scholarly journals 165 Generating enhanced tumor infiltrating lymphocytes through microfluidic cell squeezing

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A175-A175
Author(s):  
Devin Bridgen ◽  
Arindam Bhattacharjee ◽  
Colin Thalhofer ◽  
Ryan Montler ◽  
Andrew Weinberg ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Tumor Infiltrating Lymphocytes ◽  
Costimulatory Molecules ◽  
Clinical Activity ◽  
High Dose ◽  
Concomitant Treatment ◽  
High Concentration ◽  
High Viability ◽  
Membrane Bound

BackgroundTumor Infiltrating Lymphocyte (TIL) therapies have shown significant solid tumor activity in patients, but current TIL compositions require patient lymphodepletion and high dose IL-2 after cell infusion to support clinical activity. Removing this requirement through ex vivo engineering of the TIL product with mRNA could enhance potency, expand the potential patient population, and potentially allow for repeat dosing and concomitant treatment with checkpoint therapies.MethodsTo transiently overexpress both membrane-bound cytokines and costimulatory molecules, we used microfluidic cell squeezing (Cell Squeeze®) to deliver mRNA directly to the cytosol of expanded tumor reactive CD8 human TILs (AGX-148). After mRNA delivery, the TILs were cultured in media with varying levels of exogenous IL-2 and characterized by flow cytometry.ResultsWe demonstrated that multiple mRNA constructs delivered simultaneously by microfluidic cell squeezing to human TILs are highly expressed (>80% of cells) for multiple days while maintaining high viability (>80%) in vitro. Membrane bound cytokines are able to support cell expansion in the absence of exogenous IL-2 at rates comparable to control cells incubated with a high concentration of IL-2 for up to 3 days. Furthermore, we have identified a membrane-bound cytokine that alters the TIL phenotype as quantified by multiple markers, including increased L-selectin (CD62L), which is an indicator of central memory T cells.ConclusionsThrough microfluidic cell squeeze delivery of mRNAs, we have created enhanced TILs with high levels of membrane-bound cytokines and/or costimulatory molecules in vitro. These cells are able to proliferate without exogenous IL-2 and have an improved phenotype.

scholarly journals In Vitro and Ex Vivo Activities of Minocycline and EDTA against Microorganisms Embedded in Biofilm on Catheter Surfaces

2003 ◽  
Vol 47 (11) ◽  
pp. 3580-3585 ◽  
Author(s):  
Issam Raad ◽  
Ioannis Chatzinikolaou ◽  
Gassan Chaiban ◽  
Hend Hanna ◽  
Ray Hachem ◽  
...  
Keyword(s):  
Low Dose ◽  
Ex Vivo ◽  
Bacterial Colonization ◽  
Rabbit Model ◽  
Confocal Laser ◽  
High Dose ◽  
High Concentration ◽  
Ex Vivo Model ◽  
Highly Active

ABSTRACT Minocycline-EDTA (M-EDTA) flush solution has been shown to prevent catheter-related infection and colonization in a rabbit model and in hemodialysis patients. We undertook this study in order to determine the activities of M-EDTA against organisms embedded in fresh biofilm (in vitro) and mature biofilm (ex vivo). For the experiment with the in vitro model, a modified Robbin’s device (MRD) was used whereby 25 catheter segments were flushed for 18 h with 106 CFU of biofilm-producing Staphylococcus epidermidis, Staphyloccocus aureus, and Candida albicans per ml. Subsequently, each of the catheter segments was incubated in one of the following solutions: (i) streptokinase, (ii) heparin, (iii) broth alone, (iv) vancomycin, (v) vancomycin-heparin, (vi) EDTA, (vii) minocycline (high-dose alternating with low-dose), or (viii) M-EDTA (low-dose minocycline alternating with high-dose minocycline were used to study the additive and synergistic activities of M-EDTA). All segments were cultured quantitatively by scrape sonication. For the experiment with the ex vivo model, 54 catheter tip segments removed from patients and colonized with bacterial organisms by roll plate were longitudinally cut into two equal segments and exposed to either saline, heparin, EDTA, or M-EDTA (with high-dose minocycline). Subsequently, all segments were examined by confocal laser electron microscopy. In the in vitro MRD model, M-EDTA (with a low concentration of minocycline) was significantly more effective than any other agent in reducing colonization of S. epidermidis, S. aureus, and C. albicans (P < 0.01). M-EDTA (with a high concentration of minocycline) eradicated all staphylococcal and C. albicans organisms embedded in the biofilm. In the ex vivo model, M-EDTA (with a high concentration of minocycline) reduced bacterial colonization more frequently than EDTA or heparin (P < 0.01). We concluded that M-EDTA is highly active in eradicating microorganisms embedded in fresh and mature biofilm adhering to catheter surfaces.

