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Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers. It has an excessive desmoplastic stroma that can limit the intratumoral delivery of chemotherapy drugs, and protect tumor cells against radiotherapy. Therefore, both stromal and tumor compartments need to be addressed in order to effectively treat PDAC. We hereby co-deliver a sonic hedgehog inhibitor, cyclopamine (CPA), and a cytotoxic chemotherapy drug paclitaxel (PTX) with a polymeric micelle formulation (M-CPA/PTX). CPA can deplete the stroma-producing cancer-associated fibroblasts (CAFs), while PTX can inhibit tumor proliferation. Here we show that in clinically relevant PDAC models, M-CPA effectively modulates stroma by increasing microvessel density, alleviating hypoxia, reducing matrix stiffness while maintaining the tumor-restraining function of extracellular matrix. M-CPA/PTX also significantly extends animal survival by suppressing tumor growth and lowering the percentages of poorly to moderately differentiated tumor phenotypes. Our study suggests that using multifunctional nanoparticles to simultaneously target stromal and tumor compartments is a promising strategy for PDAC therapy.

Vincristine (VCR) is an integral part of chemotherapy regimens in the US and in developing countries. There is a paucity of information about its disposition and optimal therapeutic dosing. VCR is preferentially metabolized to its major M1 metabolite by the polymorphic CYP3A5 enzyme, which may be clinically significant as CYP3A5 expression varies across populations. Thus, it is important to monitor both VCR and M1 and characterize their dispositions. A previously developed HPLC-MS/MS method for VCR quantification was not sensitive enough to quantify the M1 metabolite beyond 1 hr. post VCR dose (not published). Establishing a highly sensitive assay is a pre-requisite to simultaneously quantify and monitor VCR and M1, which will enable characterization of drug exposure and dispositions of both analytes in a pediatric cancer population. The addition of formic acid during the extraction process enhanced M1 extraction from DBS samples. A sensitive, accurate, and precise UPLC-MS/MS assay method for the simultaneous quantification of VCR and M1 from human dried blood spots (DBS) was developed and validated. Chromatographic separation was performed on Inertsil ODS-3 C18 column (5 μm, 3.0 x 150 mm). A gradient elution of mobile phase A (methanol-0.2% formic acid in water, 20:80 v/v) and mobile phase B (methanol-0.2% formic acid in water, 80:20 v/v) was used with a flow rate of 0.4 mL/min and a total run time of 5 min. The analytes were ionized by electrospray ionization in the positive ion mode. The linearity range for both analytes in DBS were 0.6-100 ng/ml for VCR and 0.4-100 ng/ml for M1. The intra- and inter-day accuracies for VCR and M1 were 93.10-117.17% and 95.88-111.21%, respectively. While their intra- and inter-day precisions were 1.05 to 10.11% and 5.78 to 8.91%, respectively. The extraction recovery of VCR from DBS paper was 35.3 – 39.4% and 10.4 – 13.4% for M1, with no carryover observed for both analytes. This is the first analytical method to report the simultaneous quantification of VCR and M1 from human DBS. For the first time, concentrations of M1 from DBS patient samples were obtained beyond 1-hour post VCR dose. The developed method was successfully employed to monitor both compounds and perform pharmacokinetic analysis in a population of Kenyan pediatric cancer patients.

In the present study, a sensitive and robust LC–MS/MS method has been developed and validated for the quantification of riluzole in human plasma and cerebrospinal fluid (CSF) in clinical samples from patients with spinal cord injury (SCI). Riluzole and its labeled internal standard (IS) were isolated from plasma and CSF by liquid–liquid extraction using ethyl acetate. Riluzole ( m / z 235 → 166) and IS ( m / z 238 → 169) were detected by electrospray ionization (ESI) using multiple reaction monitoring (MRM) in a positive mode. The assay was linear in the concentration range of 0.5 (LLOQ, signal/noise ratio > 10)–800 ng/ml in plasma, and 1.0 (LLOQ)–800 ng/ml in CSF samples. The intra- and inter-day accuracy in plasma were 94.2–110.0% and 97.8–102.0%, respectively, and those in CSF were 87.6–105.1% and 91.9–98.8%, respectively. The intra- and inter-day precision were 2.2–7.2% and 4.0–9.1%, respectively, in plasma, and 1.4–14.1% and 2.6–11.5%, respectively in CSF. Matrix effect was negligible from both matrices with signal percentages of 97.6–100.6% in plasma and 99.4–106.4% in CSF. The recoveries were >75% in plasma, >84% in CSF with low protein (53.9 mg/dl), and >68% in CSF with high protein (348.2 mg/dl). This method was successfully applied to quantify riluzole concentrations in plasma and CSF from patients with SCI.

