<p> </p>
<p>Learning about ‘behavior’ has always been at the heart of my research endeavors. While my undergraduate work in evolution and ecology exposed me to the science behind why a behavior exists, in my graduate work, I intended to explore how to use something’s behavior to widen its applicability. In this thesis, <em>I will present three works that utilize some of the fundamental</em></p>
<p><em>behaviors (i.e., properties) of elastin-like polypeptides (ELP) to improve existing protein purification methods or explore their applicability in bladder cancer imaging and immunotherapy. </em></p>
<p>Bladder cancer has high recurrence rates (60-70 % annually) that necessitate multiple follow-up therapies making it one of the costliest cancers per patient. In this work, we have attempted to address two leading causes of the recurrence. First is a low sensitivity (62-84 %) and variable specificity (43-95 %) of white light cystoscopy used to diagnose and remove tumors. We aimed to address the heart of this problem, i.e., the non-specific mode of detection using white light. Only the trained eyes can discern abnormal from normal-appearing tissues even then, leaving up to 45% of tumors unresected to colonize and spread. <em>We developed and characterized near infrared dye-peptide-ligand conjugates (NIR-ELP-ligand) that undergo receptor-mediated binding and internalization to human bladder cancer cells in vitro and tissues ex vivo.</em> By using a molecular target-based probe in combination with NIR imaging, we can aid in improving the detection limit via selective binding to the tumor and reduction in background autofluorescence.</p>
<p>Bacillus-Calmette Guérin (BCG) instillation in the bladder is the gold-standard</p>
<p>immunotherapy used after surgical removal of bladder tumors. This was approved as a response to the inefficiency of surgery alone in improving cancer status. It has succeeded by reducing the recurrence rate to 30-50 %. But it comes with the complications of putting a live mycobacterium</p>
<p>in the human body and giving a patient a urinary tract infection right after surgical tumor resection. <em>Thus, we aimed to deliver nucleic acid as immunotherapeutic cargo in a selective manner to elicit robust anti-tumor immune responses while minimizing the side effects due to its carrier.</em> Towards</p>
<p>this goal, we have developed a highly modular and adaptable ELP-ligand fusion protein-based nucleic acid delivery carrier targeted toward bladder cancer. Before developing targeted peptide-based cancer imaging and nucleic acid delivery modalities, we addressed the Achilles heel of peptide-based approaches. The peptide and protein industry suffers</p>
<p>through complex, time-consuming, inconsistent, and low-yielding purification methods. <em>We have developed a scalable, facile, and reproducible protein purification method that delivers ELP and ELP fusion proteins free of host cell proteins and nucleic acids and has low lipopolysaccharide</em></p>
<p><em>content in just 3 h starting from a bacterial pellet. </em>Thus, for a coherent narrative, the thesis is structured as follows:</p>
<p>1. Introduction</p>
<p>2. ELP as a protein purification tag: Development of a rapid purification method for ELPs and ELP fusion proteins.</p>
<p>3. ELP as a cancer imaging agent: Development of NIR-ELP-Ligand imaging probe targeting bladder cancer.</p>
<p>4. ELP as a drug delivery agent: Utilizing ELP-ligand fusion protein in the formulation of targeted nucleic acid delivery carrier to bladder cancer.</p>
| Identifer | oai:union.ndltd.org:purdue.edu/oai:figshare.com:article/22696009 |
| Date | 26 April 2023 |
| Creators | Aayush Aayush (15354604) |
| Source Sets | Purdue University |
| Detected Language | English |
| Type | Text, Thesis |
| Rights | CC BY 4.0 |
| Relation | https://figshare.com/articles/thesis/PhD_Dissertation-Chemistry-Aayush-2023/22696009 |
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