Maytansinoids are potent antitumor agents found in plants and microorganisms. To elucidate their biosynthesis at the biochemical and genetic level and to set the stage for their structure modification through genetic engineering, we have cloned two gene clusters required for the biosynthesis of the maytansinoid, ansamitocin, from a cosmid library of Actinosynnema pretiosum ssp. auranticum ATCC 31565. This is a rare case in which the genes involved in the formation of a secondary metabolite are dispersed in separate regions in an Actinomycete. A set of genes, asm22-24, asm43-45, and asm47, was identified for the biosynthesis of the starter unit, 3-amino-5-hydroxybenzoic acid (AHBA). Remarkably, there are two AHBA synthase gene homologues, which may have different functions in AHBA formation. Four type I polyketide synthase genes, asmA-D, followed by the downloading asm9, together encode eight homologous sets of enzyme activities (modules), each catalyzing a specific round of chain initiation, elongation, or termination steps, which assemble the ansamitocin polyketide backbone. Another set of genes, asm13-17, encodes the formation of an unusual ''methoxymalonate'' polyketide chain extension unit that, notably, seems to be synthesized on a dedicated acyl carrier protein rather than as a CoA thioester. Additional ORFs are involved in postsynthetic modifications of the initial polyketide synthase product, which include methylations, an epoxidation, an aromatic chlorination, and the introduction of acyl and carbamoyl groups. Tentative functions of several asm genes were confirmed by inactivation and heterologous expression.

Rifamycin B biosynthesis by Amycolatopsis mediterranei S699 involves a number of unusual modification reactions in the formation of the unique polyketide backbone and decoration of the molecule. A number of genes believed to be involved in the tailoring of rifamycin B were investigated and the results confirmed that the formation of the naphthalene ring moiety of rifamycin takes place during the polyketide chain extension and is catalysed by Rif-Orf19, a 3-(3-hydroxyphenyl)propionate hydroxylase-like protein. The cytochrome P450-dependent monooxygenase encoded by rif-orf5 is required for the conversion of the D12, 29 olefinic bond in the polyketide backbone of rifamycin W into the ketal moiety of rifamycin B. Furthermore, Rif-Orf3 may be involved in the regulation of rifamycin B production, as its knock-out mutant produced about 40 % more rifamycin B than the wild-type. The work also revealed that many of the genes located in the cluster are not involved in rifamycin biosynthesis, but might be evolutionary remnants carried over from an ancestral lineage. INTRODUCTIONRifamycin continues to play a significant role in clinical medicine. Synthetically modified derivatives, such as rifampicin, rifabutin and rifapentine, remain the principal chemotherapeutic agents used for combating tuberculosis, leprosy and AIDS-related mycobacterial infections (Maggi et al., 1966;Ramos-e-Silva & Rebello, 2001;Sepkowitz et al., 1995). The potent antibacterial activity of this class of antibiotics is due to their specific inhibition of bacterial DNA-dependent RNA polymerases (Campbell et al., 2001;Wehrli & Staehelin, 1969). At the same time, they have relatively low if any activity against eukaryotic RNA polymerases. Due to their high selectivity for their molecular target, the rifamycins have become a safe and effective medication. Unfortunately, as with many other antibiotics, the incidence of resistance of Mycobacterium tuberculosis, the causative agent of tuberculosis, to rifamycins is continuing to increase over time, due largely to mutational alterations of the target molecule, the b subunit of RNA polymerase (Kirschbaum & Gotte, 1993;Suzuki et al., 1995). This high-level resistance has contributed to the recent reemergence of tuberculosis as a major health problem and the consequent increase in the death toll among world populations (Dye et al., 2002). Therefore, new drug discovery and continued development of the existing drugs to combat tuberculosis is indispensable.Despite the preparation of a large number of rifamycin derivatives by semi-synthetic approaches, the structural modifications have been limited primarily to one region of the molecule, the C-3 or C-4 positions of the aromatic core unit (Cricchio et al., 1974(Cricchio et al., , 1975Maggi et al., 1965; Wehrli et al., 1987). Alterations at other locations are difficult to accomplish chemically due to the complexity of the molecule, and require the implementation of alternative methodology, including combinatorial biosynthesis or mutasynthesis, to achieve additional...

NACT is a suitable option for patients with cervical cancer, especially for NACT responders and patients with stage IB, which provides a new concept of fertility preservation for young patients.

Considerable evidence suggests that monocytes/macrophages play a crucial role in the process of liver injury and repair. Recent investigations have focused on the function of various macrophage-produced cytokines in liver disease. Much is still unknown, however, about the mechanism of macrophage recruitment and activation during liver disease. To further define this process, the gene expression of the monocyte chemoattractant monocyte chemoattractant protein 1 (MCP-1) was examined in animal and human liver disease. MCP-1 mRNA was not found in normal rat liver by Northern blot analysis. After single-dose treatments with the hepatotoxins carbon tetrachloride and galactosamine, MCP-1 mRNA was detectable beginning at 2 and 4 h after treatment, respectively, and was expressed continuously until 60-72 h. During chronic carbon tetrachloride administration, MCP-1 mRNA levels were elevated for the entire 10 weeks of treatment with peak levels of expression occurring early (weeks 1-3) and late (weeks 8-10) in this model. Isolated liver cell fractions from rats treated for 3 weeks with carbon tetrachloride revealed the major cellular source of MCP-1 mRNA to be fat-storing or Ito cells, with some expression occurring in the endothelial cell fraction. Studies of potential inducers of hepatic MCP-1 expression showed that lipopolysaccharide, tumor necrosis factor-alpha, and interleukin-1 alpha and beta treatments all led to MCP-1 expression. Finally, studies of human liver samples revealed MCP-1 gene expression in nondiseased liver and greatly increased levels in livers from patients with fulminant hepatic failure. These data implicate MCP-1 from fat-storing cells as a modulator of the process of liver injury and further support a role for MCP-1 in the pathogenesis of human disease.