Volume-regulated Cl and oxidation-activated K channels in invasiveness and immunotherapy efficiency in human melanoma
The Biophysics Institute in Genoa has recently been granted funding by the AIRC to carry out a research project focusing on “Volume-regulated Cl and oxidation-activated K channels in invasiveness and immunotherapy efficiency in human melanoma”. The project leader is Dr. Michael Pusch.
Background: Chloride and potassium ion channels involved in volume regulation likely play an important role in the invasiveness of melanoma cells and in the homing of T-lymphocytes, both processes related to cell migration and volume regulation. However the specific role of the newly discovered volume-regulating LRRC8 proteins forming the VRAC channel is unknown. In addition, we have identified a novel oxidation activated K channel, KROS, in human melanoma cells. The currents mediated by this channel are of extremely large amplitude and their role in melanoma is unknown. These findings open a new interesting perspective in the development of new strategies to target circulating tumor cells (CTC) to reduce their metastatic potential that is closely related to the ability to migrate.
Hypothesis: In the present project we plan to investigate the role of LRRC8 mediated VRAC channels and KROS channels in tumor invasiveness using as model a highly metastatic tumor such as melanoma. Since the role of the adaptive immune system is critical in the acquisition of an aggressive phenotype we have also included in our study the involvement of LRRC8 for the homing of T-lymphocytes.
Aims: The overall idea of the present project is to obtain information on the role of LRRC8 and KROS in the processes of melanoma cell invasion and T-cell homing, and to target LRRC8 proteins developing synthetic peptides designed by computational studies to interfere with the capability of the cells to spread and invade.
Experimental Design: We intend to characterize the expression profile of LRRC8 channels (hexamers composed of 5 different subunits with variable stoichiometry) in melanoma and T-lymphocytes, to determine the molecular identity of KROS and its expression profile in melanoma cells. Next, we will use melanoma and T-cells in which the genes encoding LRRC8/KROS have been knocked down and/or knocked out to test their importance for melanoma invasiveness and for T-cell homing. Finally we intend to develop rationally designed synthetic peptides targeting LRRC8 proteins and test their ability to interfere with melanoma invasiveness and T-cell homing.
Expected Results: The project will reveal if LRRC8 proteins (and which subunit combinations) and/or KROS channels are involved in melanoma invasiveness, and which role LRRC8 proteins play in T-cell homing. In addition the project will provide highly specific molecular tools (rationally designed peptides) targeted at LRRC8 proteins and able to interfere with melanoma invasion.
Impact On Cancer: VRAC has been implicated in many aspects of cancer. However its role in melanoma invasiveness remains unknown. Our results will be of importance in determining the role of specific LRRC8 subunits in melanoma invasiveness and T lymphocyte homing. In addition we will explore the importance of the large amplitude KROS current that we have identified in melanoma Igr39 cells (see preliminary results). The identification of specific chemical tools targeting LRRC8 proteins (activators and inhibitors) could pave the way for novel chemotherapeutic strategies for melanoma and for the enhancement of the efficiency of immunotherapy.
The research will be conducted in the Biophysics Institute of the Cnr and at the University of Milan (Oncolab, www.oncolab.unimi.it).