Vacuolar ATPases Inhibitors

Roush et al. applied the diastereoselective crotylboration methodology in the total synthesis of bafilomycin Ai (66), a potent vacuolar ATPase inhibitor that displays broad antibiotic activity27 (Scheme 3.lx). In the synthesis, the known aldehyde (R)-67 was treated with ( )-crotylboronate (R,R)-43E to provide an 85 15 mixture of the homoallylic alcohol 68 and the undesired 3,4-anti-4,5-syn diastereomer with an isolated 78% yield of 68. Alcohol protection as a TBS ether followed by hydroboration mediated by Wilkinson s catalyst efficiently provided the primary alcohol 69. 

This tandem procedure is useful for synthesis of fhe highly functionahzed vacuolar ATPase inhibitor, bafilomycin Vi (Scheme 12.107) [209]. Couphng was effected wifh Pd(O), AsPh3, and LiCI in NMP and proceeded to completion in 5 h at room temperature. Alfhough fhe indole nucleus occurs in a wide range of natural products, consecutive cychzation of 2-isocyanocinnamate by hydrostannation, de-... 

The ease of hydrolysis can vary enormously in some cases hydrolysis is so easy that inadvertent loss of the protecting group during chromatography becomes a nuisance (such as acetals of a, f3-unsaturated ketones). On the other hand acetals may be so robust that forcing conditions (mineral acid and heat) are required. For example, substrates that contain a basic amino function — even if it is remote — can be quite resistant to hydrolysis, because protonation first takes place at the more basic amino function,18 The resultant positive charge repels the second Q-protonation required to set in motion the hydrolysis. An acid-catalysed hydrolysis of a basic acetal that required refluxing with 6 M HC1 in acetone for 6-10 h19 is illustrated in Scheme 2,3, Perhaps the mildest conditions employ pyridinium p-toluenesulfonate (PPTS, pKa 5.2) in methanol or aqueous acetone exemplified by the transacetalisation reaction taken from a synthesis of the vacuolar H+-ATPase inhibitor Bafilomycin Ai [Scheme 2.4, 20... 

The enzyme proved to be composed of four types of subunits (87, 60, 29 and 20 kDa). The subunit ratio was assumed to be equal to 3 3 1 1 [66]. The sequences of major subunits showed significant (more than 50%) homology with the detachable sector of eukaryotic vacuolar (V-type) H -ATPase. On the other hand, homology with bacterial Fi ATPase proved to be less than 30%. Since the above-mentioned inhibitor pattern is, in fact, identical with those of the vacuolar ATPase, one might assume that halobacterial ATPase corresponds to the V-type. However, the sequence of proteolipid (c subunit) of... 

Three types of ATP-driven cation pumps can be distinguished on the basis of their structure and their sensitivity to inhibitors. They are the E E2-ATPases, the FiFo-proton-translocating ATP synthase, and the vacuolar ATPases which are designated as P-, F-, and V-type ATPases, respectively [37]. Several of the distinguishing characteristics of these enzymes are summarized in Table 1. The F- and V-ATPases can be differentiated by the sensitivity of the former to azide and the latter to nitrate and A-ethylmaleimide (NEM)[38]. In addition, the V-ATPases are exquisitely sensitive to the antibiotic bafilomycin A [39,40]. 

Beutler JA, McKee TC. Novel marine and microbial natural product inhibitors of vacuolar ATPase. Curr. Med. Chem. 2003 10(9) 787-796. 

One closely related family to resorcinylic macroHdes are the saUcylate enamide macrolides, which are secondary marine metabolites with a 12-membered ring benzolactone and a dienyl enamide side chain. SaHcyhhalamide A (34) is a potent inhibitor of the mammalian vacuolar ATPase and exerts great diSerential cytotoxicity in the National Cancer Institute (NCI) 60 ceU lines [23]. Other members... 

In addition to direct inhibition of the vesicular transport protein, storage of neurotransmitters can be reduced by dissipation of the proton electrochemical gradient. Bafilomycin (a specific inhibitor of the vacuolar H+-ATPase), as well as the proton ionophores carbonyl cyanide m-chlorophenylhydrazone (CCCP) and carbonylcyanide p-(trifluoromethoxy) phenylhy-drazone (FCCP) are used experimentally to reduce the vesicular storage of neurotransmitters. Weak bases including amphetamines and ammonium chloride are used to selectively reduce ApH. 

The macrolide antibiotic bafilomycin Al and concanamycin A are specific inhibitors of vacuolar-type H(- -)-ATPase, which prevents the acidification of endosomes and lysosomes and increases the intralysosomal pH from about 5.1-5.5 to about 6.3 (61,84-87). Bafilomycin Al is applied in concentrations ranging from 25-1000 nM and is incubated for 30 to 60 minutes in the presence or absence of serum. We observed maximal effects in COS-7 and HUVEC with concentrations of 100-200 nM and an incubation time of 60 minutes. 

Yoshimori T, Yamamoto A, Moriyama Y, Futai M, Tashiro Y. Bafilomycin Al, a specific inhibitor of vacuolar-type H(+)-ATPase, inhibits acidification and protein degradation in lysosomes of cultured cells. J Biol Chem 1991 266(26) 17707-17712. 

Adociasulfates 1-6 (380-385) were isolated from a Haliclona (aka Adocia) sp. from Palau and were all inhibitors of kinesin motor proteins [331]. Adociasulfate 2 (381) had earlier been shown to inhibit the activity of the motor protein kinesin by interference with its binding to microtubules [332], An Adocia sp. from the Great Barrier Reef contained adociasulfates 1 (380), 7 (386) and 8 (387), which inhibit vacuolar H -ATPase [333]. Adociasulfates 5 (384) and 9 (388) were obtained from Adocia aculeata from the Great Barrier Reef [334], The structure of adociasulfate 1 (380) was confirmed by an enantioselective total synthesis [335]. Adociasulfate 10 (389) from Haliclona sp. from Palau also inhibits the kinesin motor proteins [336]. 

The effect of CCCP on NMR-detected PolyP was investigated in S. cerevisiae grown on lactate (Beauvoit el al., 1991). The cells were incubated in a resting medium in aerobiosis with lactate or glucose or in anaerobiosis with glucose. For each case, in vivo 31P NMR spectroscopy was used to measure the levels of phosphorylated compounds. A spontaneous PolyP breakdown occurred in anaerobiosis and in the absence of CCCP. In aerobiosis, PolyP hydrolysis was induced by the addition of either CCCP or a vacuolar membrane ATPase-specific inhibitor, bafilomycin Al (Beauvoit el al., 1991). 

Sato, N. Suzuki, N. Yamaguchi, M. Yamaguchi, N. Okuma, K. Proteolipid subunits of vacuolar H" -ATPase (ATP6F) as tumor antigens, application to cancer therapy, and use of proton pump inhibitor as anticancer agent. Jpn. Kokai Tokkyo Koho JP 2001286284,2001 Chem. Abstr. 2001,135, 314438. 

Farina C, Gagliardi S. Selective inhibitors of the osteoclast vacuolar proton ATPase as novel bone antiresorptive agents. Drug Discov. Today 1999 4(4) 163-172. 

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