A phase I study (E011-MEL) of a TriMix-based mRNA immunotherapy (ECI-006) in resected melanoma patients: Analysis of safety and immunogenicity.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. 2641-2641
Author(s):  
ANA Maria Arance Fernandez ◽  
Jean-Francois Baurain ◽  
Christof Vulsteke ◽  
Annemie Rutten ◽  
Ainara Soria ◽  
...  
Keyword(s):  
Adverse Events ◽  
Ex Vivo ◽  
Intracellular Cytokine Staining ◽  
Cell Receptor ◽  
Immune Monitoring ◽  
Clinical Activity ◽  
High Dose ◽  
Serious Adverse Events ◽  
Melanoma Patients

2641 Background: ECI-006 is a combination of TriMix (mRNAs encoding for dendritic cell [DC] activating molecules [CD40L, CD70 and caTLR4]), and mRNAs encoding for melanoma-specific tumor-associated antigens (TAAs): tyrosinase, gp100, MAGE-A3, MAGE-C2, and PRAME. DCs transfected ex vivo with TriMix and TAAs mRNAs showed significant clinical activity in combination with ipilimumab in metastatic melanoma without increasing toxicity. This study aims to assess the safety and immunogenicity of ECI-006 vaccine administered intranodally (i.n.) in an adjuvant setting for patients with resected melanoma. Methods: Twenty patients who underwent resection of stage IIc/III/IV cutaneous melanoma received 5 administrations of ECI-006 (either 600 µg or 1800 µg [n = 10, each]) injected i.n. on Day 1 and after 2, 4, 6 and 14 weeks. Treatment-emergent adverse events (TEAEs) were graded using CTCAE version 4.0.3. Blood samples for immune monitoring (ELISPOT and intracellular cytokine staining [ICS]) were collected pre-dose and at weeks 4, 7, 14 and 15. Results: Nineteen patients completed the treatment. One patient in the low dose group discontinued the study after 4 doses due to disease relapse. Administration of ECI-006 was well tolerated. No serious adverse events or TEAEs Grade 3 or higher were reported. Of all TEAEs, myalgia and fatigue were the most reported in 3 (15%) and 5 (25%) patients, respectively. ELISPOT and ICS were performed on T cells pre-stimulated in vitro for 10-12 days, using a previously in-house validated protocol. Vaccine-induced immune responses according to predefined criteria were detected in 4/10 and 3/9 patients treated with the low and high dose, respectively. Samples from these patients are currently being subjected to T-cell receptor repertoire analysis. Conclusions: Among patients undergoing resection of stage IIc/III/IV melanoma, i.n. administration of ECI-006 at 600 or 1800 µg was generally well tolerated. ECI-006 demonstrated to be immunogenic in a proportion of patients. These results warrant further development of ECI-006 in combination with anti-PD-1 therapy in melanoma patients. Clinical trial information: NCT03394937.

Significance of Platelet β-Adrenoceptors for Platelet Responses In Vivo and In Vitro

1992 ◽  
Vol 68 (06) ◽  
pp. 687-693 ◽  
Author(s):  
P T Larsson ◽  
N H Wallén ◽  
A Martinsson ◽  
N Egberg ◽  
P Hjemdahl
Keyword(s):  
Ex Vivo ◽  
High Dose ◽  
Platelet Aggregability ◽  
Adrenoceptor Agonist ◽  
Dose Infusion ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Factor Antigen

SummaryThe significance of platelet β-adrenoceptors for platelet responses to adrenergic stimuli in vivo and in vitro was studied in healthy volunteers. Low dose infusion of the β-adrenoceptor agonist isoprenaline decreased platelet aggregability in vivo as measured by ex vivo filtragometry. Infusion of adrenaline, a mixed α- and β-adrenoceptor agonist, increased platelet aggregability in vivo markedly, as measured by ex vivo filtragometry and plasma β-thromboglobulin levels. Adrenaline levels were 3–4 nM in venous plasma during infusion. Both adrenaline and high dose isoprenaline elevated plasma von Willebrand factor antigen levels β-Blockade by propranolol did not alter our measures of platelet aggregability at rest or during adrenaline infusions, but inhibited adrenaline-induced increases in vWf:ag. In a model using filtragometry to assess platelet aggregability in whole blood in vitro, propranolol enhanced the proaggregatory actions of 5 nM, but not of 10 nM adrenaline. The present data suggest that β-adrenoceptor stimulation can inhibit platelet function in vivo but that effects of adrenaline at high physiological concentrations are dominated by an α-adrenoceptor mediated proaggregatory action.

