A first-in-man phase I study of R306465, a histone deacetylase (HDAC) inhibitor exploring pharmacokinetics (PK) and pharmacodynamics (PD) utilizing an electrochemiluminescent immunoassay in patients (p) with advanced tumours

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 3578-3578 ◽  
Author(s):  
P. C. Fong ◽  
S. Settatree ◽  
R. Sinha ◽  
A. Hardcastle ◽  
P. W. Hellemans ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Plasma Concentrations ◽  
Hdac Inhibitors ◽  
Fold Increase ◽  
Maximum Tolerated Dose ◽  
Transcriptional Level ◽  
Peripheral Blood Mononuclear ◽  
Fed State ◽  
Class 1 ◽  
Ecog Ps

3578 Background: R306565 is an aromatic hydroxamic acid with predominant inhibitory effects on Class 1 HDAC enzymes (with IC50 ∼10 nM). HDAC inhibitors (HDACi) affect gene expression at the transcriptional level, leading to cell cycle arrest and induction of apoptosis. Methods: P with solid tumours or lymphoma were given R306465 orally daily for 3 weeks (w) out of 4 in an escalating schedule. Objectives include safety, tolerability, PK (including food effect exploration), PD evaluation and circulating tumour cell (CTC) quantitation. Results: Four dose levels (100, 200, 300 and 400 mg) have been evaluated involving 15 p (7 male), age range 29–72 (median 59 y) and ECOG PS 0–2. A total of 37 cycles have been administered. Most common adverse events (AE) were Grade (G) 1–3 fatigue (87%), G1–2 nausea (66%), G1–2 vomiting (33%), G1–2 diarrhoea (40%), and G1–2 anorexia (40%). Dose limiting toxicity of G3 fatigue was seen in 1/6 p in the 400mg cohort. PK parameters were approximately dose proportional. Plasma concentrations increased in the fed state. PD effect of histone H3 acetylation (AcH3) in peripheral blood mononuclear cells (PBMC) was determined quantitatively with a novel validated electrochemiluminescent immunoassay developed in-house (applying Mesoscale Discovery technology). Although some interpatient variability exists, increased AcH3 was observed in 2/6 p in the 400 mg cohort, while the percentage rise in AcH3 was minimal for cohorts 1–3. Peak AcH3 achieved in 2 p dosed at 400 mg was approximately 5–10 fold increase over baseline. Using CellSearch technology for quantitation of CTCs, 8/14 p had detectable CTCs at baseline; the CTC trend will be presented. 4 p had stable disease (SD) for = 4 months. Conclusions: R306465 could be safely administered on a daily dosing schedule for 3 of 4 w up to 400 mg. Common toxicities seen were gastrointestinal and fatigue. Maximum tolerated dose has not been reached. PK suggests dose proportionality. Promising PD data showing increased acetylation in PBMC at 400 mg, further supports the utilization of the immunoassay platform in HDACi clinical trials. No significant financial relationships to disclose.

Phase I trial of panobinostat (P) and imatinib (IM) in patients with treatment-refractory gastrointestinal stromal tumors (GIST).

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. 10032-10032
Author(s):  
Sebastian Bauer ◽  
Ralf A. Hilger ◽  
Florian Grabellus ◽  
James Nagarajah ◽  
Mathias Hoiczyk ◽  
...  
Keyword(s):  
Phase I ◽  
Mononuclear Cells ◽  
Metabolic Response ◽  
Combined Treatment ◽  
Plasma Concentrations ◽  
Maximum Tolerated Dose ◽  
Clinical Activity ◽  
Peripheral Blood Mononuclear

