Apatite nanoparticles strongly improve red blood cell cryopreservation by mediating trehalose delivery via enhanced membrane permeation.

Cryopreservation of red blood cells (RBC) is an important method for maintaining an inventory of rare RBC units and managing special transfusion circumstances. Currently, in a clinical setting, glycerol is used as cryoprotectant against freezing damage. After thawing and before transfusion, glycerol must however be removed to avoid intravascular hemolysis, via a complex and time-consuming deglycerolization process which requires specialized equipment. Improved cryopreservation methods using non-toxic agents are required to increase biocompatibility and decrease processing time. Biocompatible cryoprotectants (e.g. trehalose) were proposed, but their low permeation through RBC membranes limits their cryoprotection efficacy. Herein, we report for the first time a glycerol-free cryopreservation approach, using colloidal bioinspired apatite nanoparticles (NP) as bioactive promoters of RBC cryopreservation mediated by trehalose. Addition of apatite NP in the medium tremendously increases RBC cryosurvival, up to 91% (42% improvement compared to a control without NP) which is comparable to FDA-approved cryoprotection protocol employing glycerol. NP concentration and incubation conditions strongly modulate the NP bioactivity. Complementary experimental and computational analyses of the interaction between apatite NP and model lipid bilayers revealed complex events occurring at the NP-bilayer interface. Apatite NP do not cross the bilayer but momentarily modulate its physical status. These changes affect the membrane behavior, and promote the permeation of trehalose and a model fluorescent molecule (FITC). This approach is a new alternative to using toxic glycerol for cells cryopreservation, and the identification of this enhancing no-pore permeation mechanism of apatite NP appears as an original delivery pathway for cryoprotectant agents and beyond.

Novel method for fabrication of samples for cell testing of bioceramics in granular form.

BACKGROUND: Bioceramic granules are a widely studied material for regeneration of human tissues, and their biological assessment with in vitro cell cultures plays a fundamental role in the development of bioceramics. Design of samples for cell testing represents an important aspect of the biological evaluation, as it dictates how cells will interact with the biomaterial. The aim of this study was to develop samples for cell testing of bioceramic granules with a novel design that would enable direct physical contacts between cells and bioceramic and improved handling properties for efficient laboratory work. The goal was to produce a bilayered polycaprolactone-bioceramic composite with polycaprolactone serving as a bottom layer and support for a uniform and dense layer of bioceramic granules (upper layer), which would be only partly embedded and physically stabilized in the polymer with at least one face of granules still free of any polymer residues and available for direct attachment of cells. METHODS: A novel method for preparation of samples in six steps was developed. A bilayered design of samples with exposed bioceramic particles was accomplished by the application of a water-soluble alginate as a sacrificial polymer in the method protocol. Samples were analyzed with SEM/EDX and ToF-SIMS. RESULTS: Bioceramic granules had a uniform and dense morphology and were partly embedded in the polycaprolactone support. Detailed ToF-SIMS study showed that granules were clean and free of any polymer residues. CONCLUSIONS: The developed samples enable direct exposure of bioceramic granules to cells and surrounding physiological solution during cell testing, and possess improved handling characteristics.

Phase I study of nintedanib incorporating dynamic contrast-enhanced magnetic resonance imaging in patients with advanced solid tumors.

BACKGROUND: This open-label phase I dose-escalation study investigated the safety, efficacy, pharmacokinetics (PK), and dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) effects of the oral angiokinase inhibitor nintedanib in patients with advanced solid tumors. METHODS: Nintedanib was administered once daily continuously, starting at 100 mg and later amended to allow evaluation of 250 mg b.i.d. The primary endpoint was maximum tolerated dose (MTD). DCE-MRI studies were performed at baseline and on days 2 and 28. RESULTS: Fifty-one patients received nintedanib 100-450 mg once daily (n = 40) or 250 mg b.i.d. (n = 11). Asymptomatic reversible liver enzyme elevations (grade 3) were dose limiting in 2 of 5 patients at 450 mg once daily. At 250 mg b.i.d., 2 of 11 patients experienced dose-limiting toxicity (grade 3 liver enzyme elevation and gastrointestinal symptoms). Common toxicities included fatigue, diarrhea, nausea, vomiting, and abdominal pain (mainly grade ≤2). Among 45 patients, 22 (49%) achieved stable disease; 7 remained on treatment for >6 months. DCE-MRI of target lesions revealed effects in some patients at 200 and ≥400 mg once daily. CONCLUSION: Nintedanib is well tolerated by patients with advanced solid malignancies, with MTD defined as 250 mg b.i.d., and can induce changes in DCE-MRI. Disease stabilization >6 months was observed in 7 of 51 patients.

