CEPHEM SULFONES AS INACTIVATORS OF HUMAN-LEUKOCYTE ELASTASE .5. 7-ALPHA-METHOXY-1,1-DIOXOCEPHEM-4-KETONE AND 7-ALPHA-CHLORO-1,1-DIOXOCEPHEM-4-KETONE

Studies on cephem sulfones as inhibitors of human leukocyte elastase (HLE) have been extended to the new class of cephem 4-ketones. tert-Butyl and phenyl ketones were prepared from 4-carboxycephem derivatives, at either the sulfide or sulfone oxidation level, by chemoselective Grignard reaction. Obtained products were functionalized with heterocyclothio and acyloxy substituents at C-3', C-2, or both positions. tert-Butyl ketones of the 7 alpha-chlorocephem series were in general at least as potent as the corresponding esters at inhibiting the enzyme, but improvements in hydrolytic stability were only marginal. On the other hand, tert-butyl ketones of the 7 alpha-methoxycephem series combined potent biochemical activity with acceptable hydrolytic stability, thus overstepping the esters, thiolesters, and amides reported previously. In particular, the tert-butyl ketones possessing a heterocyclothio group at C-3' or C-2 were at least as active as the corresponding tert-butyl esters but 1 order of magnitude more stable in physiologic buffers (pH 7.4, 37 degrees C). Introduction of acyloxy groups at C-2 delivered the most potent HLE inhibitors of the cephem class ever reported, with inhibition parameters often outside the determination limits of our standard protocol (second-order rate constant k(on) > 2 000 000 M(-1) s(-1); K-i at steady state <2 nM). Keto-enol tautomerism was found to depress activity and boost hydrolytic stability. Thus, double substitution with heterocyclic thiols produced compounds with diverging properties, according to the extent of enolate formation at the investigated pH (7.4): the weakly acidic tert-butyl ketones (pK(a) greater than or equal to 5.8) proved to be potent inhibitors (k(on) over 10(4) M-l s(-1)) with reasonable hydrolytic stability (t(1/2) = 30-75 h), while the phenyl ketones (pK(a) < 4) were fair inhibitors (k(on),, over 10(3) M(-1) s(-1); K-i at steady state approximate to 50 nM) with hydrolytic half-lives exceeding 1000 h. Selected compounds efficiently inhibited the degradation of insoluble bovine neck elastin by HLE in a concentration-dependent manner. Intracellular HLE of polymorphonuclear leukocytes was in general unaffected; however, a lipophilic cephem sulfone apparently able to inactivate the enzyme in living cells was identified.