EXTH-48. NOVEL COMBINATION THERAPY IN PRECLINICAL GLIOMA MODELS USING THE CELL CYCLE STABILIZING COMPOUND ARGYRIN F AND CHECKPOINT INHIBITION

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi174-vi174
Author(s):  
Bianca Walter ◽  
Denis Canjuga ◽  
Simge G Yuez ◽  
Michael Ghosh ◽  
Przemyslaw Bozko ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Ex Vivo ◽  
Tumor Infiltrating Lymphocytes ◽  
Clonogenic Survival ◽  
Autologous Tumor ◽  
Cycle Arrest ◽  
Resected Tissue ◽  
Infiltrating Lymphocytes

Abstract Glioblastoma are incurable aggressive tumors and remain a therapeutic challenge. Glioblastoma frequently harbor alterations in the retinoblastoma pathway with subsequent cell cycle abnormalities. Here, we aimed to investigate the anti-glioma activity of the cell cycle-stabilizing compound Argyrin F and its potential treatment-induced vulnerabilities to exploit possibilities for novel combination therapies. We investigated cell viability, clonogenic survival, cell cycle status and immunoblots of human and murine glioma cells treated with Argyrin F. Moreover, we established an ex vivo glioma model using residual freshly resected tissue from patients, i.e. patient-derived microtumors (PDMs). Additionally, we extracted autologous tumor infiltrating lymphocytes (TILs) to perform co-culturing experiments. We performed mass spectrometry-based immunopeptidomics and used the orthotopic syngeneic SMA560/VM/Dk glioma mouse model. Argyrin F displayed anti-glioma efficacy in glioma cell lines in vitro and in PDM models ex vivo. Moreover, Argyrin F treatment induced cell cycle arrest, reduced clonogenic survival in vitro and prolonged survival in vivo. Argyrin F-treated SMA560 glioma displayed 4.6-fold more glioma-infiltrating CD8+ T cells. We discovered a distinctive treatment-induced immunopeptidome. Combination of Argyrin F plus PD-1 antibody increased cellular toxicity in PDM/TILs co-cultures ex vivo and prolonged overall survival compared with monotherapies in vivo. We conclude that our experimental data suggest a novel combination of Argyrin F plus PD-1 blockade and its clinical translation.

CD138 Regulates Myeloma Cell Survival, Proliferation, BM Engraftment and Tumor Organization

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2974-2974
Author(s):  
David R Fooksman ◽  
Amitabha Mazumder ◽  
Mark McCarron
Keyword(s):  
Plasma Cells ◽  
Conflicts Of Interest ◽  
Ex Vivo ◽  
Early Stage ◽  
Serum Starvation ◽  
High Dose ◽  
Rapid Reduction ◽  
Myeloma Cells

Abstract Multiple myeloma is the 2nd most common blood cancer in adults with a median survival time of 5 years despite high-dose chemotherapy and bone marrow transplantation interventions. Syndecan-1 or CD138, is a heparan-sulfate coated glycoprotein, which is highly expressed on the surface of plasma cells and myeloma cells, important for adhesion and accumulating survival signals. Expression of CD138 is heterogeneous in myeloma tumors, in vivo and in vitro leading some to speculate it may distinguish stem-like subpopulations. While this role is highly disputed, we investigated the effect of CD138 expression on tumor pathology in vivo. To characterize CD138neg and CD138high subpopulations, we used GFP+ Vk*myc myeloma model from Leif Bergsagel, which develops myeloma tumors in BM and spleen of C57Bl/6 mice. We found CD138high populations were more proliferative in vivo based on EdU incorporation experiments. We transferred equal numbers of sorted subpopulations into hosts and found that CD138high cells generated larger tumors in the BM than CD138neg cells after 12 weeks. Analysis of these tumor-bearing mice revealed that all tumors contained both subpopulations, indicating that these two subsets are hierarchically equivalent. We find that in mice with small tumors, the majority of cells (80% or more) are CD138high cells, while in large tumors, the level drops (to 30-50% of tumor) with higher composition of CD138neg cells. We also find lower CD138 levels on myeloma cells found in the blood compared to BM. Using intravital two-photon time-lapse imaging in the tibial BM, we find that tumor cells from smaller, early stage tumors are physically arrested within the BM parenchyma, while in larger, more advanced tumors, myeloma cells are more motile and active. CD138neg cells were more apoptotic based on ex vivo Annexin V staining following serum starvation. Interestingly, serum starvation led to rapid reduction in CD138 surface expression. Taken together, we propose a model where CD138 expression regulates localization and survival in the BM niche, but is downregulated from the plasma membrane when tumor size outgrows the necessary resources, allowing myeloma cells to migrate and metastasize to distant new locations. Disclosures No relevant conflicts of interest to declare.

