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Purpose: This single-arm, open-label phase II study evaluated the safety and efficacy of taselisib (GDC-0032) plus fulvestrant in postmenopausal women with locally advanced or metastatic HER2-negative, hormone receptor (HR)-positive breast cancer.Patients and Methods: Patients received 6-mg oral taselisib capsules daily plus intramuscular fulvestrant (500 mg) until disease progression or unacceptable toxicity. Tumor tissue (if available) was centrally evaluated for PIK3CA mutations. Adverse events (AE) were recorded using NCI-CTCAE v4.0. Tumor response was investigator-determined using RECIST v1.1.Results: Median treatment duration was 4.6 (range: 0.9–40.5) months. All patients experienced ≥1 AE, 30 (50.0%) had grade ≥3 AEs, and 19 (31.7%) experienced 35 serious AEs. Forty-seven of 60 patients had evaluable tissue for central PIK3CA mutation testing [20 had mutations, 27 had no mutation detected (MND)]. In patients with baseline measurable disease, clinical activity was observed in tumors with PIK3CA mutations [best confirmed response rate: 38.5% (5/13; 95% CI, 13.9–68.4); clinical benefit rate (CBR): 38.5% (5/13; 95% CI, 13.9–68.4)], PIK3CA-MND [best confirmed response rate: 14.3% (3/21; 95% CI, 3.0–36.3); CBR: 23.8% (5/21; 95% CI, 8.2–47.2)], and unknown PIK3CA mutation status [best confirmed response rate: 20.0% (2/10; 95% CI, 2.5–55.6); CBR: 30.0% (3/10; 95% CI, 6.7–65.2)].Conclusions: Taselisib plus fulvestrant had clinical activity irrespective of PIK3CA mutation status, with numerically higher objective response rate and CBR in patients with PIK3CA-mutated (vs. -MND) locally advanced or metastatic HER2-negative, HR-positive breast cancer. No new safety signals were reported. A confirmatory phase III trial is ongoing. Clin Cancer Res; 24(18); 4380–7. ©2018 AACR.

Background: The phosphatidylinositol 3-kinase (PI3K) pathway is frequently dysregulated in hormone receptor (HR)-positive breast cancer (BC), with activating mutations of PIK3CA detected in ~35–45% of patients (pts). Acquired mutations in the ESR1 gene, which encodes estrogen receptor α, may be associated with resistance to aromatase inhibitor (AI) therapy. Taselisib is a potent and selective PI3K inhibitor, with greater selectivity against mutant PI3Kα isoforms than wild-type (WT) via a unique mechanism. In phase I studies, taselisib plus fulvestrant had clinical activity and manageable tolerability in pts with HR-positive BC. We report exploratory analyses of PIK3CA and ESR1 from circulating tumor DNA (ctDNA). Methods: In this phase II, open-label, single-arm study (PMT4979g; NCT01296555), pts were postmenopausal with HER2-negative, HR-positive locally advanced or metastatic BC and progression or non-response to ≥1 prior endocrine therapy in the adjuvant or metastatic setting. Pts received taselisib (6 mg capsule orally, daily) plus fulvestrant (500 mg intramuscular on Days 1 and 15 of Cycle 1, then Day 1 of each 28-day cycle) until disease progression or unacceptable toxicity. PIK3CA-mutation testing on archival tumor tissue used the cobas® PIK3CA Mutation Test. The Sysmex Inostics' BEAMing Digital PCR platform was used for ctDNA analysis of ESR1 and PIK3CA mutations (pre-dose on Cycle 1, Day 1). Primary endpoints were objective response rate (ORR) and clinical benefit rate (CBR) in all pts and those with PIK3CA mutations. ORR was confirmed complete response (cCR) and confirmed partial response (cPR). CBR was cCR, cPR, or stable disease for ≥6 months. Secondary endpoints included safety, efficacy, pharmacokinetics, and exploratory biomarker analysis. Results: 60 pts were enrolled. Median age was 61.5 years (range 31–82). In the metastatic setting, pts had received prior chemotherapy (21.7%) and prior hormonal therapy (50.0%). 86.7% of pts had received prior treatment with an AI. 45 pts had PIK3CA mutation status from archival tumor tissue and ctDNA testing; concordance was 86.7% (39/45). ctDNA analysis, vs archival tumor tissue testing, identified 4 pts and 9 pts with PIK3CA mutations from pts with WT and unknown PIK3CA mutation status, respectively. Based on ctDNA analysis (N=60), 13 pts (21.7%) had mutations in both ESR1 and PIK3CA, 21 pts (35.0%) were 'mutation not detected' (MND) for both genes, 8 (13.3%) had ESR1 mutations and PIK3CA MND, and 18 (30.0%) had ESR1 MND and PIK3CA mutations. In pts with measurable disease at baseline, confirmed responses (all partial) were: PIK3CA mutation, 38.1% (8/21); PIK3CA MND, 8.7% (2/23); all pts, 22.7% (10/44). CBRs were: PIK3CA mutation, 42.9%; PIK3CA MND, 17.4%; all pts, 29.5%. ORR and CBR from ctDNA analyses were similar to archival tumor tissue data. Conclusions: ctDNA analysis identified PIK3CA mutations in pts with previously unknown or WT mutation status from archival tumor tissue; ORR and CBR were similar to those from archival tumor tissue suggesting that PIK3CA mutation testing from ctDNA may be used as a surrogate when tissue is unavailable. 21.7% of pts had mutations in both ESR1 and PIK3CA. Citation Format: Dickler MN, Saura C, Oliveira M, Richards DA, Krop IE, Cervantes A, Stout TJ, Jin H, Savage HM, Wilson TR, Baselga J. Phase II study of taselisib (GDC-0032) plus fulvestrant in HER2-negative, hormone receptor-positive advanced breast cancer: Analysis by PIK3CA and ESR1 mutation status from circulating tumor DNA [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P6-12-01.