Synthesis of bioactive ß-TCP coatings with tailored physico-chemical properties on zirconia bioceramics.

The objective of this work was to develop a synthesis procedure for the deposition of ß-TCP coatings with tailored physico-chemical properties on zirconia bioceramics. The synthesis procedure involved two steps: (i) a rapid wet-chemical deposition of a biomimetic CaP coating and (ii) a subsequent post-deposition processing of the biomimetic CaP coating, which included a heat treatment between 800 and 1200 °C, followed by a short sonication in a water bath. By regulating the heating temperature the topography of the ß-TCP coatings could be controlled. The average surface roughness (Ra) ranged from 42 nm for the coating that was heated at 900 °C (TCP-900) to 630 nm for the TCP-1200 coating. Moreover, the heating temperature also affected the dissolution rate of the coatings in a physiological solution, their protein-adsorption capacity and their bioactivity in a simulated body fluid.

Phase I study of the angiogenesis inhibitor BIBF 1120 in patients with advanced solid tumors.

PURPOSE: BIBF 1120 is an oral, potent angiokinase inhibitor targeting receptors of the vascular endothelial growth factors, platelet-derived growth factors, and fibroblast growth factors. This phase I, accelerated titration study assessed the maximum tolerated dose, safety, pharmacokinetics, and pharmacodynamic effects of BIBF 1120. PATIENTS AND METHODS: Sixty-one patients with advanced cancers received BIBF 1120 in successive cohorts. Twenty-five received 50 to 450 mg once daily and 36 received 150 to 300 mg twice daily in 4-week treatment courses interspersed by 1 week of washout. Dynamic contrast-enhanced magnetic resonance imaging assessed antiangiogenic effect in 42 patients. RESULTS: Most frequent BIBF 1120-related adverse events were mostly mild to moderate (Common Toxicity Criteria grade 1-2) nausea (68.9%), vomiting (45.9%), and diarrhea (44.3%). The majority of dose-limiting adverse events of Common Toxicity Criteria grade 3 or 4 were reversible liver enzyme elevations. The maximum tolerated dose was 250 mg of BIBF 1120 for once and twice daily dosing. BIBF 1120 was absorbed moderately fast (t(max) = 1-3 hours at steady state), with no deviation from dose linearity and no decrease of exposure over time. The gMean terminal half-life was from 13 to 19 hours. One complete and two partial responses occurred in patients with renal cell cancer (n = 2) and colorectal cancer (n = 1). Dynamic contrast-enhanced magnetic resonance imaging showed a significant reduction in tumor blood flow in 55% of evaluable patients. CONCLUSIONS: BIBF 1120 dosed continuously displayed a favorable safety and pharmacokinetics profile, and first efficacy signals were observed. Twice daily dosing permitted increased drug exposure without additional toxicity. Two hundred milligrams BIBF 1120 twice daily is the recommended dose for phase II monotherapy studies.

An open-label dose-escalation study of BIBF 1120 in patients with relapsed or refractory multiple myeloma.

BACKGROUND: To determine the maximum tolerated dose (MTD) and dose-limiting toxicities (DLTs) of BIBF 1120, a triple angiokinase inhibitor administered once-daily in patients with advanced multiple myeloma. PATIENTS AND METHODS: This Phase I study included 17 patients. Planned dose escalations of BIBF 1120 were 100, 200, 250 and 300 mg. Safety and pharmacokinetic (PK) assessments were performed. RESULTS: Two DLTs (200 and 250 mg) occurred due to increased gamma-glutamyltransferase levels (CTC grade 3). The 250 mg dose was well tolerated; no dose escalation beyond 250 mg was made. The most common adverse events included diarrhoea, nausea and vomiting. No detectable deviation from dose linear PKs was observed. Regarding tumour control, two patients had stable disease for > or = 4 months. CONCLUSION: BIBF 1120 was safe and well tolerated up to 250 mg/day. The MTD was not reached.

Bortezomib, dexamethasone, and fibroblast growth factor receptor 3-specific tyrosine kinase inhibitor in t(4;14) myeloma.

