Reconstituting the 3D Breast Cancer Microenvironment for Chemo and Immunotherapy
Restricted (Penn State Only)
- Author:
- Dey, Madhuri
- Graduate Program:
- Chemistry (PHD)
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- January 18, 2022
- Committee Members:
- Paul Cremer, Major Field Member
James Connor, Outside Unit & Field Member
Christine Keating, Co-Chair & Dissertation Advisor
Lauren Zarzar, Major Field Member
Philip Bevilacqua, Program Head/Chair
Ibrahim Ozbolat, Co-Chair & Dissertation Advisor - Keywords:
- Breast cancer
3D tumor models
Chemotherapy
Immunotherapy
MAIT-MR1
3D bioprinting
Perfusion
Tumor Angiogenesis
Cancer Invasion - Abstract:
- Metastatic breast cancer is one of the deadliest forms of malignancy, primarily driven by its characteristic microenvironment comprising of cancer cells interacting with immune and stromal components. These interactions induce genetic and metabolic alterations creating a conducive environment for tumor growth. Immune-cancer interactions resulting in an immunosuppressive microenvironment further reduces the anti-tumor efficacy of immune-therapeutics. Thus, despite several advancements in cancer treatment, it still remains elusive with only a 5% success rate of new therapeutics entering clinical trials. This necessitates the development of an in vitro tumor model which would not only help us disseminate the steps in cancer progression but also aid in rapid pre-clinical screening of therapeutics. Traditionally, two-dimensional (2D) tumor models or animal models have been widely used for studying cancer growth; however, these models lack the essential cell-cell and cell-matrix interactions of a native three-dimensional (3D) tissue. A tumor in human body grows in 3D and is surrounded by a dynamic microenvironment, which marks the different stages of tumor progression. Thus, it is essential to fabricate 3D in vitro platforms which are capable of incorporating intricacies of the tumor interaction with its microenvironment. To this end, we have developed a 3D vascularized breast cancer microenvironment comprising of metastatic MDA-MB-231 breast cancer cells and human umbilical vein endothelial cells (HUVECs) loaded in human dermal fibroblast (HDF)-laden fibrin, representing the tumor stroma. Employing these cell types enabled us to study the impact of matrix as well as stromal cell density on tumor angiogenesis and cancer invasion, two of the major hallmarks of cancer. Specifically, presence of fibroblasts impacted the transcriptional profile of genes involved in tumor angiogenesis and cancer invasion, which further led to the identification of cancer-specific canonical pathways and activated upstream regulators in these complex 3D cultures. Additionally, immune-cancer crosstalk was explored employing different 3D tumor models to assess the efficiency of a novel T cell receptor- (TCR) modified primary human T cells in identifying and killing MR1 expressing MDA-MB-231 cells. TCR-modified T cells were effective in eradicating ~90% cancer cells in MDA-MB-231-only spheroids and ~70% cancer cells in MDA-MB-231/HDF spheroids, over three days of in vitro culture. The effect of T cell localization on tumor growth was also studied using a hybrid bioprinting approach. MDA-MB-231/HDF spheroids bioprinted proximal to T cells exhibited higher expression of IFN-gamma, granzymes, CCL2 and other cytokines, all indicative of T cell activation. Immune-cancer crosstalk was also explored in a dynamic-flow based 3D bioprinted vascularized tumor model. Heterotypic tumor spheroids, comprising metastatic breast cancer cells (MDA-MB-231), human umbilical vein endothelial cells (HUVECs) and human dermal fibroblasts (HDFs) were precisely bioprinted in a collagen/fibrin biomimetic matrix. Spheroids were bioprinted at proximal (~100 micrometer) and distal (~500 micrometer) locations from a perfused vasculature which revealed enhanced capillary sprouting, angiogenesis and anastomosis for the proximally bioprinted spheroids. Proximally bioprinted spheroids also exhibited higher invasion of cancer cells compared to the distal groups. Doxorubicin, a commonly used chemotherapy drug was perfused through the vasculature at varying dosages. A dose dependent drug response behavior with gradual reduction in tumor growth was observed after 72 h of doxorubicin perfusion. Furthermore, overexpression of several cancer biomarkers was observed for doxorubicin treated groups. Immune-cancer interaction in this complex dynamic microenvironment was studied by perfusing anti HER2-Chimeric Antigen Receptor (CAR) T cells through the central vasculature. Perfusing CAR-T cells for 24 h resulted in extensive T cell recruitment to the endothelium as well as T cell infiltration to the tumor site resulting in decreased in tumor volume. This physiologically-relevant 3D platform paves the way for a robust, high throughput and clinically relevant 3D tumor microenvironment platform for future translation of anti-cancer therapies to personalized medicine for cancer patients.