Summary Background Endocrine therapy-based neoadjuvant treatment for luminal breast cancer allows efficient testing of new combinations before surgery. The activation of the phosphatidylinositol-3-kinase (PI3K) pathway is a known mechanism of resistance to endocrine therapy. Taselisib is an oral, selective PI3K inhibitor with enhanced activity against PIK3CA-mutant cancer cells. The LORELEI trial tested whether taselisib in combination with letrozole would result in an increased proportion of objective responses and pathological complete responses. Methods In this multicentre, randomised, double-blind, parallel-cohort, placebo-controlled phase 2, study, we enrolled postmenopausal women (aged ≥18 years) with histologically confirmed, oestrogen receptor (ER)-positive, HER2-negative, stage I–III, operable breast cancer, from 85 hospitals in 22 countries worldwide. To be eligible, patients had have an Eastern Cooperative Oncology Group (ECOG) performance status 0–1, adequate organ function, and had to have evaluable tumour tissue for PIK3CA genotyping. Patients were randomly assigned (1:1) by means of a permuted block algorithm (block size of four) via an interactive voice or web-based response system, to receive letrozole (2·5 mg/day orally, continuously) with either 4 mg of oral taselisib or placebo (on a 5 days-on, 2 days-off schedule) for 16 weeks, followed by surgery. Randomisation was stratified by tumour size and nodal status. Site staff, patients, and the sponsor were masked to treatment assignment. Coprimary endpoints were the proportion of patients who achieved an objective response by centrally assessed breast MRI and a locally assessed pathological complete response in the breast and axilla (ypT0/Tis, ypN0) at surgery in all randomly assigned patients and in patients with PIK3CA-mutant tumours. Analyses were done in the intention-to-treat population. This trial is registered with ClinicalTrials.gov , number NCT02273973 , and is closed to accrual. Findings Between Nov 12, 2014, and Aug 12, 2016, 334 participants were enrolled and randomly assigned to receive letrozole and placebo (n=168) or letrozole and taselisib (n=166). Median follow-up was 4·9 months (IQR 4·7–5·1). The study met one of its primary endpoints: the addition of taselisib to letrozole was associated with a higher proportion of patients achieving an objective response in all randomly assigned patients (66 [39%] of 168 patients in the placebo group vs 83 [50%] of 166 in the taselisib group; odds ratio [OR] 1·55, 95% CI 1·00–2·38; p=0·049) and in the PIK3CA-mutant subset (30 [38%] of 79 vs 41 [56%] of 73; OR 2·03, 95% CI 1·06–3·88; p=0·033). No significant differences were observed in pathological complete response between the two groups, either in the overall population (three [2%] of 166 in the taselisib group vs one [1%] of 168 in the placebo group; OR 3·07 [95% CI 0·32–29·85], p=0·37) or in the PIK3CA-mutant cohort (one patient [1%) vs none [0%]; OR not estimable, p=0·48). The most common grade 3–4 adverse events in the taselisib group were gastrointestinal (13 [8%] of 167 patients), infections (eight [5%]), and skin–subcutaneous tissue disorders (eight [5%]). In the placebo group, four (2%) of 167 patients had grade 3 or worse vascular disorders, two (1%) had gastrointestinal disorders, and two (1%) patients had grade 3 or worse infections and infestations. There was no grade 4 hyperglycaemia and grade 3 cases were asymptomatic. Serious adverse events were more common in the taselisib group (eight [5%] patients with infections and seven [4%] with gastrointestinal effects) than in the placebo group (one [1%] patient each with grade 3 postoperative wound and haematoma infection, grade 4 hypertensive encephalopathy, grade 3 acute cardiac failure, and grade 3 breast pain). One death occurred in the taselisib group, which was not considered to be treatment-related. Interpretation The increase in the proportion of patients who achieved an objective response from the addition of taselisib to endocrine therapy in a neoadjuvant setting is consistent with the clinical benefit observed in hormone receptor-positive, HER2-negative, metastatic breast cancer. Funding Genentech and F Hoffmann-La Roche.