scholarly journals Ex Vivo Evaluation of the Effect of Plasma-Derived Factor VIII/Von Willebrand Factor in Patients with Severe Hemophilia_A on Prophylaxis with Emicizumab By Thrombin Generation Assay

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4233-4233
Author(s):  
Maria-Isabel Bravo ◽  
Aida Raventós ◽  
Alba Pérez ◽  
Elena G Arias-Salgado ◽  
María Teresa Alvarez Román ◽  
...  
Keyword(s):  
Von Willebrand Factor ◽  
Research Funding ◽  
Thrombin Generation ◽  
Ex Vivo ◽  
Concomitant Treatment ◽  
Healthy Controls ◽  
Normal Range ◽  
Von Willebrand ◽  
Willebrand Factor

Abstract Introduction: Hemophilia A (HA) patients under emicizumab prophylaxis treatment may require the concomitant use of procoagulant factors for breakthrough bleedings or immune tolerance induction. Thromboembolic events have been described with the concomitant use of emicizumab and activated prothrombin complex concentrate (aPCC), but not with recombinant activated factor VII (rFVIIa). Previous studies showed that the in vitro combination of emicizumab and plasma-derived Factor VIII/Von Willebrand Factor (pdFVIII/VWF) had a non-additive effect on thrombin generation (TG)(Bravo M-I, et al J Thromb Haemost. 2020;18:1934-39). The aim of this study was to evaluate the TG resulting from ex vivo combination of plasma samples from HA patients treated with emicizumab, with a pdFVIII/VWF concentrate (Fanhdi ®, Grifols). Methods: Twelve adult patients with severe HA without inhibitors on prophylaxis with emicizumab and nine healthy controls were included in the study. Blood samples were drawn in citrate plus corn trypsin inhibitor tubes. Then, platelet poor plasma (PPP) was collected for the TG assay, which measures the whole kinetics of TG. Thrombin peak (TP) and endogenous thrombin potential (ETP) were calculated using calibrated automated thrombogram (Thrombinoscope ™ software, Stago) after in vitro activation of coagulation by trigger solution, PPP Reagent LOW TM (4 μM phospholipids/1 pM tissue factor), fluorogenic substrate and CaCl 2 (FLUKAkit TM) reagents (Diagnostica Stago). Fluorescence was read in a Fluoroskan Ascent reader (Thermo) equipped with a 390/460 filter set. Samples were spiked with increasing concentrations of pdFVIII/VWF (10 to 400 IU/dL), rFVIIa (0.9 µg/mL) or aPCC (0.5 U/mL). Results: TG from healthy control samples was measured to establish TP and ETP normal ranges. TP and ETP results obtained from HA plasma with emicizumab were lower than in healthy controls. The addition of pdFVIII/VWF as of 25 IU/kg (prophylaxis dose in HA w/o inhibitors) to samples from HA patients concomitantly treated with emicizumab restored TP and ETP levels within healthy controls normal range (Table 1). Increasing ex vivo concentrations of pdFVIII/VWF maintained TP and ETP similar to healthy controls. The highest concentration of concomitant treatment with pdFVIII/VWF (200 IU/kg) and emicizumab did not result in excessive TP and, importantly, ETP levels were always within the normal range. The combination with the bypassing agent rFVIIa moderately increased TP and ETP values up to normal range. However, when HA plasma was spiked with aPCC in the presence of emicizumab, TP and ETP dramatically increased above normal range resulting in a synergistic procoagulant profile. Conclusions: The concomitant use of pdFVIII/VWF in patients with prophylaxis with emicizumab did not trigger a multiplying effect on TG. These results were aligned with previous in vitro data and suggested the low risk of overdose and thrombotic events of concomitant treatment emicizumab with the pdFVIII/VWF concentrate in HA patients. Figure 1 Figure 1. Disclosures Bravo: Grifols: Current Employment, Other: Grifols is a manufacturer of the pdFVIII/VWF concentrate, Fanhdi®. Raventós: Grifols: Current Employment, Other: Grifols is a manufacturer of the pdFVIII/VWF concentrate, Fanhdi®. Pérez: Grifols: Current Employment, Other: Grifols is a manufacturer of the pdFVIII/VWF concentrate, Fanhdi®. Alvarez Román: Grifols: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria, Research Funding; Novo-Nordisk: Consultancy, Honoraria, Research Funding; Sobi: Consultancy, Honoraria, Research Funding; Octapharma: Consultancy, Honoraria, Research Funding; Bayer: Consultancy, Honoraria, Research Funding; CSL-Behring: Consultancy, Honoraria, Research Funding; Biomarin: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding. Butta: CSL-Behring: Research Funding; Roche: Speakers Bureau; Takeda: Research Funding, Speakers Bureau; Novo-Nordisk: Speakers Bureau. Jiménez-Yuste: Bayer: Consultancy, Honoraria, Research Funding; F. Hoffmann-La Roche Ltd: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria, Research Funding; CSL Behring: Consultancy, Honoraria, Research Funding; BioMarin: Consultancy; Sobi: Consultancy, Honoraria, Research Funding; Octapharma: Consultancy, Honoraria, Research Funding; Sanofi: Consultancy, Honoraria, Research Funding; NovoNordisk: Consultancy, Honoraria, Research Funding; Grifols: Consultancy, Honoraria, Research Funding. Costa: Grifols: Current Employment, Other: Grifols is a manufacturer of the pdFVIII/VWF concentrate, Fanhdi®. Willis: Grifols: Current Employment, Other: Grifols is a manufacturer of the pdFVIII/VWF concentrate, Fanhdi®.