10032^ Background: Panobinostat (LBH589; P) is a pan-deacetylase-inhibitor that has preclinical activity in combination with IM in GIST models in vitro and in vivo. Aim of this study was to determine the maximum tolerated dose (MTD) and dose-limiting toxicities (DLT) of escalating doses of P in combination with IM in patients with GIST who have failed IM and sunitinib treatment. Methods: This was a two-center phase I study using a 3+3 design with a prespecified expansion of the MTD cohort. IM was administered at a dose of 400mg qd. Following a 7 day run-in phase, escalating doses of P were added. The starting dose for P was 20 mg given as a three-times-per-week (MWF schedule) oral dose for 3 out of 4 weeks. Doses were increased by 10 mg if no dose limiting toxicities emerged. Blood samples were drawn for PK and biomarker assessments of IM, its main metabolite N-desmethyl-IM, and P using a validated RP-HPLC method. Acetylation of histone A3 was evaluated in peripheral blood mononuclear cells (PBMNC) as pharmacodynamic marker for P activity. Metabolic response using PET (EORTC-PET study criteria) was assessed on day 7 of IM run-in and after 3 weeks of combined treatment with IM and P. Results: In total 12 extensively pretreated (median 5 pretreatments) pts (4 f, 8 m; median age 56 y, 34-75 y) received study treatment at 2 dose levels (DL). 2 dose-limiting toxicities (grade 4 thrombocytopenia) occurred at DL 2 (30 mg). Most common AEs were thrombocytopenia, anemia, fatigue, nausea, emesis, diarrhea, creatinine elevation, abdominal cramping, and weight loss. DL 1 (20mg) was declared MTD, and 5 additional pts were enrolled at DL1. Analysis of P and IM PK revealed mean peak concentration of 14.8 +/- 9.5 ng/ml for P (20 mg). IM plasma concentrations with 400 mg once-daily administration were 2.8 ± 1.1 μg/mL at peak and 1.2 ± 0.4 μg/mL at trough. Histone A3 acetylation was demonstrated in PBMNC from pts treated at DL 1. 11 pts were evaluable for PET response: 1 had mPR, 7 had mSD and 3 had mPD. Longest treatment duration was 17 weeks (median: 6wks). Conclusions: P in combination with IM is moderately tolerated. Evidence of target inhibition at the MTD was associated with limited clinical activity in heavily pretreated pts with GIST.

Phase I Trial of Histone Deacetylase Inhibition by Valproic Acid Followed by the Topoisomerase II Inhibitor Epirubicin in Advanced Solid Tumors: A Clinical and Translational Study

2007 ◽  
Vol 25 (15) ◽  
pp. 1979-1985 ◽  
Author(s):  
Pamela Münster ◽  
Douglas Marchion ◽  
Elona Bicaku ◽  
Morgen Schmitt ◽  
Ji Hyun Lee ◽  
...  
Keyword(s):  
Valproic Acid ◽  
Histone Deacetylase ◽  
Topoisomerase Ii ◽  
Mononuclear Cells ◽  
Plasma Concentrations ◽  
Median Number ◽  
Maximum Tolerated Dose ◽  
Peripheral Blood Mononuclear ◽  
Histone Deacetylase Inhibition

Purpose To determine the safety, toxicity, and maximum-tolerated dose of a sequence-specific combination of the histone deacetylase inhibitor (HDACi), valproic acid (VPA), and epirubicin in solid tumor malignancies and to define the clinical feasibility of VPA as an HDACi. Patients and Methods Patients were treated with increasing doses of VPA (days 1 through 3) followed by epirubicin (day 3) in 3-week cycles. The study evaluated pharmacokinetic and pharmacodynamic end points, toxicities, and tumor response. Results Forty-eight patients were enrolled, and 44 received at least one cycle of therapy. Patients (median age, 54 years; range, 39 to 78 years) received the following doses of VPA: 15, 30, 45, 60, 75, 90, 100, 120, 140, and 160 mg/kg/d. Dose-limiting toxicities were somnolence (n = 1), confusion (n = 3), and febrile neutropenia (n = 1). No exacerbation of epirubicin-related toxicities was observed. Partial responses were seen across different tumor types in nine patients (22%), and stable disease/minor responses were seen in 16 patients (39%), despite a median number of three prior regimens (range, zero to 10 prior regimens). Patients received a median number of four treatment cycles (range, one to 10 cycles), and treatment was stopped after reaching maximal epirubicin doses rather than progression in 13 (32%) of 41 patients patients. Total and free VPA plasma concentrations increased linearly with dose and correlated with histone acetylation in peripheral-blood mononuclear cells. Conclusion The maximum-tolerated dose and recommended phase II dose was VPA 140 mg/kg/d for 48 hours followed by epirubicin 100 mg/m2. Sustained plasma concentrations of VPA exceeding those required for in vitro synergy were achieved with acceptable toxicity. Noteworthy antitumor activity was observed in heavily pretreated patients and historically anthracycline-resistant tumors.

