Toxicological profiling of clinically used chemotherapeutics in zebrafish

Zebrafish patient-derived xenografts (zPDX) are a powerful emerging platform for personalized oncology, offering a rapid in vivo system for high-throughput chemoprofiling. Recent studies have demonstrated a strong predictive correlation between drug response in zPDX and patient clinical outcomes. However, current protocols show significant variability in drug concentrations, which can hinder comparison across studies. We aimed to establish toxicity profiles for commonly used clinical chemotherapeutic agents in wild-type Tübingen (TU) and Casper (CSP) zebrafish (ZF) strains. Embryos aged 48-72 hours post-fertilization (hpf) were exposed to a clinically relevant chemotherapeutic panel, including standard combination regimens, for 72 hours. Toxicity was assessed using two parameters: mortality rate and any adverse effects (AAE), defined as embryos exhibiting either mortality or morphological abnormalities. Screening of single agents was similar between the two strains, but the combination regimes revealed toxicity disparities, with AAE proving to be the more sensitive endpoint. Highly toxic agents, such as paclitaxel, caused rapid, dose-dependent lethality, whereas antimetabolites like 5-fluorouracil (5-FU) showed high safety margins. Multi-agent protocols demonstrated synergistic toxicity, with more complex regimens correlating with increased adverse effects, particularly in the CSP strain. This study establishes a toxicological framework for standardizing chemotherapy dosing in ZF larvae and recommends AAE as the primary metric for defining non-toxic concentrations. Our results also underscore the necessity of testing the exact clinical drug formulation, due to potential excipient effects, and of screening multi-agent protocols for synergistic toxicity. We hope these findings will contribute to the further standardization of zPDX models for clinical applications.