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Publication Date

2024-5

First Advisor

Sarah J. Moore

Document Type

Honors Project

Degree Name

Bachelor of Arts

Department

Engineering

Keywords

targeted therapy, spheroids, cancer, mesothelin

Abstract

Cancer is a major burden of disease worldwide, and continues to be the second most common cause of death in the US, after heart disease. Even though the mortality rate of cancer keeps decreasing, there remains an unmet need for more precise and patient-specific treatment with fewer side effects and better efficacy. Targeted therapy for cancer is a more recent approach that can effectively kill cancer cells while also providing patients with less damage to healthy cells compared to traditional therapy. Targeted therapy works by targeting specific molecules or pathways that are involved in the growth, progression, and spread of cancer cells. Mesothelin (MSLN) is a biomarker of cancer cells because of its overexpression on the surface of many types of cancer cells. The Moore lab has engineered multiple variants of a non-antibody protein scaffold, derived from the tenth domain of human fibronectin type III and referred to as Fn3, to have the ability to bind MSLN with high affinity. The interaction of MSLN with another tumor surface protein, CA125, has been observed to mediate cancer cell adhesion, motility, and invasion. The goal of this thesis was to characterize the penetration ability of an engineered Fn3 protein into tumor spheroids in vitro, providing spatial information about the potential of the Fn3 protein to reach throughout a solid tumor in vivo. To enable the Fn3 to be readily visualized in multiple assays, first, recombinant DNA methods were used to add a Flag tag to an existing Fn3 variant. The Flag tag is a short, hydrophilic peptide sequence for which specific antibodies are commercially available for various measurements. Later, the cancer cells were cultured into spherical shape, known as cancer spheroids, and cryosectioned into slices. The slices were treated with protein and stained with anti-Flag Alexa Fluor 488 on the glass slide. The slides were imaged to confirm the ability of Fn3 binding to MSLN. Meanwhile, the parameters for using a mathematical model to estimate the penetration of protein were gathered through experiments, calculations, and literature review. In summary, this thesis reports progress toward measuring and modeling the penetration of a MSLN-binding candidate therapeutic into a solid tumor mass, towards the long term goal of developing these engineered proteins for targeted therapy applications.

Rights

©2024 Sicheng Pang. Access limited to the Smith College community and other researchers while on campus. Smith College community members also may access from off-campus using a Smith College log-in. Other off-campus researchers may request a copy through Interlibrary Loan for personal use.

Language

English

Comments

76 pages: color illustrations. Includes bibliographical references (pages 68-71).

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