Summary The antitumour effects of lipoteichoic acids (LTA) extracted from Streptococcus pyogenes were studied in comparison with other streptococcal cellular components. LTA suppressed the tumour growth of both solid-and ascites-type MethA fibrosarcoma as did the whole cells of S. pyogenes . No OK-432, a streptococcal preparation, has been widely used to increase the resistance against tumours in experimental animals and cancer patients (Kurokawa et al., 1972;Sakurai et al., 1972). We have reported that OK-432, consisting of the whole organisms of S. pyogenes Su strain, significantly suppressed the growth of both ascites-and solid-type Meth A fibrosarcoma and Ehrlich carcinoma. The cytoplasmic membrane fraction prepared from the Su strain had an antitumour effect against the ascites variant but not against the solid tumour. In contrast, the isolated cell wall fraction acted as an antitumour agent against the solid tumour but not the ascitic variant (Koshimura et al., 1977;Yamamoto et al., 1980). These facts indicate that the antitumour principle of streptococcal whole cells lies in the cell wall and the cytoplasmic membrane. A possible candidate for such an antitumour principle is lipoteichoic acids (LTA), in view of the study by Wicken and Knox (1980) showing that LTA transects both the cell wall and the cytoplasmic membrane. There have been no previous reports on the antitumour effect of LTA.The purpose of the present study was to examine the antitumour effects of LTA in comparison with other streptococcal cellular components. Materials and methodsAnimals BALB/c (female, 6-8 weeks old) and CD-1 mice (female, 6-8 weeks old) were purchased from Charles River Japan Inc. (Atsugi, Kanagawa, Japan). water and then extracted with an equal volume of 95% phenol at room temperature for 1h. The water phase containing the LTA was separated by centrifugation at 6,000 g for 30 min, concentrated by evaporation under reduced pressure, and lyophilized. The resulting crude LTA was dissolved in 0.2 M ammonium acetate at 50 mg ml-1 and applied to a Sepharose 6B (Pharmacia, Uppsala Sweden) column (2.6 x 87 cm). The column was eluted with 0.2 M ammonium acetate to separate the LTA from polyglycerophosphate (PGP) and nucleic acids. The LTA fraction thus purified was lyophilized, and dissolved in pyrogen-free 0.85% NaCl solution before use. The purity of the LTA preparation was assessed by gas-liquid chromatography, and by chemical and immunochemical analyses as previously described (Hamada et al., 1985). Chemical analysis showed that the ratio of glycerol, fatty acids and alanine was consistent with the analytical data reported by Ofek et al. (1975). The colorimetric Limulus lysate assay with Toxicolor Test (Seikagakukogyo, Tokyo, Japan) indicated that 1 mg of the LTA preparation contained < 280 pg equivalent to a standard reference LPS (Bacto lipopolysaccharide W derived from Salmonella enteritidis). Other streptococcal components were obtained as follows: Peptidoglycan was prepared from S. pyogenes A374 according to the method d...