Phase II Randomized Study of Ixabepilone Versus Observation in Patients With Significant Residual Disease After Neoadjuvant Systemic Therapy for HER2-Negative Breast Cancer.

BACKGROUND: Residual disease (RD) after neoadjuvant chemotherapy carries an increased risk for recurrence. Ixabepilone has activity in anthracycline/taxanes-resistant breast cancer. We explored adjuvant ixabepilone in patients with significant RD HER2-negative breast cancer. METHODS: A phase II study in patients with residual cancer burden II or III randomized to ixabepilone versus observation was conducted. Circulating tumor cells (CTCs) were measured at baseline and at 9 and 18 weeks. Survival probabilities were estimated by Kaplan-Meier product limit. Toxicities were reported as proportions in the ixabepilone arm. RESULTS: Accrual was stopped because of ixabepilone toxicity. Sixty-seven patients were registered; 43 were randomized, 19 received ixabepilone, and 24 went to observation. One patient (9.1%) in the observation arm versus 2 patients (18.2%) in the ixabepilone arm had CTCs at 18 weeks (P = 1.0). Three-year recurrence-free survival and overall survival were 94% and 82%, and 100% and 79% in the observation and ixabepilone arms (P = .35 and .18), respectively. Most common adverse events (AEs) included fatigue, pain, neuropathy, constipation, nausea, rash, anorexia, and diarrhea. Serious AEs included pain (63.2%), fatigue (31.6%), and neuropathy (31.6%). CONCLUSIONS: Adjuvant ixabepilone in patients with significant RD after neoadjuvant chemotherapy was difficult to administer because of AEs and did not change the presence of CTC or affect survival outcomes. NCT00877500.

Functional consequence of the MET-T1010I polymorphism in breast cancer.

Major breast cancer predisposition genes, only account for approximately 30% of high-risk breast cancer families and only explain 15% of breast cancer familial relative risk. The HGF growth factor receptor MET is potentially functionally altered due to an uncommon germline single nucleotide polymorphism (SNP), MET-T1010I, in many cancer lineages including breast cancer where the MET-T1010I SNP is present in 2% of patients with metastatic breast cancer. Expression of MET-T1010I in the context of mammary epithelium increases colony formation, cell migration and invasion in-vitro and tumor growth and invasion in-vivo. A selective effect of MET-T1010I as compared to wild type MET on cell invasion both in-vitro and in-vivo suggests that the MET-T1010I SNP may alter tumor pathophysiology and should be considered as a potential biomarker when implementing MET targeted clinical trials.

Mechanism of 2-hydropropyl-beta-cyclodextrin in the stabilization of frozen formulations.

Freezing of commonly used parenteral products to increase pharmaceutical stability for cost-saving purposes is a common practice in patient care. However, frozen meropenem, a model drug, in saline has a shelf life of less than a month due to the low glass transition temperature (Tg'): below -40°C. When meropenem is formulated with the 2-hydroxypropyl-ß-cyclodextrin (HPBC) the shelf life (⩾90% potency) is extrapolated to be greater than one year at -25°C based on data for storage at 6months. The mechanisms that may explain meropenem-HPBC formulation frozen stability include vitrification and/or formation of an inclusion complex. Although NMR data indicated complexation of meropenem by HPBC in a ratio of 0.6:1, inclusion was unlikely to be the mechanism as stability was not extended to the thawed solutions. Therefore, vitrification is concluded to be the stabilization mechanism. The Tg' for meropenem-HPBC (13.3%) formulation at pH 7.9 was -17.75°C which was similar to that of a meropenem solution formulated with a known vitrifying agent, Dextran 40. This higher Tg' for HPBC was unexpected based on trends predicted by the Fox-Flory equation. Trial formulations containing either Dextran 1, Dextran 40, hydroxyethyl starch, or sulfobutyl-beta-cyclodextrin heptasodium (Captisol®) were also unable to stabilize meropenem as the Tg' values were below the frozen storage temperature. Upon 6-month storage, potency losses were -3.0% and -7.7% for meropenem frozen premix formulated in 13.3% HPBC (pH 7.9) at -25 and -20°C storage, respectively; versus -31.2% and -60.8% for controls. Frozen premixes with high ionic strength (containing NaCl or Captisol®) and/or at pH 7.3 were also found to be unstable.

