Transcripts of the UGT1A gene, encoding half of human UDP-glucuronosyltransferase (UGT) enzymes, undergo alternative splicing, resulting in active enzymes named isoforms 1 (i1s) and novel truncated isoforms 2 (i2s). Here, we investigated the effects of depleting endogenous i2 on drug response and attempted to unveil any additional biologic role(s) for the truncated novel UGT proteins. We used an integrated systems biology approach that combines RNA interference with unbiased global genomic and proteomic screens, and used HT115 colorectal cancer cells as a model. Consistent with previous evidence suggesting that i2s negatively regulate i1s through protein-protein interactions, i2-depleted cells were less sensitive to drug-induced cell death (IC50 of 0.45 ± 0.05 μM versus 0.22 ± 0.03 μM; P = 0.006), demonstrating that modulation of i2 levels meaningfully impacts drug bioavailability and cellular response. We also observed reduced production of reactive oxygen species by 30% (P < 0.05), and an enhanced expression (>1.2-fold; P < 0.05) of several proteins, such as hemoglobin α genes and superoxide dismutase 1, that have network functions associated with antioxidant properties. Interaction proteomics analysis of endogenous proteins from the cellular model, mainly in human intestine but also in kidney tissues, further uncovered interactions between i2s (but not i1s) and the antioxidant enzymes catalase and peroxiredoxin 1, which may influence antioxidant potential through sequestration of these novel partners. Our findings demonstrate for the first time dual roles for i2s in the cellular defense system as endogenous regulators of drug response as well as in oxidative stress.