scholarly journals Optimizing the Procedure to Manufacture Clinical-Grade NK Cells for Adoptive Immunotherapy

Cancers ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 577
Author(s):  
Adrián Fernández ◽  
Alfonso Navarro-Zapata ◽  
Adela Escudero ◽  
Nerea Matamala ◽  
Beatriz Ruz-Caracuel ◽  
...  
Keyword(s):  
Nk Cells ◽  
Cell Activation ◽  
Mononuclear Cells ◽  
Nk Cell ◽  
Fold Increase ◽  
Clinical Grade ◽  
Transcriptome Profile ◽  
Expansion Process ◽  
Peripheral Blood Mononuclear ◽  
Activation Status

Natural killer (NK) cells represent promising tools for cancer immunotherapy. We report the optimization of an NK cell activation–expansion process and its validation on clinical-scale. Methods: RPMI-1640, stem cell growth medium (SCGM), NK MACS and TexMACS were used as culture mediums. Activated and expanded NK cells (NKAE) were obtained by coculturing total peripheral blood mononuclear cells (PBMC) or CD45RA+ cells with irradiated K562mbIL15-41BBL or K562mbIL21-41BBL. Fold increase, NK cell purity, activation status, cytotoxicity and transcriptome profile were analyzed. Clinical-grade NKAE cells were manufactured in CliniMACS Prodigy. Results: NK MACS and TexMACs achieved the highest NK cell purity and lowest T cell contamination. Obtaining NKAE cells from CD45RA+ cells was feasible although PBMC yielded higher total cell numbers and NK cell purity than CD45RA+ cells. The highest fold expansion and NK purity were achieved by using PBMC and K562mbIL21-41BBL cells. However, no differences in activation and cytotoxicity were found when using either NK cell source or activating cell line. Transcriptome profile showed to be different between basal NK cells and NKAE cells expanded with K562mbIL21-41BBL or K562mbIL15-41BBL. Clinical-grade manufactured NKAE cells complied with the specifications from the Spanish Regulatory Agency. Conclusions: GMP-grade NK cells for clinical use can be obtained by using different starting cells and aAPC.

Exercise elevates plasma levels but not gene expression of IL-1β, IL-6, and TNF-α in blood mononuclear cells

2000 ◽  
Vol 89 (4) ◽  
pp. 1499-1504 ◽  
Author(s):  
Andrei I. Moldoveanu ◽  
Roy J. Shephard ◽  
Pang N. Shek
Keyword(s):  
Gene Expression ◽  
Oxygen Uptake ◽  
Mononuclear Cells ◽  
Plasma Concentrations ◽  
Peak Oxygen Uptake ◽  
Cytokine Gene Expression ◽  
Mrna Accumulation ◽  
Peripheral Blood Mononuclear ◽  
Tnf Α ◽  
Blood Mononuclear Cells

