CHF5074 reduces biomarkers of neuroinflammation in patients with mild cognitive impairment: a 12-week, double-blind, placebo-controlled study.

As neuroinflammation is an early event in the pathogenesis of Alzheimer' s disease, new selective antiinflammatory drugs could lead to promising preventive strategies. We evaluated the safety, tolerability, pharmacokinetics and pharmacodynamics of CHF5074, a new microglial modulator, in a 12-week, double-blind, placebo-controlled, parallel groups, ascending dose study involving 96 MCI patients. Subjects were allocated into three successive study cohorts to receive ascending, titrated doses of CHF5074 (200, 400 or 600 mg/day) or placebo. Vital signs, cardiac safety, neuropsychological performance and safety clinical laboratory parameters were assessed on all subjects. Plasma samples were collected throughout the study for measuring drug concentrations, soluble CD40 ligand (sCD40L) and TNF-α. At the end of treatment, cerebrospinal fluid (CSF) samples were optionally collected after the last dose to measure drug levels, ß- amyloid1-42 (Aß42), tau, phospho-tau181, sCD40L and TNF-α. Ten patients did not complete the study: one in the placebo group (consent withdrawn), two in the 200-mg/day treatment group (consent withdrawn and unable to comply) and seven in the 400-mg/day treatment group (five AEs, one consent withdrawn and one unable to comply). The most frequent treatment-emergent adverse events were diarrhea, dizziness and back pain. There were no clinically significant treatmentrelated clinical laboratory, vital sign or ECG abnormalities. CHF5074 total body clearance depended by gender, age and glomerular filtration rate. CHF5074 CSF concentrations increased in a dose-dependent manner. At the end of treatment, mean sCD40L and TNF-α levels in CSF were found to be inversely related to the CHF5074 dose (p=0.037 and p=0.001, respectively). Plasma levels of sCD40L in the 600-mg/day group were significantly lower than those measured in the placebo group (p=0.010). No significant differences between treatment groups were found in neuropsychological tests but a positive dose-response trend was found on executive function in APOE4 carriers. This study shows that CHF5074 is well tolerated in MCI patients after a 12-week titrated treatment up to 600 mg/day and dose-dependently affects central nervous system biomarkers of neuroinflammation.

Pharmacokinetics and pharmacodynamics of CHF5074 after short-term administration in healthy subjects.

CHF5074 has been shown to inhibit brain ß-amyloid deposition and attenuate memory deficits in different transgenic mice models of Alzheimer disease. We evaluated the safety, pharmacokinetics, and pharmacodynamics of 3 ascending dose regimens of CHF5074 (200, 400, and 600 mg/d for 14 d) in a double-blind, placebo-controlled, parallel group study involving 48 healthy subjects. Plasma, urine, and cerebrospinal fluid (CSF) samples were collected for measuring drug and main metabolite concentrations and potential biomarkers of pharmacodynamic activity (ß-amyloid1-40, ß-amyloid1-42, soluble CD40 ligand, and tumor necrosis factor-α). All subjects completed the study, and no serious or severe adverse events were reported. The maximum tolerated dose was close to 600 mg/d with mild diarrhea being the most frequent adverse event at this dose. CHF5074 reached peak plasma levels 2 to 3 hours after drug administration and then was slowly eliminated (t(1/2z)=30 h) in the urine as glucoronide. Systemic exposure to the drug appeared to be dose-proportional with a 2-fold accumulation ratio at steady state. Metabolite plasma levels peaked at 4 to 5 hours and accounted for about 25% of the parent compound. Drug levels in the CSF were dose-proportional. The drug dose-dependently lowered the levels of the soluble CD40 ligand, a marker of microglia activation, in both plasma and CSF samples.

Aggregation properties of a HPMA-camptothecin copolymer in isotonic solutions.

Copolymers of camptothecin (CPT) and [N-(2-hydroxypropyl) methacrylamide] (HPMA) are novel anticancer drugs that show improved pharmacological profile in animal models as compared to the free drug CPT. We investigate here the aggregation properties of a HPMA-glycyl-6-aminohexanoyl-glycyl-CPT copolymer ( approximately 20,000 Da). The molecular size of HPMA-copolymer CPT is followed over 5 orders of magnitudes of concentration in isotonic buffer by measuring either the time resolved fluorescence anisotropy (FA) of CPT or the autocorrelation function of the light scattered by the copolymer. A detailed analysis of these data suggests the presence of elongated structures with axial ratio approximately 3 in the range 0.1-0.5 microg/ml and aggregates with association number higher than 2 in more concentrated solutions (up to 10 mg/ml). The binding affinity of HPMA-copolymer CPT for serum albumin is inversely dependent on the degree of aggregation of the copolymer. We also show that the copolymer concentration in plasma from mice treated with an active, non-toxic, dose of HPMA-copolymer CPT, decreases from 3 to 0.01 mg/ml in 72 h. In the same range of concentrations in vitro, we do not detect hydrophobic aggregates of polymers with high (>3) association number. Our study indicates that the circulating HPMA-copolymer CPT in mice should not undergo extensive aggregation and should interact with serum albumin more weakly than free CPT.

LC-MS-MS determination of brostallicin in human plasma following automated on-line SPE.

LC-MS-MS method using automated on-line solid-phase extraction (SPE) has been developed and validated for the quantitation of brostallicin (I), a new distamycin derivative, in human plasma. I is a DNA minor groove binder currently under phase I-II clinical evaluation as an anticancer drug. Plasma (0.4 ml) was spiked with 0.2 ml stable label IS solution and placed in a 96-well plate maintained at +4 degrees C. Aliquots of 0.1 ml of prepared samples were loaded into the on-line SPE HySphere Resin SH cartridges (10 mm x 2 mm ID) and the analytes back eluted with the mobile phase into LC-MS-MS system. A Platinum Cyano column (100 mm x 4.6 mm, 3.6 microm) was used to perform the chromatographic analysis. The mobile phase was acetonitrile-ammonium formate buffer (pH 3.5; 20 mM) (70:30, v/v) at a flow rate of 1.0 ml/min. LC flow was split so that 300 microl/min was directed toward the mass spectrometer interface. Retention time of I was 2.6 min and the total cycle time was 8 min. MS detection used an Applied Biosystems/MDS SCIEX API 365 with a TurboIonSpray interface and MRM (m/Z: 725/257 for I and m/Z: 729/257 for IS) operated in positive ion mode. The method was validated over the calibration range 0.124-497 ng/ml. A negligible carry-over effect from the system was observed. In spite of the known instability of I in human plasma (about 20% decrease over 12 h), the ratio analyte/IS peak area showed good stability over the analysis time required for 96 samples. The automated on-line SPE method can be considered as a valid alternative to the off-line manual SPE procedure previously developed.