title: Absorbable microparticulate cation exchanger for immunotherapeutic delivery.
abstract: An absorbable microparticulate cation exchanger was synthesized as a versatile carrier for biologically active proteins. In this work, acid-terminated polyglycolide (or polyglycolic acid) microparticulates (PG-MP) were surface modified for either sustained release of cytokines or as a platform for immunomodulation. The intended goal was to achieve in situ recruitment/maturation of dendritic cells and activation of T cells for tumor immunotherapy. PG-MP were prepared with a volume weighted mean diameter of 7.02 micro (range: 2.09-14.58 micro). Accessible carboxylic acid groups were determined to be 0.3 mmol/g with a corresponding zeta potential of -21.87 mV in phosphate-buffered saline. Under low magnification, scanning electron microscopy (SEM) revealed a highly textured surface due to processing from repetitive jet milling. However, a moderately porous architecture was noted at higher magnification. Electron spectroscopy for chemical analysis was used to characterize the PG-MP surface before and after adsorption of human granulocyte-macrophage colony stimulating factor (GM-CSF). Adsorption of GM-CSF on PG-MP (PG-GMCSF) resulted in a modest increase in the surface atomic concentration of nitrogen (0.97%). Pretreating the surface with poly-L-lysine (PG/Lys-GMCSF) prior to adding GM-CSF produced a nearly threefold increase in the surface nitrogen concentration (4.20% compared to 1.47%). This manipulation not only increased loading content, but also prolonged the release of GM-CSF released from 6 days to 26 days. ESCA on the post-release PG-MP samples (PG-GMCSF and PG/Lys-GMCSF) revealed a similar residual surface nitrogen concentration (2.26% vs. 2.35%). The observation was consistent with irreversibly adsorbed GM-CSF. It is postulated that irreversibly bound GM-CSF is released over time as a function of microparticulate degradation. Biological activity of released GM-CSF was confirmed by the proliferation of a GM-CSF-dependent cell line (TF-1) in the presence of microparticulates. PG-MP mediated activation of T cells was achieved through irreversible adsorption of either antimouse cd3 plus antimouse cd28 monoclonal antibodies (alpha-cd3/cd28-MP) or antihuman CD3 plus antihuman CD28 monoclonal antibodies (alpha-CD3/CD28-MP) on PG-MP. Irreversibly adsorbed antibodies were capable of activating both resting mouse and human T cells. Intracellular flow cytometry on mouse T cells revealed that nearly 50% of the activated cells produced interferon-gamma (IFN-gamma). This was consistent with a TH-1 or cell-mediated response. In vivo efficacy was evaluated in a mouse flank tumor model showing a significant antitumor effect both alone and in combination. Combination therapy was most effective at preventing tumor implantation (8/8 mice) and was able induce tumor regression (4/7 mice) and/or stable disease (3/7 mice) in a regression model. In these studies, immunohistochemistry was used to confirm local recruitment of dendritic cells. In conclusion, the PG-MP represents a novel absorbable cation exchanger that can be readily manipulated to deliver biologically active proteins for immunotherapy.
authors.affiliation: Division of Gynecologic Oncology, Christiana Care Health Services, Newark, Delaware 19713, USA. wshalaby@christianacare.org
authors.forename: Waleed S W
authors.initials: WS
authors.lastname: Shalaby
authors.forename: Heidi
authors.initials: H
authors.lastname: Yeh
authors.forename: Edward
authors.initials: E
authors.lastname: Woo
authors.forename: Joel T
authors.initials: JT
authors.lastname: Corbett
authors.forename: Heidi
authors.initials: H
authors.lastname: Gray
authors.forename: Carl H
authors.initials: CH
authors.lastname: June
authors.forename: Shalaby W
authors.initials: SW
authors.lastname: Shalaby
mesh_terms: D000042:Absorption; D000818:Animals; D018106:CD28 Antigens / Q000378:metabolism; D017252:CD3 Complex / Q000378:metabolism; D002455:Cell Division; D016207:Cytokines / Q000378:metabolism; D003713:Dendritic Cells / Q000166:cytology / Q000378:metabolism; D016503:Drug Delivery Systems; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism; D006801:Humans; D007167:Immunotherapy / Q000379:methods*; D016623:Ion Pumps / Q000737:chemistry / Q000378:metabolism* / Q000627:therapeutic use; D008213:Lymphocyte Activation; D051379:Mice; D008807:Mice, Inbred BALB C; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000276:immunology / Q000473:pathology / Q000517:prevention & control / Q000628:therapy; D010316:Particle Size; D011100:Polyglycolic Acid / Q000737:chemistry / Q000378:metabolism; D013499:Surface Properties; D013601:T-Lymphocytes / Q000166:cytology / Q000378:metabolism
chemical_list: D018106:CD28 Antigens; D017252:CD3 Complex; D016207:Cytokines; D016623:Ion Pumps; D011100:Polyglycolic Acid; D016178:Granulocyte-Macrophage Colony-Stimulating Factor