Oxidative Stress and Cocaine Intoxication as Start Points in the Pathology of Cocaine-Induced Cardiotoxicity

Psychomotor stimulants are the most commonly used prohibited substances after cannabis. Globally, their use reaches epidemiological proportions and is one of the most common causes of death in many countries. The use of illicit drugs has negative effects on the cardiovascular system and is one of the causes of serious cardiovascular pathologies, ranging from abnormal heart rhythms to heart attacks and sudden cardiac death. The reactive oxygen species generation, toxic metabolites formation, and oxidative stress play a significant role in cocaine-induced cardiotoxicity. The aim of the present review is to assess acute and chronic cocaine toxicity by focusing on the published literature regarding oxidative stress levels. Hypothetically, this study can serve as a basis for developing a rapid and effective method for determining oxidative stress levels by monitoring changes in the redox status of patients with cocaine intoxication.

Evaluation of Cocaine Effect on Endogenous Metabolites of HepG2 Cells Using Targeted Metabolomics

Cocaine toxicity has been a subject of study because cocaine is one of the most common and potent drugs of abuse. In the current study the effect of cocaine on human liver cancer cell line (HepG2) was assessed. Cocaine toxicity (IC50) on HepG2 cells was experimentally calculated using an XTT assay at 2.428 mM. The metabolic profile of HepG2 cells was further evaluated to investigate the cytotoxic activity of cocaine at 2 mM at three different time points. Cell medium and intracellular material samples were analyzed with a validated HILIC-MS/MS method for targeted metabolomics on an ACQUITY Amide column in gradient mode with detection on a triple quadrupole mass spectrometer in multiple reaction monitoring. About 106 hydrophilic metabolites from different metabolic pathways were monitored. Multivariate analysis clearly separated the studied groups (cocaine-treated and control samples) and revealed potential biomarkers in the extracellular and intracellular samples. A predominant effect of cocaine administration on alanine, aspartate, and glutamate metabolic pathway was observed. Moreover, taurine and hypotaurine metabolism were found to be affected in cocaine-treated cells. Targeted metabolomics managed to reveal metabolic changes upon cocaine administration, however deciphering the exact cocaine cytotoxic mechanism is still challenging.

Bioprinting transgenic plant cells for production of a recombinant biodefense agent

Transgenic rice cells (Oryza sativa) producing recombinant butyrylcholinesterase (BChE) as a prophylactic/therapeutic against organophosphate nerve agent poisoning, cocaine toxicity, and neurodegenerative diseases like Alzheimer’s were immobilized in a polyethylene glycol-based hydrogel. The cells were sustained for 14 days in the semi-solid matrix, undergoing a growth phase from days 0-6, a BChE production phase in sugar-free medium from days 6-12, and a growth/recovery phase from days 12-14. Throughout this period, the cells maintained similar viability to those in suspension cultures and displayed analogous sugar consumption trends. The rice cells in the bioprintable hydrogel also produced a significant amount of active BChE, comparable to the levels produced in liquid cultures. A considerable fraction of this BChE was secreted into the media, allowing for easier product separation. Overall, we demonstrate a simple, efficient, robust, modular, and potentially field-deployable bioreactor system for the manufacture of biologics.

4141 Molecular Signatures of Cocaine Toxicity in Postmortem Human Brain and Neurons

OBJECTIVES/GOALS: The goal of this project is to identify new therapeutic targets and biomarkers to treat or prevent cocaine toxicity by investigating proteomic, transcriptomic and epigenetic signatures of cocaine exposure in human subjects. METHODS/STUDY POPULATION: Cocaine is a highly addictive neurotoxic substance, and it is estimated that 1.9 million Americans are current users of cocaine. To study the molecular effects of cocaine, we generated preliminary proteomics and next-generation RNA sequencing (RNAseq) data from human postmortem dorsolateral prefrontal cortex (Broadmann area 9 or BA9) of 12 cocaine-exposed subjects and 17 controls. Future directions for this project include RNAseq and DNA methylation analysis of neuronal nuclei sorted from human postmortem BA9 and a human induced pluripotent stem cell-derived neuron (hiPSN) model of cocaine exposure from the same postmortem subjects from whom we have brain samples. RESULTS/ANTICIPATED RESULTS: We found alterations in neuronal synaptic protein levels and gene expression, including the serotonin transporter SLC6A4, and synaptic proteins SNAP25, SYN2, SYNGR3. Pathway analysis of our results revealed alterations in specific pathways involved with neuronal function including voltage-gated calcium channels, and GABA receptor signaling. In the future, we expect to see an enhancement in neuron-specific gene expression signatures in our sorted neuronal nuclei and our hiPSN model of cocaine exposure. The hiPSN model will help elucidate which effects are due to acute versus chronic exposure of cocaine. DISCUSSION/SIGNIFICANCE OF IMPACT: Neuronal signatures found with this analysis can help us understand mechanisms of cognitive decline in long-term cocaine users as well as the acute effects on the brain of cocaine taken in overdose. With this work and future proposed studies, we can discover novel clinical biomarkers for cocaine neurotoxicity in patients with cocaine use disorder and determine readouts for future therapeutic development on cocaine addiction and overdose.

Preventative effects of aripiprazole and quetiapine on seizure and lethality in a mice cocaine toxicity model: an experimental study

Objective To assess the effectiveness of pre-treatment with aripiprazole and quetiapine to prevent acute cocaine toxicity in a mouse model of cocaine toxicity. Methods This experimental study included three groups ( n = 25 per group) of mice that were intraperitoneally injected with normal saline solution, 10 mg/kg quetiapine or 10 mg/kg aripiprazole 15 min before 105 mg/kg cocaine hydrochloride. When the cocaine administration was completed, researchers blinded to the study groups observed the mice in terms of seizures and death for a further 30 min. Results In the cocaine + quetiapine group, the mean ± SE time to the first seizure was 10.80 ± 2.27 min and seizure activity was detected in 18 mice (72%) by the end of the 30 min. In the cocaine + aripiprazole group, the mean ± SE time to the first seizure was 18.10 ± 1.94 min and seizure activity was detected in 15 mice (60%) by the end of the 30 min. When compared with the control group, there was a significant difference between the cocaine + quetiapine and cocaine + aripiprazole groups in terms of seizure activity. Survival time was increased in the cocaine + aripiprazole group compared with the control and cocaine + quetiapine groups. Conclusion Quetiapine and aripiprazole pre-treatment reduced seizure activity and delayed the onset of seizures compared with the control group.