Janorkar, Amol V.

Glioblastoma (GBM) is a primary brain tumor that carries a dismal prognosis, which is primarily attributed to tumor recurrence after surgery and resistance to chemotherapy. Since the tumor recurrence appears near the site of surgical resection, a concept of immediate and local application of chemotherapeutic after initial tumor removal could lead to improved treatment outcome. With the ultimate goal of developing a locally-applied, injectable drug delivery vehicle for GBM treatment, we created elastin-like polypeptide (ELP) hydrogels. The ELP hydrogels can be engineered to release anti-cancer drugs over an extended period. The purpose of this study was to evaluate the biomechanical properties of ELP hydrogels, to characterize their ability to release doxorubicin over time, and to investigate, in vitro, the anti-proliferative effect of Dox-laden ELP hydrogels on GBM. Here, we present microstructural differences, swelling ratio measurements, drug release characteristics, and in vitro effects of different ELP hydrogel compositions. We found that manipulation of the ELP–collagen ratio allows for tunable drug release, that the released drug is taken up by cells, and that incubation with a small volume of ELP-Dox hydrogel drastically reduced survival and proliferation of GBM cells in vitro. These results underscore the potential of ELP hydrogels as a local delivery strategy to improve prognosis for GBM patients after tumor resection.

AbstractCulture conditions that induce hepatic spheroidal aggregates sustain liver cells with metabolism that mimics in vivo hepatocytes. Here we present an array of elastin‐like polypeptide conjugate coating materials (Aminated‐ELPs) that are biocompatible, have spheroid‐forming capacity, can be coated atop traditional culture surfaces, and maintain structural integrity while ensuring adherence of spheroids over long culture period. The Aminated‐ELPs were synthesized either by direct conjugation of ELP and various polyelectrolytes or by conjugating both ELP and various small electrolytes to the reactive polymer poly(2‐vinyl‐4,4‐dimethyl azlactone) (PVDMA). Spheroid morphology, cellular metabolic function, and liver‐specific gene expression over the long‐term, 20‐day culture period were assessed through optical microscopy, measurement of total protein content and albumin and urea production, and quantitative real‐time (qRT) PCR. We found that the amine content of the Aminated‐ELP coatings dictated the initial hepatocyte attachment, but not the subsequent hepatocyte spheroid formation and their continued attachment. A lower amine content was generally found to sustain higher albumin production by the spheroids. Out of the 19 Aminated‐ELP coatings tested, we found that the lysine‐containing substrates comprising ELP‐polylysine or ELP‐PVDMA‐butanediamine proved to consistently culture productive spheroidal hepatocytes. We suggest that the incorporation of lysine functional groups in Aminated‐ELP rendered more biocompatible surfaces, increasing spheroid attachment and leading to increased liver‐specific function. Taken together, the Aminated‐ELP array presented here has the potential to create in vitro hepatocyte culture models that mimic in vivo liver functionality and thus, lead to better understanding of liver pathophysiology and superior screening methods for drug efficacy and toxicity. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 377–388, 2017.

AbstractSeveral studies in the past have formed 3‐dimensional (3D) spheroids of primary hepatocytes in suspension culture. Unfortunately, primary hepatocytes in a suspension environment tend to lose their differentiated function over time, generally due to damage from fluid shear stress and eventual spheroid settling. We have therefore created a novel suspension culture system, by seeding H35 rat hepatoma cells, a hepatocyte‐derived cell line, in a 24‐well tissue culture polystyrene (TCPS) plate placed atop an orbital shaker to create 3D spheroids. To provide stability to the formed spheroids, we used a long‐chain polymer, bovine serum albumin (BSA), dissolved in the cell culture medium and/or coated on TCPS surfaces placed in suspension configurations. Our results demonstrate that BSA coating of culture surfaces resulted in uniform and well‐defined spheroids with little spheroid settling or “flattening” of cell colonies in either static or suspension configurations. In BSA‐coated suspension systems, spheroid size scaled with the amount of BSA dissolved in culture medium. In static uncoated cultures, the normalized rat albumin production levels were enhanced by addition of BSA within culture medium. Thus, both addition of BSA to culture medium and application of BSA as a surface coating appear to be meaningful avenues for tailoring spheroid morphology and function. This 24‐well plate suspension culture system may be a valuable tool for high throughput investigations of liver cell behavior in a stable, uniform, 3D spheroid state. Biotechnol. Bioeng. 2013; 110:2548–2555. © 2013 Wiley Periodicals, Inc.