Porter Lab – Porter Lab

From the Bench to Helping the Planet: Going Green for a Better Future

fidalgo — Wed, 04 Mar 2026 19:59:55 +0000

Our names are Jeffery Martin and Vanessa Riolo, and we are the Porter lab’s Green Lab Ambassadors! Together with Dr. Fidalgo, our Green Lab representative, and Dr. Porter’s full support, we are very excited to share more about sustainability initiatives and the various steps our lab is actively taking to reduce our environmental impact and sustainability. Hopefully, we inspire others to do the same!

Porter Lab has taken the initiative to join the Green Labs Program at the University of Windsor. The Green Labs campus initiative is a certification program that aims to fuel student, staff, and faculty participation to operate with more sustainable, efficient, and responsible laboratory practices. Green Labs and the Chemical Control Centre (CCC) empower participants with tools, knowledge, and peer learning to encourage sustainable behaviours while maintaining high-quality teaching and research spaces. Key areas of this program focus on water conservation, energy conservation, waste management, green chemistry, procurement, engagement, education and awareness.

Did you know that research laboratories generate an estimated 2-3% of global plastic waste every year (ACS Publications)? Or that a single ultra-low temperature lab freezer can use as much as 14,000 kWh of energy per year, the same amount of energy as an average family home (ThermoFisher)? Statistics like these were quite alarming to discover, and that is why Porter Lab leaped to participate in the Green Labs initiative happening within the University of Windsor.

As participants in this program, we are provided with a scorecard that has actionable items which address each of the key areas listed above, and points are awarded based on completion of these items. The points are summed to then assign a ranking, ranging from bronze, silver, gold, or the top level, platinum.

As a crucial method of reducing plastic waste in our everyday work, we give a second life to single-use plastic items, which would traditionally be quickly discarded. One of the best examples of how we do this can be seen through our reuse of pipette tip boxes. Whenever a tip box runs out, we simply fill it with more tips, bake it at 140 degrees Celsius to sterilize them, and then they’re ready to be reused!

The pipette tips themselves unfortunately come in plastic bags, but luckily, we have a solution for that too! See below a list of all the ways we reuse our plastic bags and other plastic items.

When running our Western blot experiments to analyze protein expression, we reuse these bags to incubate our membranes with antibodies.
We save plastic bags to help seal and dispose of virus and cytotoxic waste.
We reuse plastic bottles (Ex. media bottles) by washing them and using them for future experiments or by using them to help dispose of virus and cytotoxic waste.
We clean and sanitize all our non-biohazardous plastic 15 mL and 50 mL conical tubes for reuse.

Further, see below a list of the ways we reuse other everyday items in our lab, like gloves and lab coats, and how we also commit to sustainable practices through using advanced technology.

Rather than using a new pair of gloves each day or each experiment, we prioritize decontamination and reuse of gloves
We reuse disposable lab coats and only replace them when the lab coat is damaged or contaminated
Using online repositories, including OneNote and Teams, we store our updated protocols and inventory for our students’ use and reference, rather than maintaining physical notebooks
At the end of each day, all equipment not in use is switched off (including biosafety cabinets, water baths, centrifuges, etc.)
As one of several labs within the CORE Research facility, the lights in our office areas go off when nobody is around.
In the CORE Research facility, we also have shared equipment rooms where expensive lab equipment can be used by many different labs. This helps to reduce the amount of equipment being produced to run each lab

Further, we truly value the efforts of each member of our Porter lab team, especially our undergraduate student volunteers who prioritize these initiatives every day, making our efforts possible. Our volunteers spend most of their time cleaning tubes and plastic bottles to allow them to be reused, baking and sterilizing tip boxes, and autoclaving biohazardous waste to ensure its proper and safe disposal. Without the help of our volunteers, our sustainability initiatives would not be possible! And last but not least, our RAs: Dr. Fifield is responsible for the fridges and freezers organization, and Dr. Lubanska takes care of the undergraduate students’ training and our lab biosafety.

We aim to nurture a lab culture that values sustainability alongside driving research advancements. One of the ways we foster this attitude is through our annual “Elf Day.” Every December, the entire lab spends one day cleaning our lab space by disposing of old items, washing items that are deemed reusable, and recycling any waste that we may find. Of course, to add a layer of holiday cheer, we do all this dressed as Santa’s elves! Additionally, when planning this event and others, we follow the UWindsor Sustainability Event Guide to ensure we host a “Green Event,” through offering sustainable food items (such as plant-based food options at our gatherings), completing zero-waste activities, and applying the 4 Rs (Reuse, Reduce, Repair, Recycle) of sustainability.

We are very pleased that, because of our sustainability initiatives listed above, we have been elevated to the Green Lab Platinum Level; however, we are not stopping there! One of the next prioritized efforts we are taking in conjunction with the goals of MyGreenLab is our participation in the ULT Freezer Challenger, to aim to reduce the energy usage of our low-temperature freezer through removing ice buildup, items that block airflow, clearing out expired or identifiable items, and more!

Going forward, we aim to continue spreading the word of this incredible and necessary program, and we encourage other labs at the University of Windsor (and nationwide) to participate! If you or someone you know works within a research lab, we encourage you to bring these program details forward to your lab supervisor and ask about signing up for this initiative. Every small task makes a big difference, and only together can we help reduce the environmental impact of research to help build a sustainable future for all.

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Groundbreaking Prostate Cancer Research at the University of Windsor

fidalgo — Mon, 22 Sep 2025 14:07:34 +0000

My name is Jeffery Martin, and I am a second-year Master’s student working on the prostate cancer project in Dr. Porter’s lab in the Biomedical Science Department at the University of Windsor. In honour of Prostate Cancer Awareness Month, I would like to share some of the exciting prostate cancer research happening at the University of Windsor! Prostate cancer is the second most common cancer in men worldwide, affecting 76 men daily in Canada (Canadian Cancer Society). Disease management and overall patient outcomes have improved as treatment options continue to evolve, and while effective in many cases, these treatments can sometimes pressure prostate cancer cells to change into a more aggressive type of cancer, known as Neuroendocrine Prostate Cancer (NEPC). NEPC can metastasize to other parts of the body, making the current treatments available for prostate cancer much more challenging. Our research focuses on studying the progression of early-stage prostate cancer adenocarcinoma to advanced NEPC and understanding the mechanism of the cell cycle’s regulation in this process.

