Design and synthesis of the stabilized analogs of belactosin A with the unnatural cis-cyclopropane structure

The belactosin A analog 2a, having the unnatural cis-cyclopropane structure instead of the trans-cyclopropane structure in belactosin A, is a much more potent proteasome inhibitor than belactosin A. However, its cell growth inhibitory effect is rather lower than that expected from its remarkable proteasome inhibitory effect, probably due to its instability under cellular conditions. We hypothesized that the instability of 2a was due to chemical and enzymatic hydrolysis of the strained beta-lactone moiety. Thus, to increase the stability of 2a by chemical modification, its analogs with a sterically more hindered beta-lactone moiety and/or cyclopropylic strain-based conformational restriction were designed and synthesized, resulting in the identification of a stabilized analog 6a as a proteasome inhibitor with cell growth inhibitory effects. Our findings suggest that the chemical and biological stability of 2a is significantly affected by the steric hindrance around its beta-lactone carbonyl moiety and the conformational flexibility of the molecule

Rational hopping of a peptidic scaffold into non-peptidic scaffolds: structurally novel potent proteasome inhibitors derived from a natural product, belactosin A

Rational scaffold hopping of a natural product belactosin A derivative was successfully achieved based on the pharmacophore model constructed. The peptidic scaffold was replaced by significantly simplified non- peptidic scaffolds, bywhich weak belactosin A (IC50= 1440nM) was converted into highly potent non- peptidic inhibitors (IC50 = 26-393 nM)

Effect of CST at sub-MICs on the expression levels of efflux pumps and biofilm-related genes in <i>A</i>. <i>baumannii</i>.

<p>Summarized results showing the mRNA levels of efflux pumps and biofilm-related genes in strains ATCC 19606, R2 and R3 cultured in LB-broth with CST at sub-MICs. The mRNA levels of (A) <i>adeB</i>, (B) <i>adeG</i>, (C) <i>adeJ</i>, (D) <i>ompA</i>, (E) <i>bap</i>, (F) <i>pgaA</i> and (G) <i>abaI</i> were analyzed by real-time PCR. Bar graph data are shown as the means ± SEM (n = 6) of 3 independent experiments. Asterisks indicate statistically significant differences (**<i>P</i><0.01; *<i>P<</i>0.05, non-treated bacteria <i>vs</i>. antibiotics-treated bacteria; One-way ANOVA).</p

Primers used for real-time PCR.

<p>Primers used for real-time PCR.</p

MICs (μg/mL) of antibiotics against <i>A</i>. <i>baumannii</i> ATCC 19606 and the clinical isolates of MDRA.

<p>MICs (μg/mL) of antibiotics against <i>A</i>. <i>baumannii</i> ATCC 19606 and the clinical isolates of MDRA.</p

Effect of PMB at sub-MICs on the expression levels of efflux pumps and biofilm-related genes in <i>A</i>. <i>baumannii</i>.

<p>Summarized results showing the mRNA levels of efflux pumps and biofilm-related genes in strains ATCC 19606, R2 and R3 cultured in LB-broth with sub-MICs of PMB. The mRNA levels of (A) <i>adeB</i>, (B) <i>adeG</i>, (C) <i>adeJ</i>, (D) <i>ompA</i>, (E) <i>bap</i>, (F) <i>pgaA</i> and (G) <i>abaI</i> were analyzed by real-time PCR. Bar graph data are shown as the means ± SEM (n = 6) of 3 independent experiments. Asterisks indicate statistically significant differences (**<i>P</i>< 0.01; *<i>P<</i>0.05, non-treated bacteria <i>vs</i>. antibiotics-treated bacteria; One-way ANOVA).</p

Relationship between biofilm formation and the expression of efflux pumps and biofilm-related genes in <i>A</i>. <i>baumannii</i> in the presence of PMB at its sub-MICs.

<p>Relationship between biofilm formation and the expression of efflux pumps and biofilm-related genes in <i>A</i>. <i>baumannii</i> in the presence of PMB at its sub-MICs.</p