Synthesis of mercaptan-based brevetoxin scavengers and evaluation of their ability to interfere with binding to voltage-gated sodium channel and reduce cytotoxicity

Mayra Tabares-Beltran¹, Jennifer R. McCall², Kathleen S. Rein³

1 Department of Chemistry and Biochemistry, Florida International University, Miami, FL 33199
2 School of Nursing, University of North Carolina Wilmington, Wilmington, NC 28403.
3 Department of Marine & Earth Sciences, Florida Gulf Coast University, Miami, FL 33965.

Brevetoxins (PbTx), neurotoxins produced by K. brevis that bloom during red tide events, can cause marine mammal poisoning, mortalities, and deaths of fish, sea birds, and turtles and exhibit adverse human health effects. Brevetoxins can bind to the voltage-gated sodium channel (VGSC) and inhibit thioredoxin reductase (TrxR), a major regulator of cellular redox homeostasis. Currently, no treatments are available for the neurotoxic or respiratory effects of brevetoxins on wildlife or humans. Synthesis and evaluation of a series of mercaptans as potential “antitoxins”, based on the hydrophilic FDA-approved acrolein scavenger, cysteamine, that may react with and thereby detoxify PbTx-2, were conducted. Sulfonamides and amide derivatives were prepared and characterized by HRMS, ¹H, and ¹³C NMR. Partition coefficients (Log K_ow) of reduced thiols were determined in an octanol-water system, and reactivity with PbTx-2 was assessed using FT-ICR-MS at different time points. Log Kow values showed an increase in lipophilicity of all cysteamine derivatives, which could improve drug uptake and half-life compared with cysteamine. Monosubstituted and disubstituted PbTx-2 adducts were detected in most of the tested reactions. To assess the ability of the potential new therapeutic mercaptans for the ability to act as a PbTx-2 “antitoxins” we performed fluorescent receptor binding assay (RBA) and cytotoxicity assays. Most of the compounds showed the ability to reduce PbTx-2 binding affinity on the RBA, exhibiting some level of “antitoxin” properties. PbTx-2 modified with bulky naphthyl and    t-butyl benzyl, and aliphatic groups have shown similar binding affinity compared to      PbTx-2. Aromatic sulfonamides with  p-bromo and p-t-butyl, showed the most statistically significant decrease in PbTx-2 cytotoxicity at 48 hours in SJCRH30 cells. Moreover, there was an inverse proportional relationship between binding affinity and cytotoxicity, as observed by conjugated adducts with p-t-butyl benzene sulfonamides. Our studies have shown that PbTx-2 conjugation with some synthesized mercaptan drugs could interfere with the binding to VGSC, and lower PbTx-2 cytotoxicity, thus providing a set of compounds that could serve as an emerging treatment for brevetoxicosis in wildlife and humans.

Hi, I would like to kindly request that my oral presentation be scheduled for Saturday, June 1st. My co-advisor, Dr. Yuan Liu is considering driving back to Miami on the same day.

I appreciate your help, thank you.