actomyosin Cytoskeleton - an in vitro study

Abstract

One of the most dangerous aspects of cancers is their ability to metastasize, which is the leading cause of death. Hence, it holds significance to develop therapies targeting the eradication of cancer cells, in parallel, inhibiting metastases in cells surviving the applied therapy. Here, we focused on two melanoma cell lines - WM35 and WM266-4 - representing the less and more invasive melanomas. We investigated mechanisms of cellular processes regulating the activation of actomyosin being the effect of colchicine treatment. Additionally, we investigated biophysical aspects of supplement therapy using Rho-associated protein kinase (ROCK) inhibitor (Y-27632), and myosin II inhibitor ((-)-blebbistatin), focusing on the microtubules and actin filaments. We analyzed their effect on the proliferation, migration, and invasiveness of melanoma cells, supported by studies on cytoskeletal architecture using confocal fluorescence microscopy and nanomechanics using atomic force microscopy and novel method based on constriction channels. Our results showed that colchicine inhibits the migration of most melanoma cells, while for a small cell population, it paradoxically increases their migration and invasiveness. These changes are also accompanied by the formation of stress fibers compensating for the loss of microtubules. Simultaneous administration of selected agents led to the inhibition of this compensatory effect. Collectively, our results highlighted that colchicine led to actomyosin activation and increased cancer cell invasiveness. We emphasized a cellular pathway of Rho-ROCK-dependent actomyosin contraction to be responsible for the increased invasive potential of melanoma cells in tubulin-targeted therapy. HighlightsO_LIColchicine strongly increased (trans)migration in the subpopulation of melanoma cells. C_LIO_LIColchicine disrupted tubulin but compensated for the activation of the actomyosin network. C_LIO_LIRho-ROCK-targeted or myosin-targeted therapies vanished the side-effect of colchicine. C_LIO_LIIncreased actin polymerisation correlated with the transit time in the constriction channel. C_LIO_LIElastic modulus variations reflected observed alterations in the actomyosin network. C_LI