Discipline: Biological Sciences
Subcategory: Biomedical Engineering
Ruthie Stokes - North Carolina Central University
Co-Author(s): Jodie Fleming,Department of Biological and Biomedical Sciences,North Carolina Central University, Durham 27707; Darlene Taylor,Department of Chemistry and Biochemistry, Biomanufacturing Research Institute and Technology Enterorise(BRITE), North Carolina Central University, Durham 27707; Robert Vacheri, Department of Chemistry and Biochemistry, Biomanufacturing Research Institute and Technology Enterorise(BRITE), North Carolina Central University, Durham 27707
Our lab previously reported the synthesis of a four-component temperature responsive copolymer involving N-isoproypl acrylamide (NIPAAm), 2-hydroxyethyl methacrylate-polylactide (HEMA-PLA), acrylic acid (AAc), and methacrylated hyperbranched polymer glycerol (HPG-MA), named 83-7-1-9 copolymer, that changes from liquid to gel at a critical temperature. 83-7-1-9 copolymer can be used as a drug delivery platform to inject a high payload of chemotherapeutic directly into breast cancer tumors (BraC). Normal delivery of chemotherapeutics is hindered in part by the tumor’s extracellular matrix. Delivery of chemotherapeutic drugs and enzymes like collagenase (CCH) directly inside the tumor should increase therapeutic activity and break down the extracellular matrix. To this end, 83-7-1-9 copolymer was evaluated in vitro, ex vitro, and in vivo as a vehicle to deliver chemotherapeutic drugs and enzymes directly to the breast cancer tumor.
In vitro, MCF 35 cells were treated with doxyrubicin (100 ug/mL) with and without 83-7-1-9 copolymer. Flow cytometry results suggest that doxyrubicin entrapped in 83-7-1-9 copolymer overlaps the cell count curves representative of control and 83-7-1-9 copolymer alone while the free drug doxyrubicin shows complete cell death in 24 hours. Parallel studies with collagenase were conducted to determine if activity of CHC was altered after release from 83-7-1-9 copolymer. A SDS PAGE was used to analyze the data collected from cells treated with Tris, 15% copolymer, copolymer+CCH, and CCH. The gel is being developed and the stains are in the process of being analyzed. Ex vitro studies with mammary tumors were used to optimize the concentration of Liberase needed to soften the tumors. A total of four different concentrations performed in triplicates within the range 0.08 units ? 0.35 units was utilized to assess the degree of tumor softness at the end of a average time of ten minutes. These results indicated that 0.2 units was the optimal Liberase concentration to induce a desired softness in the tumors based on visual appearance and touch. In vivo studies to determine the toxicity of 83-7-1-9 copolymer have been conducted. Fluorescein (FITC) was covalently linked to 83-7-1-9 copolymer and injected at one dose (200 ug/mL) into mammary glands of mice that had a full competent immune system. At the end of two time points (2 days and 30 days) the mice were sacrificed and mammary glands retrieved, and will be assessed by histological analysis to determine toxicity. 83-7-1-9 copolymer is very promising and with further testing can be the vehicle that carries us into the future of cancer treatment.
Funder Acknowledgement(s): NSF MRSEC (DMR-1121107)
Faculty Advisor: Darlene Taylor, firstname.lastname@example.org
Role: I conducted the Liberase Activity Test with the various concentrations to determine the ideal concentrations to soften the tumor. I also conducted the SDS PAGE to see if the collagenase is still active and the amount released at target time points.