Studies in humans on the mechanism of potent spermicidal and apoptosis-inducing activities of vanadocene complexes

We previously demonstrated that bis-cyclopentadienyl (Cp) complexes of vana dium(IV) (vanadocenes) are potent spermicidal and apoptosis-inducing agents . To gain further insight into the structure-function relationships control ling these two properties of vanadocenes, we have synthesized analogues in which the bis-Cp rings were substituted with one or five electron-donating methyl groups. The three complexes included vanadocene dichloride (VDC), bi s(methylcyclopentadienyl) vanadium dichloride (VMDC), and bis(pentamethylcy clopentadienyl) vanadium dichloride (VPMDC). The concentration-dependent ef fect of these vanadocenes on sperm-immobilizing activity (SIA), mitochondri al membrane potential (Delta Psi m), axonemal dynein ATPase activity, and t yrosine phosphorylation of global and axoneme-specific sperm proteins was a ssessed by computer-assisted sperm analysis, flow cytometry, colorimetry, a nd immunoblotting, respectively. Apoptosis-inducing ability was quantitated by the two-color flow cytometric terminal dideoxynucleotidyl transferase-b ased assay that labels 3'-hydroxyl ends of fragmented DNA. All three vanado cenes induced rapid sperm immobilization (T-1/2 < 15 sec). Substitution of the bis-Cp rings by five methyl groups augmented the SIA of VDC by 10-fold. The EC50 values (50% inhibitory concentration) for VDC, VMDC, and VPMDC we re 7.5 mu M, 4.3 mu M, and 0.7 mu M, respectively. Whereas SIA of vanadocen es was apparent at low micromolar concentrations, the apoptosis-inducing pr operty was evident only at higher micromolar concentrations. The concentrat ions of VDC, VMDC, and VPMDC required for 50% apoptosis were 49 mu M, 67 mu M, and 153 mu M, and for 50% reduction in sperm Delta Psi m were 435 mu M, 173 mu M, and 124 mu M, respectively. Spermicidal activity of vanadocenes was not dependent on the inhibition of ATPase or tyrosine phosphorylation o f global and sperm axonemal proteins. Due to the ability of these vanadocen e complexes to rapidly generate hydroxyl radicals in the presence of oxidan t, our findings provide unprecedented evidence for a novel mechanism of act ion for spermicidal vanadocenes. The differential concentration-dependent s permicidal and apoptosis-inducing properties of vanadocenes gives them part icular utility as a new class of vaginal contraceptives.