We believe that through better understanding prostate cancer progression and the cell cycle’s role in this process, we will become more informed on potential treatment options. As a result of our findings, we are now testing selected drugs on prostate cancer cells to try to develop novel therapeutics that can treat and prevent the progression of prostate cancer.

Additionally, we are also looking for novel biomarkers that can detect prostate cancer progression, and we are using non-invasive liquid biopsies to do so! Biomarkers are measurable indicators, like genes or proteins found within the body, that reveal a biological process or disease within the body. Common biomarkers for prostate cancer include PSA (prostate-specific antigen), PSMA (prostate-specific membrane antigen), and AR (androgen receptor). Although these markers are very effective at detecting early stages of cancer, unfortunately, as the disease progresses, these markers are often lost. This leads to the need for the development of new biomarkers to track disease progression.

Further, we hope to find biomarkers that can be detected in early stages of prostate cancer and indicate whether or not disease progression is expected. This would help to guide treatment plans and create a more personalized medicine approach going forward. As mentioned, we plan to use non-invasive liquid biopsies instead of traditional biopsies to detect these biomarkers. Although they are effective, traditional biopsies are invasive, painful for patients, and can lead to potential risks down the road. With liquid biopsies, we can detect these biomarkers in a less invasive way by using a blood, urine, or saliva sample. Through a collaboration with Dr. Kanjeekal at Windsor Regional Hospital, we will be searching for these new biomarkers with liquid biopsies collected from prostate cancer patients.

The research we conduct is mainly done in vitro, which means it’s done in a controlled environment, like in a test tube, instead of a living organism. Specifically, we work in a cell culture room for a lot of our experiments. At the moment, I’m working with 4 different prostate cancer cell lines, which are all derived from prostate cancer patients. We use these cell lines to mimic the progression of disease, and also for drug testing to see how the cells respond to treatment.

We also do some in vivo experiments, which means using living organisms. We specifically use zebrafish as a model system for testing the drugs! Using Zebrafish is becoming increasingly common around the world for drug testing and there are many benefits of this model system. First, Zebrafish are genetically similar to humans, and they are a cost-effective model that requires easy maintenance. Next, they develop quite rapidly, which is great for experimentation, and they have a transparent embryo, allowing for easy visualization. Finally, and most importantly, Zebrafish lack a functional immune system for the first 10 days of their life! This allows us to transplant human cells directly into zebrafish and then observe how these prostate cancer cells behave in a living system. From there, we can test various drugs in the Zebrafish with the prostate cancer cells and see how the cells respond to treatments in this environment. This will also help the results from our studies translate more easily and rapidly to use in humans.

This fundamental work has been supported by grants from Cancer Research Society (CRS), Ride for Dad and WE-SPARK Health Institute. Through this funding, we have made some excellent progress so far, and I look forward to the progress that we will continue to make in the upcoming years! Additionally, through submitting my project proposal to various organizations, I have been lucky enough to receive funding from NSERC (Natural Sciences and Engineering Research Council of Canada), the Windsor Prostate Cancer Scholarship, the Dr. Michael L. Petras Memorial Scholarship, and the Doctor Family Health Research Student Support Award. Each of these awards has helped fund my education and allowed me to spend more time working in the lab. Further, another Master’s student working on this project, Christian Kassa, was recently awarded a Canadian Cancer Society Research Training Award. This is a very prestigious award with only 6 recipients across all of Canada. Christian just started his graduate studies this year and will be helping advance this project over the next 2 years!

A highlight of my project has been the various connections I have made with the community and other researchers at various conferences and local events. At the WE-SPARK Health Research Conference, I had the privilege of being a LEARN program host, which stands for Lived Experience Accelerating Research Knowledge. As a LEARN host, I was paired up with two local prostate cancer survivors who were able to share their lived experiences with me and give insight into the patient perspective. Oftentimes, when working in the lab, I forget about the direct impact my work can have, and these connections remind me of the importance of research. Further, the two patient participants that I was paired with are both members of Windsor’s local Prostate Cancer Support Group, and through making a connection with them, I was invited to speak to their group this upcoming November! Presenting our important work at various conferences has attracted the interest of other researchers and physicians for possible collaborations.

It’s important to recognize everyone involved in this project as none of this work would be possible without the help of our multi-disciplinary team. This project is supervised by Principal Investigator, Dr. Lisa Porter, and Research Associates, Dr. Elizabeth Fidalgo and Dr. Bre-Anne Fifield. Our clinical collaborator is Dr. Sindu Kanjeekal, the Chief of the Department of Oncology at the Windsor Regional Cancer Centre. In-lab experiments are carried out by myself, Jeffery Martin, fellow graduate student, Christian Kassa, and undergraduate student, Kaitlin Ferraro. Previous lab work was completed by Dr. Martin Bakht while a PhD student in our lab, and undergraduate thesis student, Maria Badalova. Knowledge translation work is completed by our Clinical RA, MSc Samavia Ahmad, and Masters of Translational Health Science student, Hannah Ferasol. Finally, none of this project’s advancements would be possible without the help of everyone else in Porter lab who help to provide meaningful feedback and insights that help drive this research further.