Background HR+/HER2- BC is the most common BC subtype, and adjuvant endocrine therapies (ET) are an integral part of its management; however, ~30% of patients still relapse. Development of ET resistance remains a challenge in HR+/HER2- BC; aberrant phosphoinositide 3-kinase (PI3K) signaling contributes to ET resistance and PIK3CA mutations occur in ~40% of HR+/HER2- BCs. Current research has led to the development of new targeted therapies, including CDK4/6 inhibitors and PI3K inhibitors (alpelisib). Preclinical models have demonstrated synergy between CDK4/6 inhibitors and PI3K inhibitors, with PI3K inhibitors blocking CDK4/6 inhibitor resistance development. GDC-0077 is a potent, selective PI3Kα inhibitor and a mutant p110α-degrader with anti-tumor activity alone and in combination with ET + P in PIK3CA-mutant preclinical models. An ongoing phase I trial showed that GDC-0077 + P + F could be combined at maximum doses. INAVO120 is a phase III, randomized, double-blind, pbo-controlled study that will assess efficacy and safety of GDC-0077/pbo + P + F in patients with PIK3CA-mutant/HR+/HER2- LA/MBC (NCT04191499). Trial design Patients are randomized 1:1 to GDC-0077 (9 mg orally; once daily [QD] continuously on a 28-day cycle) + P (125 mg orally; QD, days 1-21 of each 28-day cycle) + F (500 mg intramuscularly every 28 days with a loading dose in Cycle 1) or pbo + P + F. Circulating tumor DNA or tumor tissue must be positive for a PIK3CA mutation. Eligibility Patients whose disease progressed during/within 12 months of adjuvant ET completion, and who have not received prior systemic therapy for LA/MBC. Aims The primary endpoint is investigator-assessed progression-free survival (PFS) using Response Evaluation Criteria in Solid Tumors v1.1. Secondary endpoints include objective response rate (ORR); best overall response (BoR); duration of response (DoR); clinical benefit rate (CBR); overall survival (OS); safety; patient-reported outcomes; and pharmacokinetics. Statistical methods Patients are stratified by visceral disease, primary vs secondary resistance, and geographic region. For the primary PFS analysis, the treatment arms will be compared using a two-sided stratified log-rank test. The treatment effect will be quantified via a hazard ratio, computed from a stratified Cox proportional-hazards regression, including a 95% confidence interval. Median PFS will be estimated using Kaplan-Meier methodology. Accrual Target enrollment is 400 patients at ~210 sites globally. The study is open for enrollment and, as of 06/04/2020, eight patients have been enrolled. Contact information For more information or to refer a patient, email global.rochegenentechtrials@roche.com or call 1-888-662-6728 (USA only). Originally submitted to the ESMO 2020 Virtual Meeting. Citation Format: Nicolas Turner, Komal Jhaveri, Kevin Kalinsky, Sibylle Loibl, Sherene Loi, Seock-Ah Im, Cristina Saura, Peter Schmid, Jennifer L Schutzman, Thomas J Stout, Guiyuan Lei, Katherine E Hutchinson, Eirini Thanopoulou, Dejan Juric. Phase III study of GDC-0077 or placebo (pbo) with palbociclib (P) + fulvestrant (F) in patients with PIK3CA-mutant/hormone receptor-positive/HER2-negative locally advanced or metastatic breast cancer (HR+/HER2- LA/MBC) [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr OT-36-01.

