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Voltage-Gated Sodium Channel Na_V1.7 Inhibitors with Potent Anticancer Activities in Medullary Thyroid Cancer Cells.

ABSTRACT: Our results from quantitative RT-PCR, Western blotting, immunohistochemistry, and the tissue microarray of medullary thyroid cancer (MTC) cell lines and patient specimens confirm that VGSC subtype Na_V1.7 is uniquely expressed in aggressive MTC and not expressed in normal thyroid cells and tissues. We establish the druggability of Na_V1.7 in MTC by identifying a novel inhibitor (SV188) and investigate its mode of binding and ability to inhibit I_Na current in Na_V1.7. The whole-cell patch-clamp studies of the SV188 in the Na_V1.7 channels expressed in HEK-293 cells show that SV188 inhibited the I_Na current in Na_V1.7 with an IC₅₀ value of 3.6 µM by a voltage- and use-dependent blockade mechanism, and the maximum inhibitory effect is observed when the channel is open. SV188 inhibited the viability of MTC cell lines, MZ-CRC-1 and TT, with IC₅₀ values of 8.47 μM and 9.32 μM, respectively, and significantly inhibited the invasion of MZ-CRC-1 cells by 35% and 52% at 3 μM and 6 μM, respectively. In contrast, SV188 had no effect on the invasion of TT cells derived from primary tumor, which have lower basal expression of Na_V1.7. In addition, SV188 at 3 μM significantly inhibited the migration of MZ-CRC-1 and TT cells by 27% and 57%, respectively.

SUBMITTER: Pukkanasut P

PROVIDER: S-EPMC10216335 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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Voltage-Gated Sodium Channel NaV1.7 Inhibitors with Potent Anticancer Activities in Medullary Thyroid Cancer Cells.

Pukkanasut Piyasuda P Whitt Jason J Guenter Rachael R Lynch Shannon E SE Gallegos Carlos C Rosendo-Pineda Margarita Jacaranda MJ Gomora Juan Carlos JC Chen Herbert H Lin Diana D Sorace Anna A Jaskula-Sztul Renata R Velu Sadanandan E SE

Cancers 20230517 10

Our results from quantitative RT-PCR, Western blotting, immunohistochemistry, and the tissue microarray of medullary thyroid cancer (MTC) cell lines and patient specimens confirm that VGSC subtype NaV1.7 is uniquely expressed in aggressive MTC and not expressed in normal thyroid cells and tissues. We establish the druggability of NaV1.7 in MTC by identifying a novel inhibitor (SV188) and investigate its mode of binding and ability to inhibit INa cur ...[more]

PMID: 37345144

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Project description:The human TE671 cell line was originally used as a model of medulloblastoma but has since been reassigned as rhabdomyosarcoma. Despite the characterised endogenous expression of voltage-sensitive sodium currents in these cells, the specific voltage-gated sodium channel (VGSC) subtype underlying these currents remains unknown. To profile the VGSC subtype in undifferentiated TE671 cells, endpoint and quantitative reverse transcription-PCR (qRT-PCR), western blot and whole-cell patch clamp electrophysiology were performed. qRT-PCR profiling revealed that expression of the SCN9A gene was ∼215-fold greater than the SCN4A gene and over 400-fold greater than any of the other VGSC genes, while western blot confirmed that the dominant SCN9A RNA was translated to a protein with a molecular mass of ∼250 kDa. Elicited sodium currents had a mean amplitude of 2.6 ± 0.7 nA with activation and fast inactivation V50 values of -31.9 ± 1.1 and -69.6 ± 1.0 mV, respectively. The currents were completely and reversibly blocked by tetrodotoxin at concentrations greater than 100 nm (IC50 = 22.3 nm). They were also very susceptible to the NaV 1.7 specific blockers Huwentoxin-IV and Protoxin-II with IC50 values of 14.6 nm and 0.8 nm, respectively, characteristic of those previously determined for NaV 1.7. Combined, the results revealed the non-canonical and highly dominant expression of NaV 1.7 in the human TE671 rhabdomyosarcoma cell line. We show that the TE671 cell line is an easy to maintain and cost-effective model for the study of NaV 1.7, a major target for the development of analgesic drugs and more generally for the study of pain. KEY POINTS: Undifferentiated TE671 cells produce a voltage-sensitive sodium current when depolarised. The voltage-gated sodium channel isoform expressed in undifferentiated TE671 cells was previously unknown. Through qRT-PCR, western blot and toxin pharmacology, it is shown that undifferentiated TE671 cells dominantly (>99.5%) express the NaV 1.7 isoform that is strongly associated with pain. The TE671 cell line is, therefore, a very easy to maintain and cost-effective model to study NaV 1.7-targeting drugs.

Dataset Information

Voltage-Gated Sodium Channel Na_V1.7 Inhibitors with Potent Anticancer Activities in Medullary Thyroid Cancer Cells.

Publications

Voltage-Gated Sodium Channel Na<sub>V</sub>1.7 Inhibitors with Potent Anticancer Activities in Medullary Thyroid Cancer Cells.

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