title: Inhibition of the class C beta-lactamase from Acinetobacter spp.: insights into effective inhibitor design.
abstract: The need to develop beta-lactamase inhibitors against class C cephalosporinases of Gram-negative pathogens represents an urgent clinical priority. To respond to this challenge, five boronic acid derivatives, including a new cefoperazone analogue, were synthesized and tested against the class C cephalosporinase of Acinetobacter baumannii [Acinetobacter-derived cephalosporinase (ADC)]. The commercially available carbapenem antibiotics were also assayed. In the boronic acid series, a chiral cephalothin analogue with a meta-carboxyphenyl moiety corresponding to the C(3)/C(4) carboxylate of beta-lactams showed the lowest K(i) (11 +/- 1 nM). In antimicrobial susceptibility tests, this cephalothin analogue lowered the ceftazidime and cefotaxime minimum inhibitory concentrations (MICs) of Escherichia coli DH10B cells carrying bla(ADC) from 16 to 4 microg/mL and from 8 to 1 microg/mL, respectively. On the other hand, each carbapenem exhibited a K(i) of <20 microM, and timed electrospray ionization mass spectrometry (ESI-MS) demonstrated the formation of adducts corresponding to acyl-enzyme intermediates with both intact carbapenem and carbapenem lacking the C(6) hydroxyethyl group. To improve our understanding of the interactions between the beta-lactamase and the inhibitors, we constructed models of ADC as an acyl-enzyme intermediate with (i) the meta-carboxyphenyl cephalothin analogue and (ii) the carbapenems, imipenem and meropenem. Our first model suggests that this chiral cephalothin analogue adopts a novel conformation in the beta-lactamase active site. Further, the addition of the substituent mimicking the cephalosporin dihydrothiazine ring may significantly improve affinity for the ADC beta-lactamase. In contrast, the ADC-carbapenem models offer a novel role for the R(2) side group and also suggest that elimination of the C(6) hydroxyethyl group by retroaldolic reaction leads to a significant conformational change in the acyl-enzyme intermediate. Lessons from the diverse mechanisms and structures of the boronic acid derivatives and carbapenems provide insights for the development of new beta-lactamase inhibitors against these critical drug resistance targets.
authors.affiliation: Department of Pathology, Case Western Reserve UniversitySchool of Medicine, Cleveland, Ohio 44106, USA.
authors.forename: Sarah M
authors.initials: SM
authors.lastname: Drawz
authors.forename: Maja
authors.initials: M
authors.lastname: Babic
authors.forename: Christopher R
authors.initials: CR
authors.lastname: Bethel
authors.forename: Magda
authors.initials: M
authors.lastname: Taracila
authors.forename: Anne M
authors.initials: AM
authors.lastname: Distler
authors.forename: Claudia
authors.initials: C
authors.lastname: Ori
authors.forename: Emilia
authors.initials: E
authors.lastname: Caselli
authors.forename: Fabio
authors.initials: F
authors.lastname: Prati
authors.forename: Robert A
authors.initials: RA
authors.lastname: Bonomo
mesh_terms: D000150:Acinetobacter / Q000201:enzymology*; D015780:Carbapenems / Q000737:chemistry / Q000494:pharmacology; D002510:Cephalosporinase / Q000737:chemistry; D002512:Cephalothin / Q000737:chemistry / Q000494:pharmacology; D015195:Drug Design; D004926:Escherichia coli / Q000187:drug effects; D006090:Gram-Negative Bacteria / Q000201:enzymology*; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D010405:Penicillinase / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D021241:Spectrometry, Mass, Electrospray Ionization; D013329:Structure-Activity Relationship; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000737:chemistry / Q000378:metabolism
chemical_list: D015780:Carbapenems; D065093:beta-Lactamase Inhibitors; D002510:Cephalosporinase; D010405:Penicillinase; D001618:beta-Lactamases; D002512:Cephalothin