The aim of this study was to develop optimal micelles loaded with calcipotriol (Cal) and paclitaxel (PTX) for the treatment of pancreatic ductal adenocarcinoma (PDAC) using a Design of Experiment (DOE) approach. The central composite design (CCD), a type of DOE was used to tune the size and drug release properties of the drug-loaded micelles. This approach yielded optimal Cal and PTX co-loaded micelles (M-Cal/PTX) with size of 40-100 nm, a polydispersity index (PDI) of 0.25 and a zeta potential (ζ) of − 6.2 ± 0.8 mV. When evaluated in vitro, drug release from the micelles showed a biphasic pattern. The initial release, defined as the cumulative 2-hr drug release was less than 25% in all relevant media. This phase was followed by a gradual release with less than 80% of drugs released after 5 days. In vivo, the micelles prolonged the apparent biological half-life of Cal by more than 3 times and a marginal increase for PTX in an orthotopic mouse model of PDAC. The micelle-encapsulated drugs showed extended tumor accumulation when compared to non-encapsulated Cal and PTX at equivalent dose levels. Future studies on the antitumor activity of this novel dual drug payload delivery system are warranted.

The determination of dopamine facilitates better understanding of the complex brain disorders in the central nervous system and the regulation of endocrine system, cardiovascular functions and renal functions in the periphery. The purpose of this study was to develop a highly sensitive and reliable assay for the quantification of dopamine in human neonate plasma. Dopamine was extracted from human plasma by strong cation exchange (SCX) solid phase extraction (SPE), and subsequently derivatized with propionic anhydride. The derivatized analyte was separated by a Waters Acquity UPLC BEH C18 column using gradient elution at 0.4 ml/min with mobile phases A (0.2% formic acid in water [v/v]) and B (MeOH-ACN [v/v, 30:70]). Analysis was performed under positive electrospray ionization tandem mass spectrometer (ESI-MS/MS) in the multiple reaction monitoring (MRM) mode. The stable and relatively non-polar nature of the derivatized analyte enables reliable quantification of dopamine in the range of 10-1000 pg/ml using 200 μl of plasma sample. The method was validated with intra-day and inter-day precision less than 7%, and the intra-day and inter-day accuracy of 91.9-101.9% and 92.3-102.6%, respectively. The validated assay was applied to quantify dopamine levels in two preterm neonate plasma samples. In conclusion, a sensitive and selective LC-MS/MS method has been developed and validated, and successfully used for the determination of plasma dopamine levels in preterm neonates.

Irinotecan is a chemotherapeutic drug used in the treatment of advanced colorectal cancer and elevated blood concentrations of its active metabolite, SN-38 leads to increased gastrointestinal (GI) toxicity and diarrhea in patients. In this study, we investigated the effects of inflammation on the pharmacokinetics (PK) of irinotecan (CPT-11) and its active metabolite, SN-38. Mice were i.p.-injected with either saline or lipopolysaccharide (LPS) to induce inflammation. After 16 h, irinotecan was administered orally. Blood was collected from the tail vein of mice from 0 to 24 h after dosing. Concentrations of irinotecan, SN-38 and SN-38G were analyzed using LC-MS/MS. The AUC, Cmax, and tmax were derived using WinNonlin® 5.2. A PK model was developed using Phoenix NLME® to describe the PK of irinotecan and SN-38 during inflammation. Results indicated a significant increase in the blood concentrations of irinotecan and SN-38 in mice during inflammation. The AUC of irinotecan and SN-38 in LPS group were 2.6 and 2-folds, respectively, of those in control saline-treated mice. The Cmax of irinotecan and SN-38 in LPS treated mice were 2.4 and 2.3-folds of those in saline-treated mice. The PK model was successfully developed and validated. The best-fit plots of individual PK analysis showed a good correlation between observed and predicted concentrations of irinotecan and SN-38. Together, this study reveals that SN-38 concentrations are elevated during inflammation, which may increase the GI toxicity and diarrhea in patients who receive irinotecan; and the developed PK model can quantitatively describe the PK of irinotecan and SN-38 during inflammation.

Polymyxin B (PB) is increasingly used as the last treatment for multidrug-resistant (MDR) Gram-negative bacterial infections. In this study, serum and epithelial lining fluid (ELF) pharmacokinetics and the efficacy of a PB liposomal formulation were investigated. Two groups of 24 Swiss Webster mice were intravenously administered PB liposomes or PB aqueous solution at ca. 3 mg/kg. Serum and ELF samples were collected for up to 6 h to quantify major PB components. Three groups of neutropenic mice (n = 6/group) were infected with a clinical MDR Pseudomonas aeruginosa strain followed by intravenous administration of PB liposomes or PB aqueous solution at 3 mg/kg every 6 h or sham (drug-free) liposomes every 6 h. Bacterial burden in animal lung tissues was quantified after 24 h of therapy and was compared using one-way ANOVA. Survival of infected animals over time (n = 10/group) was evaluated by Kaplan-Meier analysis and log-rank test. In the pharmacokinetic study, the AUC ratio in ELF between liposome and aqueous solution groups ranged from 4.6 to 11.1 for various major PB components. In the efficacy study, for strain PA 9019 a significantly lower bacterial burden was seen in the liposomal group (3.8 ± 0.7 vs. 7.9 ± 0.8 log(10)CFU/g in the aqueous solution group), which subsequently prolonged survival of infected animals. In this study, treatment with a PB liposomal formulation yielded higher drug penetration into pulmonary ELF, which resulted in superior efficacy. However, further investigations on the clinical utility of the PB liposomal formulation are warranted.