I hope you enjoyed learning a little bit more about the prostate cancer research taking place at the University of Windsor. Feel free to watch the video here if you want to see more of the work we do. If you have any questions, please leave a comment below, and I will get back to you! And if you enjoy reading it, please share it below! Jeffery

Undergrads – 2023

fidalgo — Thu, 04 May 2023 18:47:44 +0000

Amy Llancari

My project focuses on differentiating the progression of Non-alcoholic Fatty Liver Disease (NALFD) into Hepatocellular Carcinoma (HCC) between male and female mice. HCC is the sixth most prevalent cancer in the world, yet it ranks as the third leading type of cancer deaths with an accelerated increase in incidence rates. NAFLD is a major risk factor for HCC, and is emerging as the most common chronic liver condition in the Western world. Prevalence of both HCC and NAFLD is significantly higher in men than women worldwide, suggesting sex-specific differences may play a role in disease susceptibility. My project will focus on investigating these sex-specific morphological and cell expression pattern differences. This will provide insight on potential protective mechanisms found in female mice inhibiting liver injury, in addition to potential diagnostic markers and molecular treatment targets during NAFLD progression.

I have been part of the Porter since my first year of undergrad. My time in the Porter Lab has been the highlight of my university career. I cannot put into words how thankful I am for the unwavering support and guidance I’ve received throughout my project. Dr. Porter and the RA’s have been exceptional role models, their dedication to the scientific community is truly inspiring, and their passion for research is contagious. Being a part of this incredible team has been an honor, and has provided me with so many opportunities both in research and community involvement. My time in the lab has taught me valuable research skills, as well as transferable skills that I will carry with me throughout my career. As I pursue my Masters in the Porter Lab, I am excited to continue to learn and grow as a researcher.

Alan Cieslukowski

Efficient targeting of multiple components of a tumour might be a successful strategy in aggressive types of cancer such as glioblastoma (GBM), which remains the most common and malignant primary brain tumour with an extremely poor patient survival of less than 15 months. The significant therapeutic challenge posed by GBM stems from its genetic and phenotypic heterogeneity fueled by characteristics such as aggressive and treatment-resistant populations of Tumour Initiating Cells (TICs) and high levels of angiogenesis contributing to tumour evolution. TICs, which are responsible for GBM patient relapse, thrive in niches close to blood vessels where they interact with endothelial cells (ECs), exit the cell cycle, and evade therapies. Targeted antiangiogenic drugs, preventing GBM cells from recruiting new blood vessels are only temporarily effective in 50% of patients, resulting in acquired secondary resistance by the tumour. This project explores the TIC-EC interplay and its role in propagating tumour aggressiveness and therapy resistance. This project investigates the impact of ECs on the aggressive characteristics of individual, specific populations of TICs using GBM patient-derived systems, including 3D organoid models and zebrafish patient-derived xenografts (PDXs). Considering that NOTCH1 signaling regulates both TIC and EC populations in GBM, this project dissects the role of NOTCH pathway in TIC-EC interaction. Elucidating the details of cell cycle signaling and specific populations of aggressive TICs with dependence on the EC component will contribute to the identification of new and better therapeutic targets and personalized approaches to treatment of patients with GBM in the future.

Depen Sharma

Tuberous Sclerosis Complex (TSC) is a genetic disorder affecting 1 in 6,000 births, leading to benign tumour formation in various body parts such as the brain, kidney, skin, and lungs. This condition arises from mutations in the TSC1 or TSC2 genes and presents a wide range of neurological symptoms, making diagnosis challenging. Tuberin, a protein encoded by the TSC2 gene, is critical in regulating cell growth, proliferation, and the cell cycle. A key element in TSC is the interaction between Tuberin and Hamartin, encoded by the TSC1 gene. Hamartin ensures Tuberin stability by inhibiting its interaction with the E3-ligase HERC1, which would otherwise lead to Tuberin degradation. Tuberin also regulates the G1/S and G2/M checkpoints in the cell cycle, which are crucial for controlling cell growth and division.

Our research explores the potential role of Cyclin B1, a cell cycle protein, in stabilizing Tuberin levels without Hamartin. Cyclin B1 is essential for mitotic entry, and its function has yet to be entirely understood. It has five serine residues that can be mutated to create a phospho-null Cyclin B1 construct (Cyclin B1 5A), which we utilize in our investigation. Previous work has shown that Tuberin can bind Cyclin B1 during the G2 phase of the cell cycle, but more is needed to know about their interactions when Hamartin is present. Cyclin B1 may be essential in Tuberin stabilization during the G2/M transition. Understanding the relationship between Cyclin B1 and Tuberin in TSC could provide valuable insights into the disorder’s molecular mechanisms and potentially lead to novel therapeutic approaches. By elucidating the role of Cyclin B1 in Tuberin stabilization, we hope to contribute to the ongoing efforts in understanding and treating Tuberous Sclerosis Complex.

Emmanuel Boujeke

My research focuses on Glioblastoma (GBM). GBM is the most frequent and aggressive primary brain tumour in adults. GBM patients have an extremely poor prognosis, with survival averaging only 14 months post-diagnosis. Our lab has been investigating the role of a novel cell cycle regulator (Spy1) in the development and progression of many cancers, including GBM. The focus of my project is to investigate the interplay between Spy1 and some of the known drivers of GBM. Using a transgenic mouse model created by the lab, I study the impact of Spy1-mediated effects on gene expression, self-renewal potential, stemness, and oncogenic transformation in the face of aberration in selected oncogenes and tumour suppressors known to drive glioma progression in neural stem cells.

I joined the Porter Lab at the beginning of my second year of University as an outstanding scholar. Having no prior research experience, I felt intimidated and unsure, worried that I would not be good enough for the lab. Fast forward to two years later, I can now say that joining the Porter Lab was one of the best decisions I made in university. Being surrounded by intelligent, motivated, and driven people inspired me to be persistent and hardworking both inside and outside of the lab. Through my thesis, I learned to think critically and creatively, mentor others, and overcome obstacles. The RA’s and Dr. Porter were exceptional role models and provided unconditional support despite my frequent mistakes. Through this experience, I learned what it means to do research and found my place in it.