Differences in gene and protein expression and the effects of race/ethnicity on breast cancer subtypes.

BACKGROUND: Differences in gene or protein expression patterns between breast cancers according to race/ethnicity and cancer subtype. METHODS: Transcriptional profiling was performed using Affymetrix HG-U133A platform in 376 patients and reverse phase protein array analysis (RPPA) was done for 177 proteins in 255 patients from a separate cohort. Unsupervised clustering was conducted, as well as supervised comparison by race and tumor subtype. Standard statistical methods, BRB-Array tools, and Ingenuity Pathways software packages were used to analyze the data. RESULTS: Median age was 50 years in both the cohorts. In the RPPA cohort, 54.5% of the tumors were hormone receptor-positive (HR-positive), 20.7% HER2-positive, and 24.71% triple-negative (TNBC). One hundred and forty-seven (57.6%), 47 (18.43%), and 46 (18.1%) of the patients were White, Hispanic, and Black, respectively. Unsupervised hierarchical clustering of the protein expression data showed no distinct clusters by race (P values were 0.492, 0.489, and 0.494 for the HR-positive, HER2-positive, and TNBC tumors respectively). In the gene expression cohort, 54.2% of the tumors were HR-positive, 16.5% HER2-positive, and 29.3% TNBC. Two hundred and sixteen (57.5%), 111 (29.52%), and 32 (8.52%) patients were White, Hispanic, and Black, respectively. No probe set with a false discovery rate (FDR) of <0.05 showed an association with race by breast cancer subtype; similar results were obtained using pathway and gene set enrichment analysis methods. CONCLUSIONS: We did not detect a significant variation in RNA or protein expression comparing different race/ethnicity groups of women with breast cancer. IMPACT: More research on the complex network of factors that result in outcomes differences among race/ethnicities is needed.

NMR of heparin API: investigation of unidentified signals in the USP-specified range of 2.12-3.00 ppm.

This article addresses the identification and quantification of the chemical species resulting in resonances at 2.17 and 2.25 ppm in the (1)H nuclear magnetic resonance (NMR) spectrum of pharmaceutical-grade heparin sodium. The NMR signals in question were first confirmed to arise from chemical moieties covalently attached to the heparin molecule through NMR diffusion experiments as well as chemical treatment of heparin active pharmaceutical ingredient (API) containing the resonances. The material responsible for the extra NMR signals was then demonstrated by NMR spiking studies to be something other than oversulfated chondroitin sulfate and was finally identified as an O-acetylation product of heparin through (13)C labeling experiments with subsequent NMR analysis. The extent of O-acetylation was quantified using three orthogonal techniques: (1)H NMR, ion chromatography, and headspace gas chromatography/mass spectrometry. The results of this work showed good agreement between the three quantitative methods developed to analyze the signals in the United States Pharmacopeia-specified region of 2.12-3.00 ppm for heparin API.

Aerosol based detectors for the investigation of phospholipid hydrolysis in a pharmaceutical suspension formulation.

A high performance liquid chromatography (HPLC) method was developed to quantify free fatty acids (FFA) in a pharmaceutical suspension formulated with phospholipids as stabilizing agents. Specifically, a suspension of crystalline itraconazole microparticles stabilized with Lipoid E80 was used as a model system to study the physicochemical stability of an aqueous, phospholipid-based suspension for injection. The hydrolysis of the phospholipids during storage at elevated temperatures (40 degrees C) necessitated the development of a suitable HPLC method for the determination of free fatty acid content in the suspension samples. HPLC methods using two types of aerosol detectors were investigated for the above purpose. Reversed-phase separation coupled with either an evaporative light scattering detector (ELSD) or a Corona(Plus) charged aerosol detector (CAD) was used. A comparison of the methods indicated that the CAD method provided better sensitivity, precision, recovery, and linearity for the parameters evaluated. As a result, this method was chosen for the stability study of itraconazole suspension and has been incorporated in subsequent formulation studies.