A series of biaryl pyrazole and imidazole Liver X Receptor (LXR) partial agonists has been synthesized displaying LXRβ selectivity. The LXRβ selective partial agonist 18 was identified with potent induction of ATP binding transporters ABCA1 and ABCG1 in human whole blood (EC50 = 1.2 μM, 55% efficacy). In mice 18 displayed peripheral induction of ABCA1 at 3 and 10 mpk doses with no significant elevation of plasma or hepatic triglycerides at these doses, showing an improved profile compared to a full pan-agonist.

1050 Background: Taselisib (T) is an oral, potent, selective inhibitor of Class I PI3-kinase with enhanced activity against PIK3CA mutant cancer cells. Results from the LORELEI trial have demonstrated a significant improvement in ORR (objective response rate) by centrally assessed magnetic resonance imaging in all randomized patients as well as in the PIK3CA mutant (MT) cohort treated with neoadjuvant T plus letrozole (L) compared to placebo (P) plus L. Here we present the results of exploratory analyses of selected pathway-related phosphoproteins. Methods: Baseline (BL) and week3 (W3) tumor biopsies were obtained from 334 patients enrolled in the trial. Phosphoproteins (pAKT, pPRAS40 and pS6) were analyzed by IHC. BL levels as well as changes from BL to W3 were correlated with response assessed either by ORR or cell cycle arrest (Ki67 at W3 < 2.7%). Results: In the overall population, BL phosphoproteins levels were similar between the T and P arms. Higher pAKT (p < 0.001) and pPRAS40 (p = 0.004) levels were observed in MT vs wild-type (WT), whereas the opposite result was found for pS6 (p = 0.03). Treatment-induced absolute changes of phosphoproteins adjusted for BL levels were not significantly different between the T and P arms in the overall population, except for pPRAS40 with higher decrease in the T arm (p = 0.014). After stratification for PIK3CA genotype, a significantly greater decrease in expression levels was observed for pPRAS40 (p < 0.001) and pS6 (p = 0.020) in MT tumors treated with T. The treatment effects were not significantly different in the WT population. A trend for an association between decrease in pS6 levels at W3 and improved ORR was observed in the MT (p = 0.08) and T (p = 0.09) subgroups. The magnitude of pS6 suppression at W3 was higher in tumors achieving a cell cycle arrest in the MT/T subgroup (biserial correlation = -0.473). Conclusions: Exploratory analyses of phosphoproteins showed bioactivity of taselisib as indicated by downstream pathway suppression. Translational research aiming to integrate these results with additional exploratory biomarkers data is currently ongoing. Clinical trial information: NCT02273973.