Physical activity induces a subclinical inflammatory response, mediated in part by leukocytes, and manifested by elevated concentrations of circulating proinflammatory cytokines, including interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α). However, the source of the cytokines that appear during exercise remains unknown. In this study, we examined exercise-induced changes in plasma cytokine concentrations and their corresponding mRNA expression in peripheral blood mononuclear cells. Ten healthy [peak oxygen uptake = 48.8 ± 6.5 (SD) ml · kg−1 · min−1] but untrained men [age = 25 ± 5 (SD) yr] undertook 3 h of exercise (cycling and inclined walking) at 60–65% peak oxygen uptake. Circulating leukocyte subset counts were elevated during and 2 h postexercise but returned to normal within 24 h. Plasma concentrations of IL-1β, IL-6, and TNF-α peaked at the end of exercise and remained elevated at 2 h (IL-6) and up to 24 h (IL-1β and TNF-α) postexercise. Cytokine gene expression in circulating mononuclear cells was measured by using the reverse transcriptase-polymerase chain reaction; mRNA accumulation did not change with exercise. In conclusion, mRNA accumulation of IL-1β, IL-6, and TNF-α in circulating mononuclear cells is not affected by 3 h of moderate endurance exercise and does not seem to account for the observed increases in plasma cytokines.

Phase 1/2 study of eprenetapopt (APR-246) in combination with pembrolizumab in patients with solid tumor malignancies.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. TPS3161-TPS3161
Author(s):  
Ecaterina Elena Dumbrava ◽  
Amit Mahipal ◽  
Xin Gao ◽  
Geoffrey Shapiro ◽  
Jason S. Starr ◽  
...  
Keyword(s):  
Solid Tumors ◽  
Mononuclear Cells ◽  
Phase 1 ◽  
Objective Response ◽  
Progression Free Survival ◽  
Maximum Tolerated Dose ◽  
Tumor Growth Inhibition ◽  
Advanced Solid Tumors ◽  
Peripheral Blood Mononuclear

TPS3161 Background: The p53 pathway has been implicated in antitumor immunity, including antigen presentation and T-cell proliferation. Loss of p53 function can increase resistance to immunotherapy across many tumor types. Eprenetapopt (eprenet) is a small molecule that stabilizes the folded structure of p53, resulting in activation of mutant p53 and stabilization of wild-type (WT) p53. It also targets the cellular redox homeostasis, resulting in induction of apoptosis in tumor cells. In vivo, mice carrying supernumerary copies of the TP53 gene harbor a pro-inflammatory tumor microenvironment, an effect recapitulated in TP53 normal-copy mice treated with eprenetapopt. Combining eprenetapopt and anti-PD1 or anti-CTLA4 therapy resulted in enhanced tumor growth inhibition and improved survival in TP53 WT mice inoculated with B16 melanoma and MC38 colon adenocarcinoma cells . Based on these results, we hypothesized that eprenet-induced p53 stabilization may augment response to immunotherapy. To test this hypothesis, we are conducting a phase 1b/2 study of eprenet in combination with pembrolizumab (eprenet+pembro) in pts with solid tumors. Methods: The primary objectives are to determine the maximum tolerated dose (MTD) and recommended phase 2 dose (RP2D) and to assess the safety and tolerability of eprenet+pembro in pts with advanced solid tumors. The secondary objectives are to estimate the anti-tumor activity and to describe the pharmacokinetics of the combination. Exploratory objectives include assessing predictive and pharmacodynamic markers of response. The study includes a safety lead-in with a 3+3 dose de-escalation design for pts with advanced solid tumors with known tumor TP53 mutation status ( TP53 WT is acceptable) (max 18 pts), followed by expansion cohorts in pts with NSCLC, gastric/GEJ and urothelial cancer (max 100 pts). In expansion, pts with urothelial and gastric cancers must be naïve to anti-PD-1/ L1 therapy. Eprenet is given IV once daily on Days 1–4 while pembro is administered on Day 3 of each 21-day cycle. The RP2D of eprenet+pembro is considered the dose at which ≤ 1 of 6 pts in a cohort has a dose-limiting toxicity (DLT). Primary endpoints are occurrence of DLTs, adverse events (AEs) and serious AEs with eprenet+pembro. Key secondary endpoints are best objective response, progression free survival and overall survival. Exploratory endpoints include gene mutations by next generation sequencing (including TP53), mRNA expression, multiplex immunohistochemistry and transcriptomics, multiplex flow cytometry on peripheral blood mononuclear cells and cytokines in serum. Continuous monitoring of toxicity will be conducted. The trial opened in May 2020 and is actively enrolling patients. Clinical trial information: NCT04383938.