Lia Oschanney

Liver cancer is one of the leading causes of cancer death globally, of which 90% of cases are Hepatocellular Carcinoma (HCC). One risk factor of HCC is Non-Alcoholic Fatty Liver Disease (NAFLD), often triggered by sedentary lifestyle, poor diet, and obesity. Most cases of HCC are cirrhotic and can start from excess fat accumulation in NAFLD. Chronic fat can trigger inflammatory responses that damage cells, leading to the liver initiating a proliferative response to recover. However, over time the liver shifts to a fibrotic state to maintain structural integrity with low levels of proliferation, characteristic of Non-Alcoholic Steatohepatitis (NASH). As fibrosis becomes extensive, the liver becomes cirrhotic, and the environment increases HCC risk; however, 20% of HCC cases are non-cirrhotic arising directly from NAFLD. The atypical cyclin-like protein Spy1 is capable of binding to CDKs, increasing cell division and overriding checkpoints. Spy1 has been shown to have a role in cancers, including HCC. Previous research has shown that mice with an overexpression of Spy1 in the liver develop NAFLD and HCC with decreased fibrosis, suggesting that elevation of Spy1 drives a proliferative response in the presence of cell damage, thus increasing HCC risk. To better understand the mechanisms that govern this response and the role of Spy1, this project examines NAFLD development in a transgenic mouse model driving Spy1 overexpression in the liver. This work may lead to the use of Spy1 as a potential drug target in the treatment of HCC.

In my first year, I knew I had a strong interest in research, but I was unsure how to take the first steps towards getting involved. Then, I discovered Dr. Porter’s website and was immediately captivated by the amazing work being conducted in the lab. However, just as I was eager to get started, the pandemic hit, and I was unable to set foot in the lab for nearly two years. Despite this obstacle, the lab kept me engaged through peer mentoring, which continued to fuel my curiosity and teach me the fundamental principles underlying research. When I finally had the opportunity to work in the lab as a third-year student, I had limited practical experience due to the majority of my undergraduate studies occurring online. However, under the guidance of the Porter lab team, I learned a multitude of techniques and quickly progressed from a hesitant and uncertain beginner to a confident and independent researcher capable of working on my own project. This experience has not only prepared me for future research endeavors but has also equipped me with invaluable skills that will undoubtedly serve me well in my future career in medicine. Completing my undergraduate thesis was an incredibly rewarding experience, and it would not have been possible without the support of my peers and the guidance of Dr. Bre-Anne Fifield and Dr. Lisa Porter. I am grateful for the opportunity to have worked in such an inspiring environment and to have learned from some of the brightest minds in the field.

Vanessa Vuong

The cell cycle contains checkpoints that delay mitotic progression and allow for DNA repair before cell division. Our cells receive thousands of DNA lesions a day and this damage can be caused by radiation, drugs, and other processes, leading to cellular mutations and carcinogenesis. There are two types of damage, single-stranded DNA breaks or the more lethal double-stranded DNA breaks (DSB). Upon receiving DSBs, the DNA damage response (DDR) pathway activates p53, which is a tumour suppressor protein that induces apoptosis or cell cycle arrest for cellular repair. Tuberin (encoded by the TSC2 gene) is another tumour suppressor protein that regulates the G2/M transition in the cell cycle and negatively regulates protein synthesis and cell growth. Mutations in tuberin can lead to the multisystem autosomal dominant disease known as tuberous sclerosis (TSC). Porter lab has previously shown that regulates mitotic onset through cellular localization of the G2/M Cyclin, Cyclin B1. My project focuses on the Tuberin/Cyclin B1 complex and its role in G2/M arrest for DNA damage repair. In this study, tuberin knockout NIH3T3 (mouse) cells were created using CRISPR editing technology. The cells will then be treated with etoposide, which is a chemotherapy agent that forms a complex with DNA and topoisomerase II to prevent DNA religation and this induces DNA damage. Wildtype NIH3T3 cells are compared to the knockout line and their cell cycle profile and DNA damage will be visualized using flow cytometry. Results have shown that without the prescence of tuberin, cells will return to G1 much faster since tuberin is not present to bind to cyclin B1 to prolong G2/M arrest. This project will provide greater insight into DNA damage induced carcinogenesis, TSC, and other proliferative diseases.

Joining Porter lab as a member of the peer mentor network and eveutally moving up to complete my undergraduate thesis was truly the best experience! I got to meet many wonderful people who help guide me throughout my struggles with getting data, especially those in the tuberin group who I had to see everyday. The problem-solving, critical thinking, and hands-on techniques I learned during my thesis are skills that I will certainly use in my future career. Best of luck to the incoming thesis students and new undergrads!

Undergrads 2022 – part 3

fidalgo — Tue, 24 May 2022 20:36:48 +0000

Determining the Dependency of Spy1 based on Rb Status in Triple Negative Breast Cancer

Breast cancer is the second most common cancer worldwide and the most common cancer among women. Triple Negative Breast Cancer (TNBC) is a particularly aggressive form of breast cancer with many subtypes based on gene expression profiles. There are currently no targeted treatments for TNBC due to its molecular characteristics, urging the discovery of new therapeutic targets. Potential therapeutic avenues are the cell cycle and its mediators which play an important role in cancer formation and progression. Spy1, a cyclin-like protein, promotes cell proliferation through the G1/S and G2/M checkpoints. Spy1 promotes proliferation even in the presence of DNA damage, overriding checkpoints and increasing cancer susceptibility. While Spy1 has been found to be elevated in breast cancer, its unique binding structure makes for an ideal candidate for cell cycle inhibition therapy. The retinoblastoma tumor suppressor protein (Rb) is known to regulate the DNA damage response system and is key in regulating the cell cycle. However, studies have shown that Rb is often mutated in TNBC inducing deregulated cell cycle progression potentially leading to tumor development. For some breast cancer subtypes the presence or absence of Rb (Rb status) can dictate response to treatment by cell cycle inhibitor drugs. Using in vitro TNBC models (MDA-MB-231 & Bt549 cell lines), this study aims to determine if Spy1 can override checkpoints independently of Rb status, and if elevated levels of Spy1 alter this response. These results could provide further guidance in developing cell cycle inhibition targeted therapies and potentially better TNBC patient outcomes.