scholarly journals Cell-autonomous FLT3L shedding via ADAM10 mediates conventional dendritic cell development in mouse spleen

2019 ◽  
Vol 116 (29) ◽  
pp. 14714-14723 ◽  
Author(s):  
Kohei Fujita ◽  
Svetoslav Chakarov ◽  
Tetsuro Kobayashi ◽  
Keiko Sakamoto ◽  
Benjamin Voisin ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Embryonic Fibroblasts ◽  
Major Pathway ◽  
Peripheral Tissues ◽  
A Cell ◽  
Membrane Bound ◽  
A Disintegrin And Metalloproteinase ◽  
Ex Vivo Culture ◽  
Culture Supernatants

Conventional dendritic cells (cDCs) derive from bone marrow (BM) precursors that undergo cascades of developmental programs to terminally differentiate in peripheral tissues. Pre-cDC1s and pre-cDC2s commit in the BM to each differentiate into CD8α+/CD103+ cDC1s and CD11b+ cDC2s, respectively. Although both cDCs rely on the cytokine FLT3L during development, mechanisms that ensure cDC accessibility to FLT3L have yet to be elucidated. Here, we generated mice that lacked a disintegrin and metalloproteinase (ADAM) 10 in DCs (Itgax-cre × Adam10-fl/fl; ADAM10∆DC) and found that ADAM10 deletion markedly impacted splenic cDC2 development. Pre-cDC2s accumulated in the spleen with transcriptomic alterations that reflected their inability to differentiate and exhibited abrupt failure to survive as terminally differentiated cDC2s. Induced ADAM10 ablation also led to the reduction of terminally differentiated cDC2s, and restoration of Notch signaling, a major pathway downstream of ADAM10, only modestly rescued them. ADAM10∆DC BM failed to generate cDC2s in BM chimeric mice with or without cotransferred ADAM10-sufficient BM, indicating that cDC2 development required cell-autonomous ADAM10. We determined cDC2s to be sources of soluble FLT3L, as supported by decreased serum FLT3L concentration and the retention of membrane-bound FLT3L on cDC2 surfaces in ADAM10∆DC mice, and by demonstrating the release of soluble FLT3L by cDC2 in ex vivo culture supernatants. Through in vitro studies utilizing murine embryonic fibroblasts, we determined FLT3L to be a substrate for ADAM10. These data collectively reveal cDC2s as FLT3L sources and highlight a cell-autonomous mechanism that may enhance FLT3L accessibility for cDC2 development and survival.

