As the first structural elucidation of a modular polyketide synthase (PKS) domain, the crystal structure of the macrocycle-forming thioesterase (TE) domain from the 6-deoxyerythronolide B synthase (DEBS) was solved by a combination of multiple isomorphous replacement and multiwavelength anomalous dispersion and refined to an R factor of 24.1% to 2.8-Å resolution. Its overall tertiary architecture belongs to the ␣͞␤-hydrolase family, with two unusual features unprecedented in this family: a hydrophobic leucine-rich dimer interface and a substrate channel that passes through the entire protein.The active site triad, comprised of Asp-169, His-259, and Ser-142, is located in the middle of the substrate channel, suggesting the passage of the substrate through the protein. Modeling indicates that the active site can accommodate and orient the 6-deoxyerythronolide B precursor uniquely, while at the same time shielding the active site from external water and catalyzing cyclization by macrolactone formation. The geometry and organization of functional groups explain the observed substrate specificity of this TE and offer strategies for engineering macrocycle biosynthesis. Docking of a homology model of the upstream acyl carrier protein (ACP6) against the TE suggests that the 2-fold axis of the TE dimer may also be the axis of symmetry that determines the arrangement of domains in the entire DEBS. Sequence conservation suggests that all TEs from modular polyketide synthases have a similar fold, dimer 2-fold axis, and substrate channel geometry. M odular polyketide synthases (PKSs) are a family of multienzyme complexes that synthesize the polyketide cores of biologically active compounds, including natural products that have become important pharmaceuticals, such as erythromycin, rifamycin, FK506, rapamycin, and avermectin (1). Their remarkable combination of substrate tolerance and selectivity is largely because of their modular architecture, in which different catalytic domains are combined into ''modules'' (Fig. 1), such that each module contains several enzymes and adds one additional building block to a growing polyketide chain. 6-Deoxyerythronolide B synthase (DEBS) is a modular PKS that catalyzes the biosynthesis of 6-deoxyerythronolide B (6-dEB, 1, Fig. 1), the macrocyclic core of the antibiotic erythromycin (2, 3). The entire DEBS is a homodimer containing two copies each of 28 catalytic domains organized into a loading didomain, six extension modules (each composed of several domains), and a terminal thioesterase (TE) that cyclizes and releases the final product, 6-dEB (1, Fig. 1). Combinatorial substitution of enzymes of DEBS, by others from rapamycin PKS, gave rise to over 100 novel compounds at varying yields (4). To date, no structure of a modular PKS component has been reported. When the structures of individual domains are determined, there will be opportunity to apply structure-based principles of protein engineering in next-generation efforts to rationally design novel polyketide products.The TE domain of...

IMPORTANCEThe humanized monoclonal antibody atezolizumab targets programmed death-ligand 1 and has demonstrated durable single-agent activity in a subset of metastatic triple-negative breast cancers. To extend the observed activity, combinatorial approaches are being tested with standard cytotoxic chemotherapies known to induce immunogenic tumor cell death.OBJECTIVE To examine the safety, tolerability, and preliminary clinical activity of atezolizumab plus nab-paclitaxel in metastatic triple-negative breast cancers. DESIGN, SETTING, AND PARTICIPANTSThis phase 1b multicohort study enrolled 33 women with stage IV or locally recurrent triple-negative breast cancers and 0 to 2 lines of prior chemotherapy in the metastatic setting from December 8, 2014, to April 30, 2017, at 11 sites in the United States. The median follow-up was 24.4 months (95% CI, 22.1-28.8 months). INTERVENTIONS Patients received concurrent intravenous atezolizumab and intravenous nab-paclitaxel (minimum 4 cycles). MAIN OUTCOMES AND MEASURESThe primary end point was safety and tolerability. Secondary end points included best overall response rate by Response Evaluation Criteria in Solid Tumors, version 1.1; objective response rate; duration of response; disease control rate; progression-free survival; overall survival; and biomarker analyses. RESULTSThe 33 women had a median age of 55 years (range, 32-84 years) and received 1 or more doses of atezolizumab. All patients (100%) experienced at least 1 treatment-related adverse event, 24 patients (73%) experienced grade 3/4 adverse events, and 7 patients (21%) had grade 3/4 adverse events of special interest. No deaths were related to study treatment. The objective response rate was 39.4% (95% CI, 22.9%-57.9%), and the median duration of response was 9.1 months (95% CI, 2.0-20.9 months). The disease control rate was 51.5% (95% CI, 33.5%-69.2%). Median progression-free survival and overall survival were 5.5 months (95% CI, 5.1-7.7 months) and 14.7 months (95% CI, 10.1-not estimable), respectively. Concurrent nab-paclitaxel neither significantly changed biomarkers of the tumor immune microenvironment (programmed death-ligand 1, tumor-infiltrating lymphocytes, CD8) nor impaired atezolizumab systemic immune activation (expansion of proliferating CD8+ T cells, increase of CXCL10 chemokine). CONCLUSIONS AND RELEVANCEIn this phase 1b trial for metastatic triple-negative breast cancers, the combination of atezolizumab plus nab-paclitaxel had a manageable safety profile. Antitumor responses were observed, including in patients previously treated with a taxane. TRIAL REGISTRATION ClinicalTrials.gov identifier: NCT01633970