Dose-escalation and pharmacodynamic study of topotecan in combination with cyclophosphamide in patients with refractory cancer.

1997 ◽  
Vol 15 (1) ◽  
pp. 148-157 ◽  
Author(s):  
J R Murren ◽  
S Anderson ◽  
J Fedele ◽  
G Pizzorno ◽  
D Belliveau ◽  
...  
Keyword(s):  
Dna Fragmentation ◽  
Mononuclear Cells ◽  
Day Treatment ◽  
Alkylating Agents ◽  
Plasma Concentrations ◽  
Fixed Dose ◽  
Sign Test ◽  
Published Data ◽  
Drug Induced ◽  
Peripheral Blood Mononuclear

PURPOSE Based on preclinical data that demonstrated synergy between alkylating agents and topoisomerase (topo) I poisons, we determined the maximum-tolerated dose (MTD) of topotecan, using a 5 day bolus schedule, that could be given in combination with a single, fixed dose of cyclophosphamide. Pharmacodynamics of this combination were explored by analyzing biochemical effects of treatment in peripheral-blood mononuclear cells (PBMCs). PATIENTS AND METHODS Patients with refractory cancer were treated with cyclophosphamide 600 mg/m2 on day 1, followed by topotecan given as a 30-minute infusion for 5 consecutive days. Cycles were repeated every 3 weeks. Once the MTD was defined, granulocyte colony-stimulating factor (G-CSF) was added to the regimen in an attempt to escalate further the dose of topotecan. Plasma concentrations of topotecan were determined during the first treatment cycle by high-performance liquid chromatography. PBMCs were sampled at baseline and throughout the 5-day treatment period for analysis of topo I protein concentrations and to determine drug-induced DNA fragmentation. RESULTS Twenty-six patients were treated with topotecan at doses that ranged from 0.5 mg/m2/d to 1.2 mg/ m2/d for a total of 74 cycles. Reversible neutropenia was dose-limiting, with mild to moderate suppression of the other blood-cell elements commonly occurring. Transfusions of RBCs and platelets were required in 24% and 7% of treatment cycles, respectively. The most prominent nonhematologic toxicities were fatigue and weight loss. Compared with previously published data in which topotecan was administered alone, cyclophosphamide did not appear to alter the pharmacokinetics of topotecan. Significant increases in topo I concentration were identified in PBMCs following the administration of cyclophosphamide on day 1 and there was a significant decrease in topo 1 during the 5-day course of treatment (P < .01, sign test). DNA fragmentation as a result of drug treatment was identified in 11 of 15 (73%) cycles analyzed. CONCLUSION For previously treated patients, the recommended dose of topotecan in this schedule is 0.75 mg/m2/d without growth factor support and 1.0 mg/ m2/d if it is administered with G-CSF. Biochemical changes in cells induced by exposure to camptothecins can be measured in vivo and these effects may have important implication in the design of combination therapies and the optimal scheduling of this class of agents.

Phase I clinical and pharmacologic study of eniluracil plus fluorouracil in patients with advanced cancer.

1998 ◽  
Vol 16 (4) ◽  
pp. 1450-1457 ◽  
Author(s):  
R L Schilsky ◽  
J Hohneker ◽  
M J Ratain ◽  
L Janisch ◽  
L Smetzer ◽  
...  
Keyword(s):  
Washout Period ◽  
Mononuclear Cells ◽  
Dihydropyrimidine Dehydrogenase ◽  
Maximum Tolerated Dose ◽  
Peripheral Blood Mononuclear ◽  
Period 2 ◽  
Daily Schedule ◽  
Starting Dose ◽  
Dose Levels