“Before transferring to the University of Windsor, I wasn’t sure of how to get involved in research. Being in second year at the time, I was intimidated by research and unsure about my capabilities as a scientist. When I enrolled in CURES (cancer undergraduate research experience students), a course with Dr. Lisa Porter, I learned about the diversity of local cancer research taking place in Windsor-Essex. This course fuelled my motivation to learn, and genuine passion to contribute in this field. I found my confidence as a researcher when I joined Porter Lab in my fourth year, where I was thrilled to be given the opportunity to complete my undergraduate thesis. Up until my last year of University, I never knew myself to be as confident and disciplined as I am now. Being in a research lab has accelerated my learning, competency, and knowledge translation in such a short time. The atmosphere of Porter Lab is extremely motivating, and the people are supportive and patient. Porter Lab offers each member numerous opportunities to move forward science within the community and has helped me adapt to becoming a leader in science- a field I never felt I fit in before. During my time in lab, I have learned a multitude about the nature of science and what is involved in basic science research, including all the time invested in planning experiments and writing grants. Completing my undergraduate thesis was extremely rewarding and I am excited to continue my research in a master’s degree with Porter Lab, working alongside all the Porter Lab rats!”

The Impact of Cyclin B1 on Tuberin Stabilization

Tuberous Sclerosis Complex (TSC) is an autosomal dominant disorder caused by mutations in either TSC1 or TSC2, genes that encode the proteins Hamartin and Tuberin respectively. Hamartomas (benign tumours), skin lesions, neurological symptoms, renal dysfunctions, and retinal malformations are often present in TSC patients with varying severity. Tuberin and Hamartin regulate protein synthesis through mTORC1 inhibition. Tuberin can also delay mitotic onset at the G2/M cell cycle transition by binding to Cyclin B1. Hamartin has been shown to stabilize Tuberin by inhibiting its ubiquitination by HERC1 and subsequent degradation. Preliminary data from our lab suggests that Cyclin B1 may also contribute to the stabilization of Tuberin levels during the G2/M transition. Phosphorylation status of the cytoplasmic retention sequence (CRS) of Cyclin B1 plays an important role in the formation of the Tuberin/Cyclin B1 complex. The unphosphorylated CRS form of Cyclin B1 (Cyclin B1 5xA) binds stronger to Tuberin compared to the phosphorylated form (Cyclin B1 5xE). My thesis investigates the role of Cyclin B1 in Tuberin stabilization. HEK293-TSC1 null cell lines (IC2) were transfected with varying concentrations of Cyclin B1 5xA DNA and Tuberin protein levels were quantified by Western blot techniques. After blocking HEK293 cells in G2/M using thymidine and collecting them at different timepoints, endogenous Tuberin levels at different stages of the cell cycle were also quantified with Western blot techniques. Understanding the role of Cyclin B1 in Tuberin stabilization will shed light on cell proliferation and growth mechanisms that underlie tumorigenic disorders.

Characterization of Multiple Myeloma Cells Treated with CDK Inhibitors

Multiple myeloma (MM) is an aggressive hematopoietic cancer that is caused by the abnormal growth of plasma cells in the bone marrow. MM has a poor prognosis and most patients will eventually relapse with a more aggressive and untreatable form of the disease.
Cyclin Dependent Kinase Inhibitors (CKis) are an emerging targeted therapeutic option for Relapsed MM and have been investigated in many clinical trials. Flavopiridol is a first-generation non-specific CKI that inhibits CDK1, CDK2, CDK4, and CDK7. Flavopiridol has had high pre-therapeutic efficacy, but limited clinical success as a monotherapy. Dinaciclib is a second-generation CKI that is more selective and inhibits CDK1, CDK2, CDK5, and CDK9. Dinaciclib has also had low efficacy in clinical trials. In this study, we compared MM cells (U266B1) treated with Dinaciclib to Flavopiridol in-vitro by examining cell proliferation, apoptosis, and cell cycle profiles. Our data shows that there is no significant difference in cell proliferation or apoptosis of U266B1 cells treated with Dinaciclib and Flavopiridol after 24 hours, 48 hours, and 72 hours. Cells treated with Flavopiridol were more frequently arrested at the G2M checkpoint compared to Dinaciclib after 72 hours. Better characterization of MM cells treated with different types of CKIs will help to better understand their lack of clinical success as a monotherapy.

Undergrads 2022 – part 2

fidalgo — Tue, 17 May 2022 23:16:14 +0000

Applying CRISPR/Cas9 and Fluorescent Tools to Dissect the Role of Tuberin in Cell Cycle Regulation

How cells regulate their growth and division involves a tightly controlled integration of many mechanisms. In cells, Tuberin (gene – TSC2) is a protein in the Tuberous Sclerosis Complex (TSC) which modulates cellular growth, size, and proliferation. Mutations in the proteins forming the TSC can cause Tuberous Sclerosis Complex, an autosomal dominant disorder characterized by multisystem pathologies, and is often associated with benign hamartomas in the brain, kidney, lungs, and skin. The focus of my research is to clarify the role of Tuberin in the regulation of cell size and proliferation at the G2/M cell cycle checkpoint. During late G2, Tuberin retains Cyclin B1 (gene – CCNB1), a mitotic cyclin, in the cytoplasm thereby prolonging mitotic onset. We constructed six TSC2 mutants that harbour clinically relevant mutations which are known to destabilize the TSC. Interestingly, these mutations fall within the Tuberin Cyclin B1 binding domain. By over-expressing these Tuberin mutants in Tuberin null cells, an increased mitotic index is observed indicating a dysregulation of the G2/M transition. The resultant phenotypes are analyzed by flow cytometry, co-immunoprecipitation, and immunofluorescence. To aid in the temporal study of the cell cycle, we aim to validate successful CRISPR/Cas9-mediated knock-in of a near infrared (iRFP720) tag within the TSC2 gene of HEK293 cells, creating an endogenously expressed fluorescent Tuberin-RFP fusion protein. This new cell line will be a powerful tool to dissect the roles of Tuberin in regulating cellular growth and division and can provide a deep understanding of proliferative diseases like TSC and cancers.

