The ratio of nicotinic acid to nicotinamide as a microbial biomarker for assessing cell therapy product sterility.

Controlling microbial risks in cell therapy products (CTPs) is important for product safety. Here, we identified the nicotinic acid (NA) to nicotinamide (NAM) ratio as a biomarker that detects a broad spectrum of microbial contaminants in cell cultures. We separately added six different bacterial species into mesenchymal stromal cell and T cell culture and found that NA was uniquely present in these bacteria-contaminated CTPs due to the conversion from NAM by microbial nicotinamidases, which mammals lack. In cells inoculated with 1 × 10 CFUs/mL of different microorganisms, including USP <71> defined organisms, the increase in NA to NAM ratio ranged from 72 to 15,000 times higher than the uncontaminated controls after 24 h. Importantly, only live microorganisms caused increases in this ratio. In cells inoculated with 18 CFUs/mL of , 20 CFUs/mL of , and 10 CFUs/mL of , significant increase of NA to NAM ratio was detected using LC-MS after 18.5, 12.5, and 24.5 h, respectively. In contrast, compendial sterility test required >24 h to detect the same amount of these three organisms. In conclusion, the NA to NAM ratio is a useful biomarker for detection of early-stage microbial contaminations in CTPs.