PURPOSE To determine the highest dose of fluorouracil (5-FU) that could be safely administered with Eniluracil (776C85; Glaxo Wellcome Inc, Research Triangle Park, NC), an inactivator of dihydropyrimidine dehydrogenase (DPD), on a daily schedule for 5 days, and to define the toxicities of the combination and the pharmacokinetics of 5-FU when administered with 776C85. PATIENTS AND METHODS Patients with advanced solid tumors refractory to standard therapy were enrolled at two institutions. The study consisted of three periods designed to evaluate the safety, pharmacokinetics, and pharmacodynamics of 776C85 alone (period 1); the effects of 776C85 on the pharmacokinetics of 5-FU (period 2); and the maximum-tolerated dose (MTD) of 5-FU, with or without leucovorin, that could be safely administered with 776C85 (period 3). Cohorts of at least three patients each received oral 776C85 alone at doses of 3.7 mg/m2/d, 18.5 mg/m2/d and 0.74 mg/m2/d. After a 14-day washout period, each patient then received 776C85 daily for 3 days, with a single intravenous (i.v.) bolus dose of 5-FU 10 mg/m2 on day 2. After a second washout period, patients were treated with 776C85 daily for 7 days and 5-FU i.v. bolus on days 2 through 6. The starting dose of 5-FU 10 mg/m2/d was escalated until the MTD was determined. After determination of the MTD of 5-FU given with 776C85, oral leucovorin 50 mg/d on days 2 through 6 was added to determine the MTD of 5-FU with leucovorin in the presence of 776C85. Near the completion of the study, additional cohorts of patients were treated with 776C85 at 50 mg/d and oral 5-FU with or without leucovorin. RESULTS Sixty-five patients were enrolled onto the study and 60 were assessable for toxicity and response. Bone marrow suppression was the primary and dose-limiting toxicity of this regimen. Other toxicities included diarrhea, mucositis, anemia, anorexia, nausea, vomiting, and fatigue. 776C85 suppressed DPD activity in peripheral-blood mononuclear cells (PBMCs) by at least 90% for at least 24 hours at all dose levels tested. In the presence of 776C85, 5-FU half-life was prolonged, clearance was reduced, and the drug displayed linear pharmacokinetics. Recommended doses for further testing on a daily for 5-day schedule are 776C85 10 mg/d with i.v. 5-FU 25 mg/m2/d; 776C85 10 mg/d with i.v. 5-FU 20 mg/m2/d plus leucovorin 50 mg/d; 776C85 50 mg/d with 5-FU given orally at 15 mg/m2/d with leucovorin at 50 mg/d. CONCLUSION 5-FU can be safely administered with 776C85; however, the MTDs are considerably lower than those conventionally used, caused, at least in part, by marked alterations in 5-FU plasma pharmacokinetics.

scholarly journals Intracellular and Plasma Trough Concentration and Pharmacogenetics of Telaprevir

2015 ◽  
Vol 18 (2) ◽  
pp. 171 ◽  
Author(s):  
Jessica Cusato ◽  
Sarah Allegra ◽  
Amedeo De Nicolò ◽  
Lucio Boglione ◽  
Giovanna Fatiguso ◽  
...  
Keyword(s):  
Triple Therapy ◽  
Mononuclear Cells ◽  
Linear Regression Analysis ◽  
Plasma Concentrations ◽  
Kinetic Studies ◽  
Pharmacokinetic Data ◽  
Nucleotide Polymorphisms ◽  
Allelic Discrimination ◽  
Peripheral Blood Mononuclear ◽  
Drug Kinetic