Phase I study of OKT3 x hu3F8 bispecific antibody (GD2Bi) armed T cells (GD2BATs) in GD2-positive tumors.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. 2533-2533 ◽  
Author(s):  
Maxim Yankelevich ◽  
Shakeel Modak ◽  
Roland Chu ◽  
Daniel W. Lee ◽  
Archana Thakur ◽  
...  
Keyword(s):  
T Cells ◽  
Phase I ◽  
Phase I Study ◽  
Residual Disease ◽  
Ex Vivo ◽  
Autologous Blood ◽  
Maximum Tolerated Dose ◽  
Clinical Activity ◽  
Minor Response

2533 Background: With the proven success of anti-GD2 monoclonal antibodies in eradicating minimal residual disease in neuroblastoma (NB), exploiting antibody based anti-GD2 in T cell mediated strategies has potential to combat higher disease burden and improve patient outcome. We hypothesized that arming of ex vivo expanded and activated, autologous, blood derived T cells (ATC) with chemically heteroconjugated GD2Bi should redirect them to target NB. In vitro, ATC coated (armed) with 50 ng/106 cells of GD2Bi exhibited specific killing of NB and osteosarcoma (OS) cell lines. Methods: In this phase I study (NCT02173093), patients with GD2-positive tumors received 8, biweekly infusions of GD2BATs + daily low-dose IL-2 and biweekly granulocyte-macrophage colony stimulating factor (GM-CSF). The study followed the standard 3+3 design with dose levels of 40, 80, and 160 x 106 GD2BATs/kg/infusion. Results: Twelve patients (NB = 7, OS = 3, Desmoplastic Small Round Cell Tumor = 2) were enrolled from 11/2013 to 12/2017 and 9 completed therapy. Adequate ATCs could not be grown in one patient and two patients did not complete 8 infusions because of rapid disease progression. Infusions were given in outpatient settings. All patients developed a mild, dose-independent and manageable form of cytokine release syndrome with grades 2-3 fevers/chills, headaches and occasional hypotension for up to 48 hours after infusion. No patients developed significant pain. Maximum tolerated dose was not reached. Evidence of activity was seen in several patients including one patient with OS who had a PET response, one patient with NB who had complete bone marrow response (this patient had remained progression free for 2.5 years after completion of infusions), and another NB patient who had a minor response on MIBG scan. Four patients with NB are currently alive after additional therapies at 12, 14, 18, and 47 months post BAT infusions. Conclusions: Autologous T cells from heavily pretreated patients could be expanded ex vivo to large numbers, armed with GD2Bi, cryopreserved and thawed for safe IV administration up to total dose of 1.28x109/kg. Ongoing phase II arm of the trial will focus on evaluation of clinical activity of GD2BATs in patients with NB. Clinical trial information: NCT02173093.

scholarly journals Efficacy of Bacteriophage Therapy in Experimental Sepsis and Meningitis Caused by a Clone O25b:H4-ST131 Escherichia coli Strain Producing CTX-M-15

2012 ◽  
Vol 56 (7) ◽  
pp. 3568-3575 ◽  
Author(s):  
Flavie Pouillot ◽  
Maryline Chomton ◽  
Hélène Blois ◽  
Celine Courroux ◽  
Julien Noelig ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Ex Vivo ◽  
Coli Strain ◽  
Neonatal Meningitis ◽  
Lytic Phage ◽  
Experimental Sepsis ◽  
High Concentration ◽  
Bacteriophage Therapy ◽  
Content Type

ABSTRACTWe evaluated phage therapy in experimental infections due to S242, a fatal neonatal meningitisEscherichia colistrain belonging to the worldwide-distributed O25b:H4-ST131 clone that produces extended-spectrum beta-lactamase CTX-M-15. A lytic phage, EC200PP, active against S242, was isolated from environmental water. After determiningin vitroandex vivostabilities and pharmacokinetic properties of EC200PPin rat pups, we assessed the therapeutic efficacy of a single dose of 108PFU using models of sepsis and meningitis in which fatality was 100%. EC200PPwas partially neutralized by human serum. In contrast to the high concentration of phage in the spleen and the kidney, low titers in urine and the central nervous system were observed. Nevertheless, in the sepsis model, EC200PPadministered 7 h or 24 h postinfection resulted in 100% and 50% pup survival, respectively. In the meningitis model, EC200PPadministered 1 h or 7 h postinfection rescued 100% of the animals. The most delayed treatments were associated with the selection of phage-resistant S242 mutants. However, a representative mutant was highly sensitive to killing serum activity and avirulent in an animal model. EC200PPis a potential therapeutic agent for sepsis and meningitis caused by the widespreadE. coliO25:H4-ST131 multidrug-resistant clone.

