BME PhD Candidate Mackenzie Scully will be defending their dissertation:
Engineering Cancer Cell Membrane-Wrapped Nanoparticles for Targeted Delivery of Cancer Therapeutics
- Location: 590 Avenue 1743, Newark, DE 19713 Conference Room 140AB
- Zoom Link: https://udel.zoom.us/j/5695263714
- Password: scully
- Date: Thursday, December 14, 2023
- Time: 11 am
- Committee: Emily Day, Catherine Fromen, Millicent O’Sullivan, Jennifer Sims-Mourtada
Triple-negative breast cancer (TNBC) is an aggressive and highly metastatic subtype of breast cancer that accounts for 10-20% of all breast cancer diagnoses. It is unsusceptible to traditional hormonal or molecular-oriented therapies because it lacks expression of the receptors these therapies target. TNBC patients are limited to nonspecific treatments such as surgery, radiation, or systemic chemotherapy, which are ineffective in combating TNBC long-term and are severely debilitating because they attack both TNBC cells and non-cancerous cells in the body. TNBC patients face higher risk of tumor recurrence, distant metastases, and adapted chemoresistance leading to an outstanding need for therapies that can precisely deliver anticancer agents to TNBC cells. Prior research has shown that nanoparticles (NPs) can be coated with phospholipid membranes isolated from cancer cells, enabling the cancer cell membrane-wrapped NPs (CCNPs) to preferentially accumulate in primary tumors and secondary metastases that match the source cell type. This homotypic targeting is mediated by “self-recognition” and adhesion molecules on the membrane surface. This thesis demonstrates that the homotypic targeting of CCNPs can be exploited to enhance the delivery of anti-cancer agents to TNBC cells in vitro and orthotopic tumors in vivo to increase apoptosis, reduce cell proliferation, and impede metastatic nodule formation. It also explores the mechanisms by which CCNPs enter targeted cancer cells and investigates the ability to coat NPs with membranes derived from both cancer cells and cancer-associated fibroblasts (CAFs) to further enhance tumor delivery. Chapter 1 introduces TNBC biology and the status of cancer nanomedicine with a focus on membrane-wrapped (i.e., biomimetic) NPs. Chapter 2 describes the materials and methods used to complete experiments in Chapter 3-5, which investigate the ability to treat TNBC using CCNPs loaded with hydrophobic drugs (Chapter 3) or hydrophilic small interfering ribonucleic acid (siRNA) molecules (Chapter 4),or using NPs that incorporate both cancer cell membranes and CAF membranes in hybrid NP coatings (Chapter 5). Finally, Chapter 6 summarizes the impact of this work and provides insights for future directions of this nanotechnology.
Overexpression of the anti-apoptotic protein B-cell lymphoma 2 (Bcl-2) is correlated with poor survival outcomes in TNBC, making Bcl-2 inhibition a promising strategy to treat this aggressive disease. Unfortunately, Bcl-2 inhibitors developed to date have limited clinical success against solid tumors owing to poor bioavailability, insufficient tumor delivery, and off-target toxicity. To circumvent these problems, we encapsulated the Bcl-2 inhibitor ABT-737 in poly(lactic-co-glycolic acid) (PLGA) NPs that were wrapped with membranes derived from 4T1 murine TNBC cells (ABT CCNPs). In Chapter 3, we show that the biomimetic cancer cell membrane coating enabled the NPs to preferentially target 4T1 TNBC cells over non-cancerous mammary epithelial cells in vitro and significantly increased NP accumulation in orthotopic 4T1 TNBC tumors in mice after intravenous injection by over 2-fold compared to poly(ethylene glycol)-PLGA (PEG-PLGA) copolymer NPs. Congruently, the ABT CCNPs induced higher levels of apoptosis in TNBC cells in vitro than ABT-737 delivered freely or in PEG-PLGA NPs. When tested in a syngeneic spontaneous metastasis model, the ABT-CCNPs increased apoptosis and reduced proliferation throughout tumors while suppressing metastatic nodule formation in the lungs. Moreover, the ABT-CCNPs did not alter animal weight or blood composition, suggesting the specificity afforded by the TNBC cell membrane coating mitigated the off-target adverse effects typically associated with ABT-737. These results demonstrate that drug-loaded biomimetic NPs have substantial potential to safely treat solid tumors like TNBC that are characterized by Bcl-2 overexpression.
TNBC’s rapid growth and metastasis are also driven by hyperactive Wnt signaling mediated through the protein β-catenin, which is unfortunately undruggable by small molecule therapeutics. Delivering siRNA into the cell cytosol offers a powerful alternative method to modulate β-catenin expression through RNA interference (RNAi). However, siRNA requires a targeted carrier for clinical use because it exhibits fast renal clearance, endogenous nuclease degradation, serum protein aggregation, and limited cellular uptake. Accordingly, in Chapter 4 we load siRNA into CCNPs and demonstrate in vitro that these NPs can suppress β-catenin in TNBC cells to reduce the cells’ tumorigenic qualities. Compared to unwrapped and non-targeted NPs, the CCNPs exhibit dramatically improved uptake by TNBC cells versus non-cancerous breast epithelial cells. The CCNPs also yield greater silencing of β-catenin at the mRNA and protein levels. Consistent with the improved gene silencing, CCNPs significantly activate senescence in 2D cultured TNBC cells and reduce proliferation in 3D spheroids. This work advances the development of nucleic acid carriers for targeted RNAi of β-catenin in TNBC.
Since the first biomimetic NP design was reported, there has been an explosive development of NPs that combine different core materials, cargoes, and membrane exteriors. While the various platforms’ efficacy at performing specific tasks, such as hindering tumor growth or improving tumor detection, has been widely reported, few studies have examined the mechanism of cellular targeting in detail. It is largely presumed that CCNPs target and enter cells via the same mechanisms cancer cells use to locate and adhere to one another during metastatic dissemination. Chapter 5 investigates the specific extracellular receptors and endocytosis mechanisms CCNPs use to enter TNBC cells by analyzing NP uptake by flow cytometry in the presence or absence of various inhibitors. As a step towards future improvements to biomimetic NP design, this Chapter also describes the development of a hybrid membrane coating that combines 4T1 TNBC cells and CAFs on the NPs’ surface. CAFs are a predominant population of cells in the tumor stroma that create a protective physical barrier that impedes NP delivery throughout the tumor volume. This research examines whether the addition of CAF membranes onto the NP surface improves siRNA-loaded NP accumulation in tumors compared to singular membrane-wrapped NPs in an orthotopic mammary fat pad tumor model. These studies provide important insight to the role of membrane proteins in mediating tumor delivery of biomimetic NPs.
In summary, this dissertation develops and investigates CCNPs loaded with hydrophobic small molecule drugs or hydrophilic siRNA cargoes for the enhanced targeted therapy of TNBC in vitro and in vivo. It also explores the specific receptors and endocytosis mechanisms utilized by CCNPs to enter cancer cells and tests the ability to further enhance delivery via hybrid cancer cell/CAF membrane coatings. Chapter 6 discusses the potential future directions for this biomimetic nanotechnology. These studies are innovative and impactful because they advance the art of membrane-wrapped NPs while also investigating promising new treatments for TNBC.