PURPOSE: Triple therapy for HCV-1 infection consists in boceprevir or telaprevir, ribavirin and PEG-interferon. Telaprevir is a P-glycoprotein substrate and it is metabolized by CYP3A4/5. No data have been published on intracellular penetration of telaprevir. We determined peripheral blood mononuclear cells (PBMCs) and trough plasma S and R telaprevir isomers concentrations; moreover, we evaluated the influence of some single nucleotide polymorphisms (SNPs) on these pharmacokinetic data after 1 month of triple therapy in humans. METHODS: Plasma and intracellular telaprevir concentrations were determined at the end of dosing interval (Ctrough) using ULPC-MS/MS validated methods; allelic discrimination was performed through real-time PCR. RESULTS: Median telaprevir Ctrough plasma concentrations were 2579 ng/mL and 2233 ng/mL for the pharmacologically more active S, and R, enantiomers, respectively, with median S/R plasma ratio of 1.11. In PBMC, the medians were 6863 ng/mL and 1096 ng/mL for S and R, respectively, with median S/R being 5.73. The PBMC:plasma ratio for S was 2.59 for R. Plasma ribavirin concentrations were directly correlated with plasma S-telaprevir concentrations. In linear regression analysis, only CYP24A1_rs2585428 SNP (p=0.003) and body mass index (p=0.038) were able to predict S-telaprevir PBMC concentrations. CONCLUSIONS: Our preliminary data could increase the understanding of mechanisms underlying telaprevir intracellular and plasma exposure, suggesting the implementation of pharmacogenetics in these drug kinetic studies. This article is open to POST-PUBLICATION REVIEW. Registered readers (see “For Readers”) may comment by clicking on ABSTRACT on the issue’s contents page.

Increased Activity of Prothrombinase Fgl-2 in Peripheral Blood Mononuclear Cells of Patients with B-Cell Lymphoma.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2665-2665
Author(s):  
Ofir Wolach ◽  
Esther Rabizadeh ◽  
Doron Lederfein ◽  
Natalia Binkovski ◽  
Pia Raanani ◽  
...  
Keyword(s):  
B Cell ◽  
Peripheral Blood Mononuclear Cells ◽  
Thrombin Generation ◽  
Mononuclear Cells ◽  
Cell Lymphoma ◽  
B Cell Lymphoma ◽  
Fold Increase ◽  
Lymphoproliferative Disorders ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

Abstract Abstract 2665 Background: Hematologic and solid tumors are associated with hypercoagulability the reason for which has not been delineated. Prothrombinase named fibrinogen-like protein 2 (FGL-2) is a 70 kD transmembrane protein that was found to have a quality of a serine protease capable of directly cleaving prothrombin to thrombin. FGL-2 is synthesized by monocytes, T-lymphocytes and endothelial cells. FGL-2 protein and its mRNA have been previously found within different tumor cells. Aim: To study the role of FGL-2 in patients with lymphoproliferative disorders. Our hypothesis is that upregulation of FGL-2 activity in patients with B-cell malignancies may contribute to tumorigenesis via generation of thrombin leading to increased angiogenesis and spread/metastasis of malignant cells. Methods: Thrombin generation reflecting FGL-2 activity was measured in homogenized peripheral blood mononuclear cells (PBMC) from 29 patients with active lymphoproliferative disorders and 107 normal controls. Informed consent was obtained from every participant. PBMC extracts were incubated with an equal volume of human prothrombin (final concentration10 μM) for 30 min at 37 °C. Thrombin generation was measured at 405 nm using an automated plate reader after addition of chromogen S-2238. The thrombin activity of each sample was calculated by comparison with absorbance curve generated by known concentrations of human thrombin. FGL-2 was immunoprecipitated (IP) from PBMC with an anti-FGL-2 antibody and EZview Red Protein A Affinity Gel (sigma # P6486). The activity of IP FGL-2 was measured by thrombin generation assay. The expression of FGL-2 was analyzed in HUVEC and PBMC in the presence or absence of IF-γ at 20 ng/ml. Total RNA was isolated using RNAqueous™ (Ambion #AM1912) and RT-PCR, analysis was performed using Rotor-gene RG-3000 (Corbett). The difference in cycle time (ΔCT) was measured by comparing FGL-2 gene with ABL-1 gene (house keeping gene). The relative quantification was calculated by the formula RQ= 2−ΔCT. HUVEC were transfected with 0.5 μM SiRNA synthesized complementary to FGL-2 (Target SiRNA) using Dharmacon ON-TARGET plus SMART pool reagent (Thermo Fisher Scientific) according to manufacturer instructions. FGL-2 expression following addition of target SiRNA was compared to that obtained following addition of non-specific (non-target) SiRNA. Student's t-test was used for all comparisons. Results: As shown in the table, almost 3-fold increase in FGL-2 activity in PBMC was observed among patients with active B-cell lymphoma, either aggressive or indolent, as compared to that of healthy controls (p<0.0001 for all comparisons). No difference in FGL-2 activity in PBMC between patients with aggressive and indolent lymphoma was observed (p=0.68). Three-fold increase in thrombin generation was obtained in PBMC from the patients following IP, similarly to that observed in non-IP PBMC. A significant increase in mRNA of FGL-2 was found in either PBMC of lymphoma patient or HUVEC treated with IF-γ. In HUVEC the increase in FGL-2 mRNA was inhibited by 80% following treatment with specific SiRNA. Conclusions: Disclosures: No relevant conflicts of interest to declare.