“When I first applied to be part of the Porter Lab back in the Spring of 2019, I knew that I was joining a well-recognized and large research lab on campus which has continuously been on the forefront of scientific discovery. However, I did not foresee the new family I would make during my time here. Over the past few years, I have grown from being a student volunteer that started off by filling ethanol bottles and pipette tip boxes to working on a project that uses CRISPR technology to create a modified HEK-293 cell line! My progress as a young scientist has been a direct result of the mentorship and teaching opportunities in the Porter lab ecosystem. It took no time for me to get my hands dirty with experiments at the start, and while the COVID-19 pandemic had put a slight pause on my progress, the lab still provided a means of learning new skills and leadership opportunities through the Peer Mentor Network! I have made many friendships during my time here, and while some of these friends are also starting new chapters in their careers and lives, our relationship will prove to withstand the test of time. As I look forward to a life in medicine, these past few years have created a special place in my heart for biomedical research and it will certainly be a pillar of my future career. I am beyond grateful for the constant support that Dr. Lisa Porter and Dr. Elizabeth Fidalgo da Silva have given me this past year while also pushing me to do expand my scholarly boundaries. While embarking on my thesis this year, I have learned so many skills from troubleshooting experiments (shoutout to Adam Pillon for literally always being there to help) to communicate my work to both a general and scientific audience. As this chapter in my life nears an end, I will forever cherish the friendships and skillsets I have developed and will always have a special place in my heart for my fellow Porter lab rats. #porterlab #lifechanging #mentorship #futureinscience”

Spy1 Exacerbates the Long-Term Effects of Parity on the Mammary Gland

Age is a significant variable in cancer development, and approximately 1/3 of breast cancer cases occur in patients older than 70. Another emerging risk factor is parity, or childbearing, which may be linked to cellular changes that affect a woman’s risk of developing breast cancer over the course of her lifetime. These changes are thought to result from the mammary gland not reverting to normal

after lactation and involution – a developmental remodeling process where the milk secreting cells are cleared and replaced with adipocytes after weaning. Aberrant expression of the cyclin-like protein Spy1 has been shown to stimulate precocious development, resulting in disrupted morphology and oncogenesis within the mammary gland. Preliminary data suggests that the mammary glands of mice overexpressing Spy1 do not fully regress following lactation and involution, which may predispose them to breast cancer. We hypothesize that the overexpression of Spy1 exacerbates the long-term effects of parity on mammary gland morphology. To investigate this, we performed hematoxylin and eosin (H&E) staining as well as immunohistochemistry (IHC) on paraffin embedded sections, and whole mount staining of MMTV-Spy1 mice, a transgenic mouse model that overexpresses Spy1 within the mammary gland. We then compared the mammary gland morphology of parous MMTV-Spy1 mice to nulliparous MMTV-Spy1 mice, parous control FVB

mice, and nulliparous control FVB mice. This research begins to improve our understanding of Spy1’s role in regulating proliferation and apoptosis, contributes to our overall knowledge of breast cancer dynamics, and further solidifies Spy1 as an important target for treatment.

“When I first joined Porter lab, I was a volunteer with very little experience under the unprecedented circumstances of a pandemic. The amount of growth I have experienced as a scientist since then is astonishing, and I could not have imagined the full extent of it before I started as a thesis student. The environment and learning opportunities at Porter lab have been exceptional thanks to everyone’s commitment to our lab’s values, and I have met people and learned skills that will stay with me for the rest of my life. It still seems surreal to me to have been lucky enough to apply concepts, techniques, and an understanding of the scientific method from undergraduate courses to a real project with Implications on the field of mammary development and breast cancer research under the guidance of my mentors. I’m excited to pass the torch onto the next group of undergrads at the end of this journey, and excited to keep up with the lab’s future accomplishments.”

Undergrads 2022 – part 1

fidalgo — Mon, 09 May 2022 23:16:25 +0000

Control of Stemness in Glioma via Nanoparticle-Mediated Regulation of CD44 Receptor

Glioblastoma, also known as glioma or GBM, is the most common and aggressive malignant primary brain tumor which continues to puzzle researchers as decades of efforts to find an effective therapy has proven to fail. Barriers to success of the available treatment include impermeability of the blood brain barrier (BBB) to drugs, tumor heterogeneity, and existence of aggressive and resistant Tumour Initiating Cells (TICs). Collaborative studies in our lab show that conjugated polymer nanoparticles (CPNs) can serve as a potential therapeutic approach to overcome these barriers. CPNs are labeled with hyaluronic acid (HA-CPNs) which is a ligand for one of the most prominent receptors found on TICs, CD44. Our data to date has demonstrated that HA-CPNs cause selective decrease in proliferation, migration potential, and expression of markers of aggressiveness and self renewal in CD44+ glioma cells. This study investigates the mechanism by which HA-CPNs exert their effects and how the uptake of the nanoparticles is regulated. We study the status of CD44 receptor on the protein and mRNA levels as well as how manipulation of its levels affects glioma aggressiveness. We explore inhibition of Clathrin Mediated Endocytosis as responsible for the uptake of HA-CPNs and use it to validate selected effects of HA-CPNs. This work will reveal molecular mechanism behind HA-CPN anti-glioma effects which will further contribute to more in- depth evaluation of this nanoparticle system as a potential therapy for patients with GBM.

“Since joining Porter Lab in my second year of undergrad, I would have never thought this experience would have been so rewarding. Before my time in the lab, I had little knowledge of what the research side of academia was like. Through the COVID-19 pandemic, I found myself thinking largely about my future and through working in the lab via virtual platforms and maintaining my interest in research, I began my thesis in my forth year in person. Although this experience was such a learning curve, I have never been so appreciative and lucky to have this opportunity. Learning and gaining valuable skills in the lab such as trouble-shooting, team work, and resiliency, I can honestly say research has become such a passion of mine.

