PR-104A sensitivity correlated with AKR1C3 mRNA, protein, and enzymatic activity. (A) A total of 17 ALL xenografts (T-ALL, n = 9; BCP-ALL, n = 8) were profiled on Illumina Human Ref 12 Beadchip arrays, and the top 25 differentially expressed genes between PR-104A–resistant and –sensitive xenografts; genes were ordered based on the P value. Red shows relative upregulation and blue shows relative downregulation. T-ALLs are shown in red and BCP-ALLs in blue. (B) mRNA expression of AKR1C3 by RT-qPCR in the panel of xenografts, and correlated with in vitro IC₅₀ and in vivo LGD. (C) Protein lysates were extracted from T-ALL and BCP-ALL xenografts and AKR1C3 was probed by western blot, quantified, and correlated with IC₅₀ and LGD. A representative immunoblot is shown; for all immunoblots, see supplemental Figure 3. (D) Through a SN34037-sensitive coumberone reduction, AKR1C3 enzymatic activity can be measured by the fluorescent product coumberol. Coumberol formation was measured in vitro in T-ALL and BCP-ALL xenografts, and correlated with in vitro IC₅₀ and in vivo LGD.