Abstract 5377: Assessing the chemotherapeutic response of nanoliposomal topotecan in conjunction with radiotherapy on tumor and its microenvironment
Authors: Jyoti Amar , Sethi Pallavi , Fugit Kyle , Langner Ulrich , Clair William St. , McGarry Ronald C. , Anderson Bradley D. , Upreti Meenakshi 
Published: May 2015

Abstract
Topotecan (TPT), a highly active, water soluble analogue of camptothecin, is currently being used in the treatment of various localized cancers from different tissue origin including the breast. Topotecan is classically known for its ability to inhibit topoisomerase I activity as well as accumulation of HIF-1α to block upregulation of pro-angiogenic factors. Despite its known anticancer and antiangiogenic properties the clinical use of TPT is limited owing to its rapid clearance and hematological toxicity. Liposomal encapsulation of TPT can potentially increase its efficacy by shielding it from systemic clearance, allowing greater uptake and extended tissue exposure in solid tumors. We hypothesize that tumor cell biology and treatment response would be more informative when done in the presence of stromal components, like endothelial cells. To that end we have developed a 3D in-vitro/ in-vivo murine model of Triple Negative Breast Cancer where 4T1-GFP tumor cells are co-cultured with their stromal component (2H11 murine endothelial cells) in ‘hanging drops’ of medium. In the present study we have determined the effective dose of radiation and TPT for 4T1 tumor and 2H11 endothelial cells by cell viability and clonogenic assays. Based on the observed toxicity treatment response of actively loaded liposomal topotecan (ALLTs) and free drug with or without radiation exposure (3Gy) in tumor-endothelial spheroids was determined by monitoring the spheroid size and quantitation of apoptotic changes observed.
100nm DSPC and m-PEG DSPE (95:5 mol: mol) nanoliposomes were prepared in ammonium besylate and then TPT was actively loaded by pH gradient method. Tumor-endothelial spheroids grown in hanging drops, were subsequently transferred to cell repellent plates for high throughput in-vitro analysis. Our studies indicate that endothelial cells are more sensitive to TPT treatment (IC50 of 0.13µM) than tumor cells (IC50=63µM). Combinatorial treatment with topotecan loaded nanoliposome (ALLT) and radiation of tumor endothelial spheroid reveals a decrease in spheroid size and severe disruption of the peripheral layers with time. ALLTs based chemotherapy in conjunction with radiation was thus observed to be more efficacious in disrupting the integrity, more specifically the extracellular matrix of tumor-endothelial spheroids. Unlike the free drug which gets hydrolyzed into its relatively inactive carboxylate form, we expect that the ability of ALLTs for extended release of active TPT (more than 24 hours) in a metronomic fashion, may contribute to the therapeutic efficacy of its combined treatment with radiation. Efforts are underway to implant these spheroids in mammary fat pad and dorsal skin fold window chamber and study the effect of ALLTs in combination with radiation on tumor growth and neovascularization and metastasis.
Supported by NCI grants R25CA153954 and CA173609
Citation Format: Amar Jyoti, Pallavi Sethi, Kyle Fugit, Ulrich Langner, William St. Clair, Ronald C. McGarry, Bradley D. Anderson, Meenakshi Upreti. Assessing the chemotherapeutic response of nanoliposomal topotecan in conjunction with radiotherapy on tumor and its microenvironment. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5377. doi:10.1158/1538-7445.AM2014-5377