In my time in Porter Lab, I have created so many friendships with my fellow lab mates that I know will last forever. If you asked me in my first year what I thought I would accomplish during my undergrad, I would have never thought I could say that on a micro level, I am contributing to the advancement of cancer research in my community. I am excited to pursue my masters degree in biological sciences with Porter lab and eager to see all the change each and every member contributes to cancer research.”

Novel Cell Cycle Therapeutic Strategy Against Type II Medulloblastoma

Medulloblastoma (MB) is one of the most common malignant brain tumours, occurring mainly in children between the ages of 1 to 9 years. The standard of care which is currently being applied for MB includes combination of radiation, surgery, and chemotherapy. This treatment is aggressive and is only effective for a small percentage of patients due to the presence of aggressive and therapy resistant populations of Tumour Initiating Cells (TICs). A recent approach in clinical trials is the use of cell cycle regulator proteins, CKIs, which are synthetic cyclin-dependent kinase inhibitors. Our lab focuses on a specific cell cycle regulatory protein, Speedy (Spy1) which promotes proliferation and overrides cell cycle checkpoints established by natural CKIs. Spy1 plays a role in the maintenance and expansion of stemlike populations of TICs in different types of brain tumour. It is our hypothesis that Spy1 can drive tumour initiating cells in medulloblastoma in face and despite ongoing therapy. In this study we investigate and reveal the role of Spy1 in MB in response to known therapeutic agents and Spy1 mediated effects on the dangerous TIC populations in the treatment setting. In addition, Sonic hedgehog (Shh) MB, a type 2 medulloblastoma, is studied and manipulated to assess potential effects of Shh pathway in the regulation of Spy1. This project is the first one to demonstrate that standard of care treatment of MB leads to increased levels of Spy1 and selection of resistant TIC populations. We show that a key molecular driver in MB plays a role in the regulation of Spy1 levels which is an important finding in further studies seeking for novel effective therapeutic strategies against MB. Overall, my project will contribute to better understanding of the role of Spy1 in MB and its application as a potential therapeutical target in the future.

“Joining the Porter Lab at the end of my first year was the best decision I made in my University experience; it was life changing. I met a lot of amazing, professional and talented people that I am proud to call them my family now. I gained a lot of unique skills like patience, problem solving and critical thinking in addition to a lot of new techniques. I learned to never give up and most importantly passion in cancer research as without hope there is no meaningful research. My work in the lab was affected by Covid-19 but with the help of Dr. Porter, the RAs, the grad students and the mentor network I was able to overcome many challenges. We are a true family that stands which each other. Joining the Porter Lab helped me better understand the importance of cancer research and value the importance of hard work. My experience in the lab helped realize that I want to pursue a career that involves research in the future. I learned something new every minute I spent in the lab and this experience increases my desire in continuing to learn more and more every day. I am very grateful for my experience in the Porter Lab and I am proud to call myself a Porter Lab rat.”

The Role of Tuberin in DNA Damage Repair During Cell Proliferation

The cell cycle contains DNA damage checkpoints that delay mitotic progression and allow for DNA repair before cell division. DNA damage can be caused by radiation, drugs, and other processes which lead to cellular mutations and carcinogenesis. The tumour suppressor protein p53 is activated in the presence of DNA damage. It induces apoptosis or cell cycle arrest which allows cells to repair themselves. Tuberin (TSC2), another tumour suppressor protein, regulates the G2/M transition in the cell cycle and negatively regulates protein synthesis and cell growth. Mutations in tuberin can lead to the multisystem autosomal dominant disease known as tuberous sclerosis (TSC). Previously, our lab has shown that Tuberin regulates mitotic onset through cellular localization of the G2/M Cyclin, Cyclin B1. My project focuses on the Tuberin/Cyclin B1 complex in relation to G2/M arrest and DNA damage repair. In this study, we will overexpress Tuberin-WT and Tuberin clinical mutants in NIH-3T3 (mouse) and U2OS (human) p53 wild type cells. Etoposide, a topoisomerase II drug, will be used to induce DNA damage. Cells will then be analyzed by flow cytometry, TUNEL assay, and western blot to assess their cell cycle profile, apoptotic levels, and protein expression. Using CRISPR-Cas9 technology, a NIH-3T3 null TSC2 cell line will be created to clarify the role of Tuberin during DNA repair. Preliminary results have determined that 4μM of etoposide treatment at 8 hours arrests 50% of NIH-3T3 cells at G2/M. This project will provide greater insight into DNA damage induced carcinogenesis, TSC, and other proliferative diseases.

“Joining the Porter lab in my first year has been one of the most rewarding experiences of my undergraduate career. Not only was I able to learn about advanced scientific techniques in class, but I was able to put them in action throughout my own research in lab. The problem-solving, critical thinking, and hands-on techniques I learned during my thesis are skills that I will carry forever. I was also lucky enough to make life-long friendships during my research journey. There were some challenging moments, but I could always rely on my lab team members and mentors to guide me. The last three years have been an amazing experience, and I wish the best for the next team of brilliant undergrads.”

The Essentiality of Spy1 in Cooperation with Hepatocellular Carcinoma Drivers to Promote Tumour Formation

Liver cancer is the fourth leading cause of cancer-related deaths worldwide, standing at an estimation of 800,000 deaths annually. Among the various subtypes, hepatocellular carcinoma (HCC) is the most common primary liver malignancy. HCC is known to develop through a series of genetic and epigenetic alterations of proto-oncogenes and tumour suppressor genes in the liver environment. These changes ultimately lead to the malignant transformation of hepatocytes, the primary cells of the liver. Various HCC drivers are known to cause disruption of cellular pathways and promote tumour formation. Importantly, several cell cycle mediators cause dysregulation, thereby stimulating tumour formation and progression. The cyclin-like protein Spy1, promotes cell cycle progression and overrides apoptosis. Recent reports have detected increased levels of Spy1 in human HCC, which directly correlates to severity of the disease and poor prognosis. We hypothesize that Spy1 plays a critical role along with hepatocellular carcinoma drivers to advance tumour development.  We will test the essentiality of Spy1 on HCC development by first investigating potential gRNAs to use for Spy1 knockout in vivo in the liver, for hydrodynamic tail vein injections in wildtype mice. Simultaneously, in vitro testing of HCC cells (HepG2) will study the importance of Spy1 in HCC cell characteristics in combination with HCC drivers such as c-Myc, p53, and 𝛽-catenin. This project will assist in understanding the essentiality of Spy1 in HCC, which may reveal insight into the molecular mechanism of the tumour suppressors and proto-oncogenes connected with this subset of liver cancer. 