Phase Ib dose escalation study of oral quisinostat, a histone deacetylase inhibitor, in combination with bortezomib and dexamethasone for patients with relapsed multiple myeloma.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. 8530-8530
Author(s):  
Xavier Leleu ◽  
Cyrille Touzeau ◽  
Lotfi Benboubker ◽  
Thierry Facon ◽  
Martine Delain ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Overall Response Rate ◽  
Mononuclear Cells ◽  
Response Rate ◽  
Complete Response ◽  
Maximum Tolerated Dose ◽  
Dose Escalation Study ◽  
Peripheral Blood Mononuclear ◽  
Overall Response ◽  
Aggresome Formation

8530 Background: Aggresome formation is a mechanism of resistance to agents (e.g., bortezomib) which block proteasome activity. HDACi (e.g., quisinostat) prevents aggresome formation by deacetylation of tubulin that allows the transport of unfolded proteins to lysosomes for degradation. Methods: Patients received quisinostat (Q) at escalated doses (6, 8, 10 and 12 mg) on days 1, 3, and 5 weekly, subcutaneous VELCADE (V) at 1.3 mg/m2on days 1, 4, 8, and 11 of a 3-week cycle, and oral dexamethasone (D) at 20 mg on the day of and the day after VELCADE dosing. The primary endpoint was the maximum tolerated dose (MTD) of Q in the combination (Q+V+D). The secondary endpoints included safety, overall response rate, and pharmacodynamic biomarkers. Results: Eighteen patients (3, 3, 6, and 6 in increasing Q doses) were enrolled: 56% male; median age = 69 (range 50-82) years; multiple myeloma stage: IA = 11% and IIIA = 89%; prior lines of therapy: 1 = 100%, 2 = 55.6%, and 3 = 11.1%; prior VELCADE treatment = 50%. At the highest dose (12 mg) 2 patients had dose-limiting toxicity, 1 with QTc prolongation and 1 with atrial fibrillation. The MTD was established at the 10 mg Q for the Q+V+D regimen. The most common adverse events (≥ 10% of patients) were diarrhea (39%), asthenia (33%), peripheral oedema (22%), nausea (17%), thrombocytopenia (17%), alopecia (11%), constipation (11%), and vomiting (11%); most were grade 2 or lower in toxicity. To date, 13 patients have discontinued treatment, of which 5 completed 11 cycles of treatment. The overall response rate was 87.5% (14/16, 95% CI: 61.7% to 98.5%), including 1 complete response, 2 very good partial response, and 11 partial responses. Most patients (9/11) showed a decrease in number of circulating multiple myeloma cells after 1 cycle. Two of 5 patients showed an increase in acetylated histone 3 from baseline as measured in peripheral blood mononuclear cells. Conclusions: The MTD is 10 mg quisinostat in combination with VELCADE and dexamethasone. The combination is active in the treatment of relapsed multiple myeloma and has an acceptable safety profile. Clinical trial information: NCT01464112.

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