A Comparative Study of a Non-Small Cell Lung Cancer Associated EGFR Missense Variant of Uncertain Significance

Author

Vanessa MejiaFollow

Date of Award

Spring 2024

Document Type

Thesis

Degree Name

Master of Science (MS) in Biology

Department

Biology

Committee Chair

Ashley Turner

Abstract

Non-small cell lung cancer (NSCLC) is a form of lung cancer that can be driven by heightened activity of epidermal growth factor receptor encoded by the EGFR gene. Genetic variants in EGFR have been identified that lead to abnormal cell growth and tumorigenesis. The objectives of this study was to 1) determine if an EGFR variant of uncertain significance (VUS) associated with NSCLC is potentially damaging based on evaluation of the ortholog let-23 in the model organism, C. elegans, and 2) identify conserved missense VUS loci associated with NSCLC. Through ClinVar, the EGFR VUS c.845G>C(p.Gly282Ala) was identified in a conserved loci through multiple sequence alignments. Additional analyses were carried out to examine the EGFR VUS’s amino acid chemical class properties and PolyPhen-2 score. Between the wildtype and mutant amino acid chemical class properties, the classification remained consistent with the initial glycine being non-polar, hydrophobic and the mutant change to alanine with similar properties. PolyPhen-2 predicts the EGFR VUS to be probably damaging to the encoded protein (HumDiv=0.999). In addition, protein models were constructed to examine potential structural changes observed between the encoded native and mutant VUS proteins in human and C. elegans. Further investigation will examine the VUS in vivo through the C. elegans let-23 ortholog and model system. Two sets of DNA primers were designed and optimized through polymerase chain reaction (PCR), amplifying the EGFR VUS loci in nematode let-23. To target let-23, a CRISPR RNA guide was designed. These molecular reagents will be utilized for future CRISPR-Cas9 experimentation and microinjection to generate the let-23 VUS C. elegans model. This study aims to provide a comprehensive evaluation of a NSCLC associated EGFR VUS, elucidating its potential clinical significance on its implication for NSCLC and human health.