# Cross platform analysis of drug targets and toxicity of bath salts

> **NIH NIH U01** · UNIVERSITY OF TOLEDO HEALTH SCI CAMPUS · 2024 · $556,470

## Abstract

Project Summary/Abstract
 The United States is in the midst of a substance abuse and mental health epidemic. Over the past decade,
abuse of psychostimulant (and sometimes hallucinogenic) cathinones, often referred to as “bath salts”, and
variants such as ‘flakka’, has become increasing popular in America’s youth. Although there has been substantial
research into the mechanisms underlying the addictive and adverse effects of some of these agents, such as
3,4-methylenedioxymethamphetamine (MDMA), a wide variety of variants are available for sale on the internet.
Furthermore, these represent only a small fraction of conceivable structures which can be produced on kilograms
scale for sale online. Gaining a more complete understanding of the drug targets and toxicity associated with
these agents is an important step to treat substance abuse disorders and episodic toxicity.
 The over-arching goal of our work is to unite whole system level analysis involving the neurobehavioral
effects of psychoactive agents with the study of binding partners at the molecular level. The focus of this
proposal is to implement a novel in vivo photoaffinity labeling (PAL) approach for cross platform analysis of
psychoactive agents, with an emphasis on structures related methylenedioxypyrovalerone (MDPV) and α-
pyrrolidinovalerophenone (α-PVP). In Aim 1 we will synthesize a carefully designed set of probes to be
cataloged in Aim 2 assays. This includes stereoselective synthesis of PAL probes, structure activity relationship
(SAR) controls, and stable isotope labeled standards for LC-MS studies in Aim 2C. The synthesized library will
be assayed for 1) receptor binding profile; 2) effect on monoamine uptake and release in rat synaptosomes;
and 3) toxicity in multiple cell lines HepG2 (liver), SHSY-5Y (neuronal), AC16 (cardiac myocyte) and PC-12
(neuronal [rat]). In Aim 2C, active compounds (and select inactive controls) will be studied in an adult zebrafish
behavioral paradigm utilizing the novel tank test with concurrent in vivo PAL. The concentration of each drug
will be measured by LC-MS/MS to correlate target tissue (brain) drug exposure with behavioral responses.
Select compounds will be re-examined in the Aim 2 assays using PAL for side-by-side comparison of binding
interactome of Aim 2 experimental systems versus the in vivo binding interactome. Chemical biology in Aim 3
will utilize click chemistry to visualize protein labeling via fluoroimaging of SDS PAGE gels. Proteomics will
determine protein identification after affinity purification to define the binding interactome of each probe. A
variety of competition experiments will be performed to characterize background PAL and recognize bona fide
targets. Bioinformatics will be used for network analysis to provide an unbiased comparison of the binding
interactomes for each experimental system to propose potential therapeutic targets to treat toxicity and episodic
lethality of these agents.

## Key facts

- **NIH application ID:** 10870155
- **Project number:** 5U01DA054330-03
- **Recipient organization:** UNIVERSITY OF TOLEDO HEALTH SCI CAMPUS
- **Principal Investigator:** Isaac T Schiefer
- **Activity code:** U01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $556,470
- **Award type:** 5
- **Project period:** 2022-09-15 → 2027-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10870155

## Citation

> US National Institutes of Health, RePORTER application 10870155, Cross platform analysis of drug targets and toxicity of bath salts (5U01DA054330-03). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10870155. Licensed CC0.

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