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Azepino[4,5-b]indoles have been identified as potent agonists of the farnesoid X receptor (FXR). In vitro and in vivo optimization has led to the discovery of 6m (XL335, WAY-362450) as a potent, selective, and orally bioavailable FXR agonist (EC(50) = 4 nM, Eff = 149%). Oral administration of 6m to LDLR(-/-) mice results in lowering of cholesterol and triglycerides. Chronic administration in an atherosclerosis model results in significant reduction in aortic arch lesions.

BackgroundCabozantinib, an orally bioavailable inhibitor of tyrosine kinases including MET, AXL, and VEGF receptors, was assessed in patients with hepatocellular carcinoma (HCC) as part of a phase 2 randomized discontinuation trial with nine tumor-type cohorts.Patients and methodsEligible patients had Child-Pugh A liver function and ≤1 prior systemic anticancer regimen, completed ≥4 weeks before study entry. The cabozantinib starting dose was 100 mg daily. After an initial 12-week cabozantinib treatment period, patients with stable disease (SD) per Response Evaluation Criteria in Solid Tumors (RECIST) version 1.0 were randomized to cabozantinib or placebo. The primary endpoint of the lead-in stage was objective response rate (ORR) at week 12, and the primary endpoint of the randomized stage was progression-free survival (PFS).ResultsAmong the 41 HCC patients enrolled, the week 12 ORR was 5%, with 2 patients achieving a confirmed partial response (PR). The week 12 disease control rate (PR or SD) was 66% (Asian subgroup: 73%). Of patients with ≥1 post-baseline scan, 78% had tumor regression, with no apparent relationship to prior sorafenib therapy. Alpha-fetoprotein (AFP) response (>50% reduction from baseline) occurred in 9 of the 26 (35%) patients with elevated baseline AFP and ≥1 post-baseline measurement. Twenty-two patients with SD at week 12 were randomized. Median PFS after randomization was 2.5 months with cabozantinib and 1.4 months with placebo, although this difference was not statistically significant. Median PFS and overall survival from Day 1 in all patients were 5.2 and 11.5 months, respectively. The most common grade 3/4 adverse events, regardless of attribution, were diarrhea (20%), hand-foot syndrome (15%), and thrombocytopenia (15%). Dose reductions were utilized in 59% of patients.ConclusionsCabozantinib has clinical activity in HCC patients, including objective tumor responses, disease stabilization, and reductions in AFP. Adverse events were managed with dose reductions.Trial registration numberNCT00940225.

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The crystal structure of E. coli Fmu, determined at 1.65 A resolution for the apoenzyme and 2.1 A resolution in complex with AdoMet, is the first representative of the 5-methylcytosine RNA methyltransferase family that includes the human nucleolar proliferation-associated protein p120. Fmu contains three subdomains which share structural homology to DNA m(5)C methyltransferases and two RNA binding protein families. In the binary complex, the AdoMet cofactor is positioned within the active site near a novel arrangement of two conserved cysteines that function in cytosine methylation. The site is surrounded by a positively charged cleft large enough to bind its unique target stem loop within 16S rRNA. Docking of this stem loop RNA into the structure followed by molecular mechanics shows that the Fmu structure is consistent with binding to the folded RNA substrate.