Xanthohumol (XN), a prenylated flavonoid found in hops, inhibits growth in a variety of cancer cell lines; however, its use raises concerns as gut microbiota and the host’s hepatic cytochrome P450 enzymes metabolize it into the most potent phytoestrogen known, 8-prenylnaringenin (8-PN). The XN derivatives dihydroxanthohumol (DXN) and tetrahydroxanthohumol (TXN) are not metabolized into 8-PN and they show higher tissue concentrations in vivo compared with XN when orally administered to mice at the same dose. Here we show that DXN and TXN possess improved anti-proliferative activity compared with XN in two colon (HCT116, HT29) and two hepatocellular (HepG2, Huh7) carcinoma cell lines, as indicated by their respective IC₅₀ values. Furthermore, XN, DXN, and TXN induce extensive apoptosis in all these carcinoma cell lines. Finally, TXN induces G₀/G₁ cell cycle arrest in the colon carcinoma cell line HT29. Our findings suggest that DXN and TXN could show promise as therapeutic agents against colorectal and liver cancer in preclinical studies without the drawback of metabolism into a phytoestrogen.

A calorie-dense diet is a well-established risk factor for obesity and metabolic syndrome (MetS), whereas the role of the intestinal microbiota (IMB) in the development of diet-induced obesity (DIO) is not completely understood. To test the hypothesis that Swiss Webster (Tac:SW) mice can develop characteristics of DIO and MetS in the absence of the IMB, we fed conventional (CV) and germ-free (GF) male Tac:SW mice either a low-fat diet (LFD; 10% fat derived calories) or a high-fat diet (HFD; 60% fat derived calories) for 10 weeks. The HFD increased feed conversion and body weight in GF mice independent of the increase associated with the microbiota in CV mice. In contrast to CV mice, GF mice did not decrease feed intake on the HFD and possessed heavier fat pads. The HFD caused hyperglycemia, hyperinsulinemia, and impaired glucose absorption in GF mice independent of the increase associated with the microbiota in CV mice. A HFD also elevated plasma LDL-cholesterol and increased hepatic triacylglycerol, free fatty acids, and ceramides in all mice, whereas hypertriglyceridemia and increased hepatic medium and long-chain acylcarnitines were only observed in CV mice. Therefore, GF male Tac:SW mice developed several detrimental effects of obesity and MetS from a high-fat, calorie dense diet.

Protein tyrosine (Y) nitration is an oxidative modification that occurs in pathological conditions such as neurodegenerative diseases and solid tumors. Depending on the location of the tyrosine residue, nitration can modify protein structure and function and affect cellular processes. We previously showed that site-specific nitration of the molecular chaperone heat shock protein 90 (Hsp90) leads to distinct pathological gain-of-function that cannot be compensated or overcome by native Hsp90. While Hsp90 nitrated on Y33 localizes in mitochondria and decreases mitochondrial metabolism, Hsp90 nitrated on Y56 activates the purinergic receptor and calcium channel P2X7, triggering downstream signaling pathways that can lead to either cell proliferation or apoptosis, depending on the cell type. Herein, using complementary biophysical, biochemical, and in silico methods, we show that nitration on Y33 and Y56 triggers significant site-dependent local and global structural changes linked to changes in Hsp90 activity. Nitration of these critical residues led to destabilization of Hsp90 dimer and formation of stable oligomeric species, with differential effects on Hsp90 ATPase and chaperone holdase activities depending on the nitrated residue. Molecular dynamics simulations further support the impact of nitration on Y33 and Y56 on the ATP-lid dynamics and the interaction of ATP with R392, critical to Hsp90 ATPase activity. Establishing the molecular basis of nitration-induced structural changes in Hsp90 leading to disease-driving functions is the first step toward the development of therapeutic approaches selectively targeting these pathological variants of Hsp90.