Recurrence of Severe ITP Following Administration of G-CSF: Remission Following Platelet and Vinca-Alkaloid Infusion.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3222-3222
Author(s):  
Eugene R. Ahn ◽  
John J. Byrnes ◽  
Vincenzo Fontana ◽  
Pamela Dudkiewicz ◽  
Carlos J. Bidot ◽  
...  
Keyword(s):  
Platelet Count ◽  
Platelet Transfusion ◽  
Ex Vivo ◽  
Vinca Alkaloid ◽  
High Dose ◽  
Vinca Alkaloids ◽  
Platelet Counts ◽  
Life Threatening ◽  
Immune Neutropenia

Abstract Introduction: In ITP, platelets opsonized with antibodies are phagocytosed by macrophages. Activation of macrophages often triggers aggravation or relapses of ITP as demonstrated following vaccination, infections. G-CSF stimulates granulocyte colonies but can stimulate macrophages at higher concentrations in vitro. We report recurrence of severe life threatening ITP following G-CSF therapy, successfully managed by selective injury of macrophages with sequential infusions of platelets and vinca-alkaloids. Case Study: A 30 year old healthy Caucasian man developed severe ITP in 9/03 with wet purpura, epistaxis, multiple hematomas in the mouth, tongue and lips and a platelet count <2 K. He suffered severe headaches, refractory gastrointestinal (GI) and genitourinary (GU) bleeding requiring numerous platelet and pRBC transfusions. Increased megakaryocytes were seen in a bone marrow biopsy. CT scans of the head and body were normal, including normal spleen size. ITP was refractory to several measures including high dose glucocorticoids, IV immunoglobulins (IVIG), danazol, rituximab, and vinca-alkaloids. Splenectomy in 5/04 induced a complete remission, lasting for over 3 years. On 2/12/07 he presented with agranulocytosis and neutropenic fever. His Hgb and platelet counts were normal but leukocyte count was 0.9 with absent granulocytes. IVIG infusions began for immune neutropenia with partial improvement of granulocytopenia. Beginning 5/31/07, he was treated with a biweekly regimen of IVIG and Neulasta with normalization of WBC. However, a month following this normalization, patient presented with a platelet count of 9K, wet purpura, epistaxis, multiple hematomas in the tongue and oral mucosa, GI and GU bleeding, headaches and dizzy spells. In spite of high dose IV steroids, daily platelet and pRBC transfusions were required, with little change in platelet counts. He also suffered hypotensive episodes from GI bleeding and pseudomonas bacteremia. Using a rationale described in our previous work (NEJM298:1101, 1978), vincristine 1mg injection was given immediately following platelet transfusion and one week later, 4mg vinblastine immediately following another platelet transfusion. Vinca rapidly binds to transfused platelets and serve as targeted therapy against the activated macrophages that phagocytose platelets. The therapy was effective. Platelet count rose to 72K 1 week after vinblastine, and then normalized. Additional vincristine 1mg was given at discharge. ITP underwent remission. Summary/Discussion: A patient with refractory ITP who underwent CR for over three years after splenectomy suffered severe life threatening thrombocytopenia following injections of G-CSF. This case report is highlighted by the following features. While ITP was in CR, severe granulocytopenia developed which responded to IVIG, indicating an autoimmune cause of leukopenia. Treatment with G-CSF for leukopenia triggered recurrence of severe ITP. Platelet transfusion immediately followed by injection of vinca-alkaloids was successful in inducing remission of life threatening ITP. G-CSF should be used with caution in patients with history of ITP, since it may activate macrophages and trigger relapse of ITP. The immediate sequence of platelet transfusion followed by vinca injection might be particularly useful in this scenario, and is less cumbersome compared to the previously described procedure of incubating platelets ex-vivo with vinca prior to infusion (NEJM298:1101, 1978; AmJHem81:423, 2006).

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