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Histopathologic examination was done on 18 cases after percutaneous ethanol injection therapy (PEIT) for hepatocellular carcinoma. In eight cases, the lesion was treated by PEIT alone; in the other ten cases, PEIT was combined with transcatheter arterial embolization. The lesion was completely necrotic in 13 cases, 90% necrotic in four cases, and 70% necrotic in the rest. In addition, PEIT seemed to be effective against intercapsular, extracapsular, and vascular invasions. In the four cases of incomplete necrosis, the viable cancer tissue remained in small tumor nodules around the main tumor, in portions isolated by septa, or along the edge of the lesion. Therefore, ethanol should be injected not only into the center of the lesion, but also into sites close to its edge. Ethanol did not damage noncancerous liver parenchyma distant from injected sites. Local dissemination of the cancer cells was not found in any case. Therefore, PEIT seems to be a valuable therapy and may be an alternative to surgery in some cases.

Crystal structures of L-2-haloacid dehalogenase from Pseudomonas sp. YL complexed with monochloroacetate, L-2-chlorobutyrate, L-2-chloro-3-methylbutyrate, or L-2-chloro-4-methylvalerate were determined at 1.83-, 2.0-, 2.2-, and 2.2-Å resolutions, respectively, using the complex crystals prepared with the S175A mutant, which are isomorphous with those of the wild-type enzyme. These structures exhibit unique structural features that correspond to those of the reaction intermediates. In each case, the nucleophile Asp-10 is esterified with the dechlorinated moiety of the substrate. The substrate moieties in all but the monochloroacetate intermediate have a D-configuration at the C 2 atom. The overall polypeptide fold of each of the intermediates is similar to that of the wild-type enzyme. However, it is clear that the Asp-10 -Ser-20 region moves to the active site in all of the intermediates, and the Tyr-91-Asp-102 and Leu-117-Arg-135 regions make conformational changes in all but the monochloroacetate intermediates. Ser-118 is located near the carboxyl group of the substrate moiety; this residue probably serves as a binding residue for the substrate carboxyl group. The hydrophobic pocket, which is primarily composed of the Tyr-12, Gln-42, Leu-45, Phe-60, Lys-151, Asn-177, and Trp-179 side chains, exists around the alkyl group of the substrate moiety. This pocket may play an important role in stabilizing the alkyl group of the substrate moiety through hydrophobic interactions, and may also play a role in determining the stereospecificity of the enzyme. Moreover, a water molecule, which is absent in the substrate-free enzyme, is present in the vicinities of the carboxyl carbon of Asp-10 and the side chains of Asp-180, Asn-177, and Ala-175 in each intermediate. This water molecule may hydrolyze the ester intermediate and its substrate. These findings crystallographically demonstrate that the enzyme reaction proceeds through the formation of an ester intermediate with the enzyme's nucleophile Asp-10.1 is a unique enzyme that catalyzes the hydrolytic dehalogenation of L-2-haloacids to produce the corresponding D-2-hydroxyacids with an inversion of the C 2 -configuration. Various L-DEXs exhibiting very similar sequences have been isolated from different bacterial sources. Of all the enzymes, L-DEX YL, isolated from a 2-chloroacrylate-utilizable bacterium, Pseudomonas sp. YL, is an unusual L-DEX, in that it is relatively thermostable even though it is derived from a mesophilic bacterium (1). It is a dimeric enzyme formed by two identical subunits of 232 amino acid residues. We have already reported the crystallization of L-DEX YL (2) and its crystal structure determined by a 2.5-Å resolution x-ray analysis (3). The enzyme has a core domain of ␣/␤-structure, which differs topologically from those of the ␣/␤ hydrolase fold family proteins (4), along with a subdomain having a four-helix-bundle structure. Our ion spray mass spectrometric study showed that the dehalogenation of the L-2-haloacid catalyzed by L-DEX YL proceeds i...