Bhuvi M's Senior Project Blog

Project Title: Potential Markers to Combat Afatinib Resistance in Esophageal Adenocarcinoma
BASIS Advisor: Wendy Sandor
Internship Location: Norton Thoracic Institute St. Joseph’s Hospital and Medical Center
Onsite Mentor: Timothy Fleming



Project Abstract

Although chemoresistance is a significant problem in cancer treatment, as resistance to a cancer therapy results in cancer-related morbidity, chemoresistance occurs more often with esophageal adenocarcinoma as specific targeted therapies have not been approved for esophageal cancer. Aftatnib is a targeted chemotherapy for the Epidermal Growth Factor Receptor family and is effective until a resistant colony forms. The properties of the resistant colony are shown via an invasion assay. Differences in activated tyrosine kinase receptors between the parental and resistant esophageal adenocarcinoma can be found using a Phosph-RTK Dot Blot Array Kit. These differences will be confirmed through Western Blot Analysis and qPCR. The goal of this study is to identify growth-factor receptors that have been activated in a resistant colony to circumvent afatinib-mediated resistance.

    My Posts:

  • Last Minute Data

    Hello everyone! This past week has been extremely busy with last-minute data collection and finishing up my presentation. Additionally, during the editing process, Dr. Fleming and I decided to change the title of my project, altering the project's outlook. We started the week with a dot blot of the inhibition titration we had initiated. Below,... Read More

  • Transfection Updates and Inhibition Studies

    Hello everyone! Last week, I began another round of Axl transfection into OE33. Here are some pictures from Day 3 of the transfection process: We have switched from serum-free media with the transfection reagent to media with the antibiotic G418. So, some things we can observe are that the kill plate is starting to die,... Read More

  • Harvesting pDNA

    Hello everyone! Last week, I initiated a new transfection for Flo with MAL and OE33 with Axl. I quickly realized that we did not have enough Axl pDNA. Thus, we needed to harvest more pDNA. Plasmid DNA, typically abbreviated as pDNA, refers to small, circular DNA molecules found in bacteria and other microorganisms. They can... Read More

  • Transfection Round 2

    Hello everyone! Last week, I discovered that we had mixed up some of the antibiotics we used for our transfected cells. We conducted two transfections: one involving Axl into OE33 and the other MAL into Flo-1. I was personally overseeing the Axl transfection for my project, while the MAL transfection was conducted for a separate... Read More

  • The Epithelial to Mesenchymal Transition (EMT)

    Hello everyone! This week, I continued to work on the transfected cells. In this post, I will discuss the epithelial to mesenchymal transition as it is a crucial part of the analysis. Epithelial-to-mesenchymal transition (EMT) is a cellular process wherein epithelial cells, typically structured and stationary, transform into mesenchymal cells, which are more flexible and... Read More

  • Transfection Updates

    Hello everyone! This week, I will discuss our transfection data results thus far. Three days following the transfection, we changed the media of the cells. This was the same serum-free media. Here is a picture of the parental cell line with the transfected Axl from day 3. On day 5, we noticed that some cells... Read More

  • Transfection Mechanism

    Hello everyone! This week, I will discuss the transfection mechanism and the protocol we are following. Transfection is a process that genetically modifies cells. There are two types: transient and stable transfection. Transient transfection involves short-term protein expression achievable through chemical or physical methods. Stable transfection, on the other hand, involves long-term protein expression and... Read More

  • The Growth Curve

    Hello everyone! This week, I will discuss the growth curve assay I completed to illustrate the difference between GD and OE33. As a reminder, OE33 is the parental cell line, an esophageal adenocarcinoma isolated from a 73-year-old Caucasian female. This cell line is commercially purchased and utilized at the Norton Thoracic Institute. GD is a... Read More

  • The Reverse Transcriptase Quantitative Polymerase Chain Reaction

    Hello everyone! This week, I will discuss the qPCR experimentation I completed to confirm the dot blot results from last week. RT-qPCR is a Reverse Transcriptase quantitative Polymerase Chain Reaction. This method is designed to identify the amount of RNA encoded within a sample with one assay. The process consists of converting the RNA to... Read More

  • The Dot Blot

    Hello everyone! This week, my primary focus has been summarizing my Trimester 1 research for presentation at SARSEF. Today, I am eager to delve into a pivotal aspect of my study—the dot blot analysis, an intricate method that unveils crucial insights into the activation status of receptor tyrosine kinases (RTKs). A dot blot can be... Read More

  • The Invasion Assay

    Hello everyone! This week, I have been working on a poster based on my original research in Trimester 1, which I will present later this week at the Junior Science and Humanities Symposium. In this blog post, I will focus on the Invasion Assay conducted during Trimester 1, as it laid the foundation for my... Read More

  • Introduction

    My name is Bhuvi, and I am a senior at BASIS Phoenix. Over the next ten weeks, I will be researching chemoresistance in esophageal cancer. I have been interning at the Norton Thoracic Lab for the past two years, primarily observing techniques such as qPCR, Western Blots, organoid creation, and cell culture assays focusing on... Read More