Background: Taselisib is an oral, potent, selective inhibitor of Class I PI3-kinase (PI3K) alpha, gamma, and delta isoforms with enhanced activity against PIK3CA mutant cancer cells. LORELEI trial demonstrated a significant improvement in ORR (objective response rate) centrally assessed by MRI with neoadjuvant taselisib plus letrozole compared to letrozole plus placebo in all randomized patients as well as in the PIK3CA mutant cohort (Saura et al, ESMO 2017). Methods: 334 postmenopausal women with newly diagnosed ER+/HER2-, untreated, Stage I-III operable breast cancer and evaluable tumor tissue for PIK3CA genotyping were randomized (1:1) to receive daily letrozole (2.5 mg) with either taselisib (4 mg on a 5 days on/ 2 days off schedule) or placebo for 16 weeks, followed by surgery. Tumor tissue collection was performed at baseline, week 3 (W3) and at surgery. Secondary objectives included, but were not restricted to, ORR assessed by MRI in patients with PIK3CA wild type (WT) tumors, ORR using alternative methods of tumor assessment (ultrasound, mammogram and clinical breast exam) in all patients and patients with PIK3CA mutant and WT tumors, central assessment of Ki67 at different timepoints (baseline, W3 and surgery), and the centrally derived PEPI score. Central Ki67 was assessed by two independent readers blinded to treatment arms and PIK3CA status (Vall D'Hebron Institute of Oncology, Barcelona). Results: ORR by centrally assessed MRI was similar in the two treatment arms in patients with PIK3CA WT tumors (45.7 vs 40.4% for taselisib and placebo, respectively). ORR assessed by breast US was also significantly higher with taselisib compared to placebo in all randomized patients and in the PIK3CA mutant cohort. The highest concordance rate between MRI and other imaging modalities was found with breast ultrasound (53.7%). Centrally assessed Ki67 changes are reported in Table 1. Ki67 values decreased from baseline to W3 and from baseline to surgery in both treatment arms. No significant differences in the decrease of Ki67 values between treatment arms were detected. Unplanned analysis of Complete Cell Cycle Arrest (CCCA) at W3 was numerically higher with taselisib than with placebo in all randomized patients (49.6% vs 38.5%) and in the PIK3CA mutant cohort (60.9% vs 47.5%). Due to the variability in timing between the last dose of taselisib (median time 11 days; interquartile range 6-16 days) and tissue collection at surgery, considering the half-life of taselisib of approximately 40 hours, centrally derived PEPI score is not interpretable. Ki67 proportional changes, %Taselisib + letrozolePlacebo + letrozoleBaseline to W3All patients-83.8-80.4PIK3CA mutant-84.5-79.1PIK3CA WT-82.8-81.1Baseline to surgeryAll patients-75.6-80.5PIK3CA mutant-71.9-79.9PIK3CA WT-78.2-81.2 Conclusion: Among the investigated alternative methods for assessing ORR, breast ultrasound performed similar to MRI. Decrease in the Ki67 values from baseline to W3 and to surgery were observed in both treatment arms. The time interval between taselisib cessation and tissue collection at surgery are being further investigated. Clinical trial information: NCT02273973 Citation Format: de Azambuja E, Saura C, Nuciforo P, Frantal S, Oliveira M, Zardavas D, Jallitsch-Halper A, de la Pena L, Dubsky P, Lombard JM, Vuylsteke P, Castaneda Altamirano C, Sanchez C, Ballestrero A, Colleoni M, Santos Borges G, Ciruelos E, Bardia A, Fornier M, Boer K, Wilson TR, Stout TJ, Hsu JY, Shi Y, Piccart M, Baselga J, Gnant M. Ki67 changes and PEPI score in the LORELEI trial: A phase II randomized, double-blind study of neoadjuvant letrozole plus taselisib versus letrozole plus placebo in postmenopausal women with ER-positive/HER2-negative early-stage breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr PD5-04.

Oncogenic BRAF alleles are both necessary and sufficient for cellular transformation, suggesting that chemical inhibition of the activated mutant protein kinase may reverse the tumor phenotype. Here, we report the characterization of SB-590885, a novel triarylimidazole that selectively inhibits Raf kinases with more potency towards B-Raf than c-Raf. Crystallographic analysis revealed that SB-590885 stabilizes the oncogenic B-Raf kinase domain in an active configuration, which is distinct from the previously reported mechanism of action of the multi-kinase inhibitor, BAY43-9006. Malignant cells expressing oncogenic B-Raf show selective inhibition of mitogen-activated protein kinase activation, proliferation, transformation, and tumorigenicity when exposed to SB-590885, whereas other cancer cell lines and normal cells display variable sensitivities or resistance to similar treatment. These studies support the validation of oncogenic B-Raf as a target for cancer therapy and provide the first evidence of a correlation between the expression of oncogenic BRAF alleles and a positive response to a selective B-Raf inhibitor. (Cancer Res 2006; 66(23): 11100-5)

