Fig. 4. APA shows anti-tumor activity in vivo and is a proof-of-concept compound for colorectal cancer chemotherapy.

a, b Treatment with liposomal APA selectively inhibits growth of slow NAT2 tumors in vivo. Athymic nude mice carrying subcutaneous slow (left flank) and rapid (right flank) NAT2 RKO cell clones received intravenous injections of liposomal APA on days 0 and 3. Results are shown for one representative experiment conducted in 9 mice for the treatment group and 10 mice for the control group. Data were analyzed using one-way ANOVA, n.s., P = 0.9013 and *P = 0.0220. c Primary CRC tumors show sensitivity toward APA treatment. The survival index (SI) of primary tumor cells and organoids from 12 different CRC patients was determined when exposed to increasing concentrations of APA (log of concentration in μM). Samples encoding rapid (blue), intermediate (orange), and slow (gray) acetylator phenotypes are shown with their respective NAT2 haplotype. d A conceptual way of exploiting NAT2 loss of heterozygosity. Anti-cancer drugs benefit from reduced toxicity and improved tumor uptake when encapsulated in stealth liposomes. First, liposomal APA is injected intravenously in a rapid/slow NAT2 patient whose tumor has lost the rapid allele¹. The stealth liposomes extravasate through the leaky tumor vessels and accumulate in tumor tissues due to the enhanced permeability and retention (EPR) effect². The lipid bilayer of the liposomes fuses with cell membranes and deliver APA to the colorectal tumor cells and adjacent normal epithelial cells. The adjacent normal epithelial cells express rapid NAT2 and are able to process APA into its nontoxic product NAPA. In contrast, cancer cells expressing only slow NAT2 are not capable of detoxifying APA and die. Liposomal drug formulations also accumulate in the liver, where APA is cleared by rapid NAT2³. Every element of (d) was created in Adobe Illustrator 2019.