Molecular Insights into Microbialβ-Glucuronidase Inhibition to Abrogate CPT-11 Toxicity

Abstract

Bacterial b-glucuronidases expressed by the symbiotic intestinal microbiota appear to play important roles in druginduced epithelial cell toxicity in the gastrointestinal (GI) tract. For the anticancer drug CPT-11 (irinotecan) and the nonsteroidal anti-inflammatory drug diclofenac, it has been shown that removal of the glucuronide moieties from drug metabolites by bacterial b-glucuronidases in the GI lumen can significantly damage the intestinal epithelium. Furthermore, selective disruption of bacterial b-glucuronidases by small molecule inhibitors alleviates these side effects, which, for CPT-11 {7-ethyl-10-[4-(1-piperidino)-1-piperidino]}, can be dose limiting. Here we characterize novel microbial b-glucuronidase inhibitors that inhibit Escherichia coli b-glucuronidase in vitro with K i values between 180 nM and 2 mM, and disrupt the enzyme in E. coli cells, with EC 50 values as low as 300 nM. All compounds are selective for E. coli b-glucuronidase without inhibiting purified mammalian b-glucuronidase, and they do not impact the survival of either bacterial or mammalian cells. The 2.8 Å resolution crystal structure of one inhibitor bound to E. coli b-glucuronidase demonstrates that it contacts and orders only a portion of the "bacterial loop" present in microbial, but not mammalian, b-glucuronidases. The most potent compound examined in this group was found to protect mice against CPT-11-induced diarrhea. Taken together, these data advance our understanding of the chemical and structural basis of selective microbial b-glucuronidase inhibition, which may improve human drug efficacy and toxicity.

Gut Microbiota-Mediated Drug Interactions between Lovastatin and Antibiotics

“…As the impact of gut microbiota on drug metabolism has gradually been gaining recognition, the potential DDIs occurring via the interactions with microbiota metabolism studied and the impact of gut microbes on DDIs has been discussed (Lindenbaum et al, 1981;Saha et al, 1983;Wilson and Nicholson, 2009). Furthermore, the role and significance of microbial b-glucuronidase in connection with enterohepatic recycling of xenobiotics have been emphasized (Roberts et al, 2013). The impact of gut microbiota on drug metabolism was evidenced by experimental studies involving animal models, in particular, with germ-free animals and animals in which human microflora had been introduced (Bowey et al, 2003).…”

Investigation of Host–Gut Microbiota Modulation of Therapeutic Outcome

“…Indeed, it has been discovered that the hydrolysis of SN-38G to SN-38 by microbial b-glucuronidase is responsible for the undesirable side effect of severe diarrhea (Takasuna et al, 1996;Mathijssen et al, 2001). The manipulation of the microbial b-glucuronidase activity with b-glucuronidase inhibitors and antibiotics has been shown to prevent the production of toxic SN-38 metabolites and protect the intestines of mice against injury (Takasuna et al, 1996;Wallace et al, 2010;Roberts et al, 2013). This underscores the therapeutic potential of targeted manipulation of gut microbes in minimizing the toxicity associated with irinotecan.…”

“…Such mechanistic understanding will aid the systematic characterization of drugs that are susceptible to the metabolic influence of gut microbiota and will provide insights for their therapeutic management (Jia et al, 2008;Wallace et al, 2010;Turnbaugh, 2012, 2013;Holmes et al, 2012;LoGuidice et al, 2012;Nicholson et al, 2012;Maurice et al, 2013;Roberts et al, 2013).…”

“…Coadministration of sorivudine and 5-fluorouracil resulted in drug interactions that led to death Deconjugation of drugs excreted in bile as inactive conjugates Digitoxin (Volp and Lage, 1978) b-glucuronidase Hydrolysis of glucuronide Indomethacin (Saitta et al, 2014) Hydrolysis of glucuronide of indomethacin release the aglycone which leads to gastrointestinal toxicity Morphine (Walsh and Levine, 1975) Hydrolysis of glucuronide Irinotecan (Roberts et al, 2013) Hydrolysis of SN-38 glucuronide of irinotecan (prodrug) release SN-38 in the intestines, which leads to gastrointestinal toxicity Removal of succinate group Succinylsulfathiazole (Sousa et al, 2008) Not reported Activation of prodrug to sulfathiazole Dehydroxylation L-Dopa (Goldin et al, 1973) Not reported Alteration of L-dopa pharmacokinetics by gut microbiota metabolism to form m-tyramine and m-hydroxyphenylacetic acid Acetylation 5-Aminosalicylic acid (Dull et al, 1987;Deloménie et al, 2001) N-acetyltransferase Acetylation to form acetylated 5-aminosalicylic acid Deacetylation Phenacetin (Smith and Griffiths, 1974) Not reported Formation of p-phenetidin from deacetylation reaction is correlated with toxicities such as methemoglobinemia and nephritis Cleavage of N-oxide bond Ranitidine (Basit and Lacey, 2001) Not reported Susceptible to N-oxide bond cleavage by gut bacteria Nizatidine (Basit et al, 2002) Not reported Susceptible to N-oxide bond cleavage by gut bacteria Proteolysis Insulin (Tozaki et al, 1997) Not reported Susceptible to proteolysis Calcitonin (Tozaki et al, 1997) Not reported Susceptible to proteolysis Denitration Glyceryl trinitrate (Abushamat, 1993;Sousa et al, 2008) Not reported Generate glyceryl-1,3-dinitrate, glyceryl-1,2-dinitrate, glyceryl-1-mononitrate, and glyceryl-2-mononitrate Isosorbide dinitrate (Sousa et al, 2008) Not reported Generate isomeric mononitrates and isosorbide Amine formation and hydrolysis of amide linkage…”

Inhibitory Effect of Heat-Treated 3-(3^|^prime;,4^|^prime;-dihydroxyphenyl)-L-alanine (DOPA) on ^|^beta;-glucuronidase Activity

“…Recently, Roberts et al (2013) reported that β-glucuronidase-inhibitor ASN0311025 contacts Tyr-472 within the active site region of the enzyme. The o-quinones could interact with the hydroxyl group of the tyrosine residue of protein (Coultate, 2009).…”