Unlike all other organisms studied to date, Bacillus subtilis expresses two different thymidylate synthases: bsTS-A and bsTS-B. bsTS-A displays enhanced enzymatic and structural thermal stability uncharacteristic of most TSs. Despite the high level of TS conservation across most species, bsTS-A shares low sequence identity (

Background: Thymidylate synthase (TS) is critical to DNA synthesis as it catalyzes the rate limiting step in the only biosynthetic pathway for deoxythymidine monophosphate (dTMP) production. TS is therefore an important target for anti-proliferative and anti-cancer drug design. The TS enzymatic mechanism involves the reductive methylation of the substrate, deoxyuridine monophosphate (dUMP), by transfer of a methylene group from the co-factor, methylenetetrahydrofolate (CH 2 H 4 folate), resulting in the production of deoxythymidine monophosphate (dTMP) and dihydrofolate (H 2 folate). Previous drug design efforts based on co-factor analogues have produced good inhibitors of TS, but poor bioavailability and toxicity have limited their usefulness. BW1843U89, a folate analogue, is a recently developed compound which is an exceptionally strong inhibitor (K i =0.09 nM), has good bioavailability and in clinical trials thus far has not demonstrated significant toxicity. Results We report the crystal structure of E. coli TS in ternary complex with dUMP and BW1843U89 at 2.0 a resolution. Although the benzoquinazoline ring system of the inhibitor binds to TS in much the same manner as previously determined for H 2 folate and CB3717, the larger size of the ligand is accommodated by the enzyme through a local distortion of the active site, that is not strictly conserved in both monomers in the asymmetric unit. Several conserved waters that had been previously implicated in mechanistic roles have been displaced. Conclusion BW1843U89 forms a ternary complex with dUMP and competes with CH 2 H 4 folate at the active site. Inhibition of TS by BW1843U89 shows four unique aspects in its mechanism of action. BW1843U89 prevents the Michael addition of dUMP to Cys146, in contrast to the mechanisms implicated from crystallography of other quinazoline based inhibitors; displaces a catalytic water from the active site; reorders a peptide loop (Leu72–Trp83) in the active site; and is unique amongst the antifolates in inactivating TS at a stoichiometric ratio of one molecule per TS dimer. Thus, it exploits the principles of negative cooperativity that are increasingly being recognized in the catalytic mechanism of the enzyme per se . The structure suggests that this ‘half-the-sites' effect is catalytic and not related to ligand binding. Therefore BW1843U89 is both a competitive inhibitor (at the binding site) and a non-competitive inhibitor at the other site.

520Background: The PI3K pathway is activated in HR+ BC, often via gain-of-function mutations in PIK3CA that occur in ~40% of HR+ BC. Taselisib is a potent and selective PI3K inhibitor, with greater selectivity against mutant PI3Kα isoforms than wild type (WT) PI3Kα. Phase Ib data demonstrated good tolerability and preliminary efficacy for taselisib + F in HR+ BC. Methods: This Phase II, open-label, single-arm study enrolled post-menopausal pts with HER2-, HR+ locally advanced or metastatic BC (mBC) who had progression or non-response to ≥ 1 prior endocrine therapy in adjuvant or mBC settings. Pts received taselisib (6 mg capsule PO qd) plus F (500 mg IM on Cycle 1 Day 1, Cycle 1 Day 15, then q4w Day 1 of each cycle) until progressive disease or unacceptable toxicity. PIK3CA mutation status was centrally confirmed retrospectively on archival tumor tissue by 'cobas' PIK3CA Mutation Test. Primary endpoints were objective response rate (RECIST version 1.1) and clinical benefit rate (CBR; confirmed CR and PR, ...

Emeniveol (1), a new pollen growth inhibitor, was isolated from the fungus, Emericella nivea. The structure was elucidated from the X-ray crystallographic analysis as well as homonuclear decoupling, COSY, COLOC and HMBC experiments, and chemical reaction.