Physicochemical stability of phospholipid-dispersed suspensions of crystalline itraconazole.

The physicochemical stability of an aqueous, phospholipid-based dispersion of itraconazole microcrystals was studied as a model water-insoluble drug suspension. The particle size, phospholipid concentrations, free fatty acid (FFA) content, pH, and zeta potential of two test suspensions were followed over 63 days at 5 and 40 degrees C storage conditions. Hydrolysis of a control suspension containing Lipoid E80 led to rapid FFA formation, pH drop, and subsequent particle aggregation. In the second suspension, sodium oleate used in conjunction with Lipoid E80 significantly enhanced the suspension physicochemical stability. Oleate anions effectively (1) increased the anionic charge of the phospholipid surface layer, (2) buffered the suspension near pH 7, and (3) reduced the specific production of oleic acid as a phosphatidylcholine (PC) degradant. The observed hydrolysis rate constants k(obs) approximately 2 x 10(-7) (Lipoid only) and k(obs) approximately 5 x 10(-8) (Lipoid and oleate) were consistent with the pH dependent behavior reported for saturated soybean PC solutions. Mechanistically, FFA formed initially in the control suspension partitioned to the aqueous phase with limited influence on the phospholipid microenvironment at the itraconazole particle surface. Phospholipid stabilization of water-insoluble drugs was demonstrated with clear benefits from fatty acid anions as co-additives to influence the surface microenvironment, reduce hydrolysis kinetics, and enhance suspension physicochemical stability.

Solid-phase extraction and HPLC analysis of methylparaben and propylparaben in a concentrated antibiotic suspension.

An accurate and precise solid-phase extraction coupled with high performance liquid chromatography (SPE/HPLC) method developed for the quantification of antimicrobial preservatives (methylparaben and propylparaben) in oxytetracycline injectable suspension is described in this article. The SPE technique was necessary to quantify the preservatives since the high concentration of the drug and excipients was masking low levels of preservatives, making quantification difficult. This developed HPLC method was stability-indicating and found to be linear between 1.3 to 2.4 mg/mL for methylparaben and 0.15 to 0.27 mg/mL for propylparaben in this concentrated antibiotic suspension formulation. The extraction recoveries were 98.8-101.6%. System precision and sample extraction precision (RSD) were less than 1%.

Comparison of electrospray ionization mass spectrometry and evaporative light scattering detections for the determination of Poloxamer 188 in itraconazole injectable formulation.

A high performance liquid chromatography (HPLC) method for Poloxamer 188 using size-exclusion chromatography (SEC) was developed and two different detection mechanisms, evaporative light scattering (ELSD) and electrospray ionization mass spectrometry (ESI-MS), were compared for their quantification capabilities in itraconazole formulation. Both detection techniques coupled with SEC separation were highly effective for the determination of Poloxamer 188, which is difficult to analyze by other common HPLC methods. As expected, ESI-MS detection provided sensitivity and selectivity superior to ELSD. But since the analyte is an excipient in the formulation, high sensitivity was not required and ELSD's simplicity and ruggedness made it more appropriate for routine analysis of this formulation.

Determination of polysorbate 80 in parenteral formulations by high-performance liquid chromatography and evaporative light scattering detection.

Drugs that are not very soluble in aqueous formulations are solubilized with surfactants such as polysorbate 80. In order to evaluate the stability of excipient such as polysorbate 80 in drug formulation, a rapid chromatographic methodology is desired; however, polysorbate 80 does not have a strong chromophore for monitoring by absorption spectrometry. A simple and fast method for the analysis of polysorbate 80 in pharmaceutical formulations was developed using high-performance liquid chromatography with evaporative light scattering detection (ELSD). Separation of polysorbate 80 as a single peak was achieved on a C18 column using a methanol/water gradient mobile phase and ELS detection. The method is specific for polysorbate 80 in the formulation as there were no interferences from the drug or other excipients. Precision, recovery, linearity and limit of quantitation/detection experiments gave acceptable results during the evaluation of the method.