PURPOSE: Novel drugs including targeted approaches have changed treatment paradigms for multiple myeloma (MM) and may also have therapeutic potential in the poor-prognosis t(4;14) subset; t(4;14) results in overexpressed and activated fibroblast growth factor receptor 3 (FGFR3). Blocking this receptor tyrosine kinase (RTK) induces apoptosis in t(4;14)+ MM cells and decreases adhesion to bone marrow stromal cells (BMSC). Using combinations of novel drugs, we investigated potential enhancement of single-agent activities within the tumor cells, targeting of the marrow micromilieu, or circumvention of drug resistance in t(4;14)+ MM. EXPERIMENTAL DESIGN: We tested effects on apoptosis and related signaling pathways in the t(4;14)+ MM subset, applying drug combinations including a FGFR3 tyrosine kinase inhibitor (RTKI), the proteasome inhibitor bortezomib, and dexamethasone. RESULTS: RTKI, bortezomib, and dexamethasone were active as single agents in t(4;14)+ MM. RTK inhibition triggered complementary proapoptotic pathways (e.g., decrease of Mcl-1, down-regulation of p44/42 mitogen-activated protein kinase, and activation of proapoptotic stress-activated protein/c-Jun NH(2)-terminal kinases). Synergistic or additive effects were found by combinations of RTKI with dexamethasone or bortezomib. In selected cases of t(4;14)+ MM, triple combinations were superior to dual combinations tested. Prevention from MM cell apoptosis by BMSC or exogenous interleukin-6 was circumvented by drug combinations. In t(4;14)+, N-ras-mutated NCI-H929 cells, resistance to RTKI was overcome by addition of dexamethasone. Notably, the combination of RTKI and dexamethasone showed additive proapoptotic effects in bortezomib-insensitive t(4;14)+ MM. CONCLUSIONS: Combining novel drugs in poor-prognosis t(4;14)+ MM should take into account at least bortezomib sensitivity and probably Ras mutational status.

Growth inhibition and induction of apoptosis in acute myeloid leukemia cells by new indolinone derivatives targeting fibroblast growth factor, platelet-derived growth factor, and vascular endothelial growth factor receptors.

In acute myeloid leukemia (AML), receptor tyrosine kinase ligands promote growth and survival and contribute to AML-associated marrow neoangiogenesis. We have tested simultaneous inhibition of vascular endothelial growth factor, fibroblast growth factor, and platelet-derived growth factor receptor signaling by novel indolinone derivatives using 14 myeloid, including 11 human leukemic, cell lines. Compounds inhibited colony formation of all cell lines in a dose-dependent fashion. Inhibitory concentrations for 50% of the colony formation/survival (IC50) for BIBF1000 were <100 nmol/L for 3 of 11,

Targeting receptor kinases by a novel indolinone derivative in multiple myeloma: abrogation of stroma-derived interleukin-6 secretion and induction of apoptosis in cytogenetically defined subgroups.

In multiple myeloma (MM), both vascular endothelial (VEGF) and basic fibroblast growth factor (bFGF) promote tumor growth and survival. We have used the novel indolinone BIBF 1000 to study effects of simultaneous inhibition of VEGF, FGF and transforming growth factor-beta on MM cells and their interactions with bone marrow stroma cells (BMSCs). Both, in the absence and presence of myeloma-stroma cell contacts, BIBF 1000 abrogated BMSC-derived secretion of interleukin-6 (IL-6). In addition, BIBF 1000 directly induced apoptosis in t(4;14)-positive cell lines as well as in CD138+ marrow cells from patients with t(4;14) myeloma. To a similar extent, BIBF 1000 induced apoptosis in MM.1S and MM.1R cells carrying the translocation t(14;16). In case of MM.1S and other dexamethasone-sensitive t(14;16) cell lines, BIBF 1000 and dexamethasone had additive proapoptotic effects. Induction of apoptosis by BIBF 1000 was associated with inhibition of the mitogen-activated protein kinases (MAPK) pathway in t(4;14) and inhibition of the phosphatidyl-inositol-3 kinase/AKT pathway in t(14;16) cells. Apoptotic effects did not occur in t(4;14)-or t(14;16)-positive MM cells carrying n- or k-Ras mutations. The data provide the rationale for clinical evaluation of this class of targeted kinase inhibitors in MM with focus on defined cytogenetic subgroups.