A convergent asymmetric synthesis of the antinoeplastic macrolide antibiotic cytovaricin has been achieved through the synthesis and coupling of the illustrated spiroketal and polyol glycoside subunits. All absolute steroechemical relationships within the target structure were ultimately controlled by the use of asymmetric aldol, alkylation, or epoxidation methodology. Union of the two subnits was accomplished by Julia-Lythgoe trans olefination, providing direct access to a suitable macrolactonization substrate

Serpins exhibit a range of physiological roles and can contribute to certain disease states dependent on their various conformations. Understanding the mechanisms of the large-scale conformational reorganizations of serpins may lead to a better understanding of their roles in various cardiovascular diseases. We have studied the serpin, plasminogen activator inhibitor 1 (PAI-1), in both the active and the latent state and found that anionic halide ions may play a role in the active-to-latent structural transition. Crystallographic analysis of a stable mutant form of active PAI-1 identified an anion-binding site between the central beta-sheet and a small surface domain. A chloride ion was modeled in this site, and its identity was confirmed by soaking crystals in a bromide-containing solution and calculating a crystallographic difference map. The anion thus located forms a 4-fold ligated linchpin that tethers the surface domain to the central beta-sheet into which the reactive center loop must insert during the active-to-latent transition. Timecourse experiments measuring active PAI-1 stability in the presence of various halide ions showed a clear trend for stabilization of the active form with F(-)Cl(-)Br(-)I(-). We propose that the "stickiness" of this pin (i.e., the electronegativity of the anion) contributes to the energetics of the active-to-latent transition in the PAI-1 serpin.

Duodenal ulcers were produced in rats by subcutaneous infusion for 48 hr of either of the following secretagogues: histamine, carbachol, pentagastrin. When any two of these secretagogues were administered together, synergism was observed in the ulcerogenic effect. Synergism, or potentiation, is defined as an effect produced by two agents given together that is greater than the sum of the effects produced by each agent when given separately. The ulcers were located opposite to the mesenteric attachment, were often multiple, and extended as far as 4 cm from the pylorus. Their severity ranged from shallow erosions to perforation with peritonitis. When two secretagogues were infused at the same time, the incidence of animals with ulcers, their severity, number, and distance from the pylorus were related to the doses given. Pentagastrin, when administered alone, was the least ulcerogenic agent, but when pentagastrin was given with either histamine or carbachol, the ulcerogenic effect was potentiated. Although the duodenum was consistently ulcerated, the stomach remained intact, except in some animals receiving high doses of secretagogues and dying or dead from peritonitis. It is concluded that: (1) the rat can develop duodenal ulcers; and (2) secretagogues, when infused for 48 hr and especially when combined, can produce such ulcers; (3) the mechanism by which these ulcers develop remains to be elucidated.

Introduction: EGFR- mutant lung cancers are sensitive to EGFR tyrosine kinase inhibitors (TKIs). Unfortunately, they develop resistance, often due to acquisition of a second-site mutation (T790M). Current EGFR TKIs select for T790M in preclinical models of acquired resistance. We explored whether all EGFR TKIs similarly select for the T790M mutation using data from early clinical trials and established in vitro models of acquired resistance. Methods: We analyzed the clinical characteristics of eight patients with metastatic EGFR-mutant lung adenocarcinoma who were treated first-line with XL647 and then progressed. XL647 is an ATP-competitive inhibitor of EGFR, HER2, KDR, and EPHB4. Additional molecular preclinical studies were performed to characterize resistance. Results: Four patients displayed confirmed partial responses (PRs), three patients had unconfirmed PRs, and one patient displayed stable disease. Only one of five patients' tumor samples available for analysis after disease progression harbored the T790M mutation. Eight patients subsequently received erlotinib, with ( n = 3) or without ( n = 5) chemotherapy. Three of five patients treated with single-agent erlotinib derived additional benefit, staying on drug up to 9 months. EGFR-mutant PC-9 cells with acquired resistance to XL647 did not harbor the T790M mutation, displayed a distinct mRNA profile from PC-9 cells with T790M-mediated resistance, and were moderately sensitive to erlotinib in growth inhibition assays. Crystal structure analyses of XL647/EGFR T790M did not reveal a different binding mode from that of erlotinib. Conclusions: The findings of this exploratory study suggest that different EGFR TKIs may select for distinct mechanisms of resistance. These results raise the possibility that different EGFR TKIs could be sequentially used to improve outcomes in patients with EGFR -mutant lung cancer. Further work investigating this hypothesis is warranted.