Date of Award

2021

Document Type

Dissertation

Degree Name

Philosophy (Ph.D)

Department

Pharmacy Administration and Public Health

First Advisor

Ketankumar KP Patel

Second Advisor

Zhe-sheng ZC Chen

Third Advisor

Vikas V. VD Dukhande

Abstract

Melanoma is one of the most aggressive and deadliest types of skin cancer. Currently, off-target toxicities and the rapid resistance development of metastatic melanoma mainly restrict the efficiency of the treatments. This thesis presents efforts towards developing liposomes to address current problems of BRAF-mutant metastatic melanoma.

EphA2-Receptor Targeted PEGylated Nanoliposomes for the Treatment of BRAF V600E Mutated Parent and Resistant Melanoma

In order to address off-target toxicities of the targeted therapy of MEK inhibitor trametinib (TMB), we developed a physically stable EphrinA1-mimicking peptide (YSA) anchored TMB-loaded PEGylated nanoliposomes (YTPLs). The YTPLs were evaluated in BRAFV600E-mutated parent cell lines (A375 and SK-MEL-28) and vemurafenib-resistant cell lines (A375R and SK-MEL-28R). A differential scanning calorimetry (DSC) study confirmed that TMB was retained in a solubilized state within the lipid bilayers. No burst release was observed of TMB in 24 h and negligible hemolysis was observed at therapeutic concentrations of TMB. YTPL showed higher intracellular uptake in parental cell lines compared to vemurafenib-resistant cell lines. Western blot analysis and a cytotoxicity study with the EphA2 inhibitor confirmed a reduction in EphA2 expression in resistant cell lines. Thus, YTPLs can be useful for metastatic melanoma-specific delivery of TMB.

Development of BRD4 PROTAC and anti-fibrotic agent co-loaded PEGylated Nanoliposome for BRAF inhibitor resistant Melanoma

In the present study, we proposed a new treatment strategy for the treatment of vemurafenib-resistant melanoma by targeting to both cancer cells and tumor stroma. A BRD4 proteolysis targeting chimera (ARV-825) and nintedanib co-loaded PEGylated nanoliposomes (ARNIPL) were developed in a synergistic cytotoxic ratio against vemurafenib-resistant melanoma. Both the molecules have extremely poor aqueous solubility. Citric acid was used to improve the loading of both the molecules in ARNIPL. ARNIPL with mean particle size 111.1 ± 6.55 nm exhibited more than 90% encapsulation efficiency (EE) for both the drugs and was found to be physically stable for a month. Both the molecules and ARNIPL showed significantly higher cytotoxicity, apoptosis and downregulation of target proteins BRD4 and c-Myc in vemurafenib-resistant cell line (A375R). Vasculogenic mimicry and clonogenic potential of A375R were significantly inhibited by ARNIPL. Tumor growth inhibition in 3D spheroid of A375R and 3D spheroid of co-culture of A375R+Dermal fibroblasts model with reduction of TGF-β1 was observed with ARNIPL treatment. Therefore, ARNIPL could be a novel therapeutic approach for the treatment of vemurafenib-resistant melanoma.

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