“Applying to Porter lab during the start of my second year was genuinely a spontaneous decision but it turned out to the best decision I’ve made in my life. Before joining the lab, I had no idea what I wanted to do after graduating but after stepping foot into Porter Lab, I was able to unlock my passion for research. My time in the lab learning new skills, meeting peers from various walks of life and finally executing my undergraduate thesis made me realize the potential for myself to pursue a career in research. I have learned an incredible amount of information during the years and the passion to learn continues to grow. I learned to appreciate research as not just a career but as a lifelong passion which at times is underapprecaited. It may be time consuming, require numerous amounts of trials and errors and force you to step out of your comfort zone but the journey is worth it. I was privileged to work alongside such an amazing team of research assistants and colleagues that supported me at every point of my journey. Seeing how much investment is made into cancer research, and how much of an impact every single attribute plays into the final goal, makes me realize that I chose the correct path and I can’t wait to delve more into it. The Porter lab has wonderful incoming students that will continue to make the lab’s name shine, can’t wait to see what more they can accomplish. I will always cheer on for the lab and its members!!. “

2021 Undergraduate Theses – Antonio and Jared

fidalgo — Tue, 04 May 2021 16:29:19 +0000

Two of our awesome undergraduate students are sharing their projects and experiences during their time in our lab. Check here to read the first post about our 2021 undergraduate students.

Antonio Roye-Azar

Evaluation of the cell cycle regulation in Tuberin knockout cell lines

Tuberin (TSC2) and its binding partner hamartin (TSC1) are tumour suppressor proteins that regulate cell growth and cell proliferation through the mTOR pathway and cell cycle, respectively. Mutations in these genes are related to many proliferative diseases such as Tuberous Sclerosis Complex (TSC) and cancers. Our lab has determined a novel role for TSC2 as a cell cycle regulator at the G2/M transition. TSC2 can bind to the G2/M cyclin, Cyclin B1 (CycB1), and retain this cyclin in the cytoplasm, delaying mitotic onset. My project focuses on further understanding the TSC2/CycB1 complex formation and the consequences of this interaction during the progression of the cell cycle. I hypothesized that cells lacking TSC2 will have an improper G2/M transition causing morphological and proliferative abnormalities. To test my hypothesis, I used TSC1 or/and TSC2 knockout HEK293 cell lines acquired from the TSC Biosample Repository to measure the rate of the G2/M transition, cell proliferation, and cell morphology. To further understand the TSC2 knockout consequences, the cells were synchronized using a thymidine block (S phase blocker) and the cell cycle profiles were analyzed following release. I found that KO cells exhibit a change in morphology and size when examined under a confocal microscope. When the cell cycle profiles were analyzed with flow cytometry, the double knockout cells are significantly decreased in the dead cell population under normal conditions while TSC1 KO cells are significantly decreased in the S phase population. Additionally, KO cells were prone to serum starvation-induced apoptosis and were never able to fully synchronize using a double thymidine block; however, a 2mM final concentration of thymidine provided a more uniform block than 0.2mM. Moreover, when analyzed with flow cytometry, the WT HEK293 were significantly larger than the KO cells and dilute BrdU faster than KO cells suggesting faster proliferation. The results of my project will shed light on the role of TSC2 as a cell cycle regulator and clarify its role as a tumour suppressor protein.

“Joining the Porter Lab has been one of the most rewarding experiences of my academic career. Before joining the Porter Lab, I wasn’t really sure in which direction I wanted to take my academic career. However, I knew that I liked science and I knew that this would be a great opportunity to explore my interests and work with highly skilled academics doing cutting-edge research. What I didn’t know, was that, on top of being mentored and pushed to achieve excellence, I would develop amazing connections with people who I’m very proud to call my colleagues. It has been a rollercoaster of emotions, to say the least. From the long frustrating days trying to troubleshoot experiments, to the many laughs and fun nights shared with lab mates, it has all been such an integral part of my university experience. I’m incredibly grateful for all of the support and guidance I have received over the years and especially during my thesis. The Porter Lab has been a place for my knowledge and interests to grow unrestricted and for me to be able to give back to the scientific community, even if only in a minute way. Even as I write this and look back on all of my successes and failures, I know it was accomplished by standing on the shoulders of giants with the unwavering support of everyone in the lab. I don’t know what the future holds for me, but I will forever look back on this experience with fond memories and will always consider myself a Porter lab rat.”

Jared Miller

Spy1: A Closer Look at its Binding Mutants

Spy1 (SPEEDY; RINGO) has been previously identified as a novel cell-cycle regulator that is elevated in breast cancer, promoting progression through the cell cycle and increased cell proliferation. This atypical, cyclin-like protein is capable of binding directly to Cdk1 and Cdk2, activating their kinase activity to override cell-cycle checkpoints; however, the mechanism by which Spy1A activates Cdks has been resolved using only Cdk2 and Spy1A, a single-member within the Spy1 family. Spy1 can also promote the degradation of p27, a Cdk2 inhibitor, through its phosphorylation and binding between p27 and the Spy1-Cdk2 complex. Amino acid residues on Spy1 have been identified to be important for p27 and Cdk2 binding and Cdk2 activation. There is interest in determining whether these residues are conserved across Spy1 family members and further resolving the impact these mutations have on Spy1 function. We expressed these mutants in HEK-293 cells in the presence and absence of p27 to examine the effects of these mutants on cell growth and binding to its effectors. To investigate sequence homology between the different Spy1 family members, their amino acid sequences were aligned and compared using the Basic Local Alignment Search Tool (BLAST). Future work can be done to examine whether the function of these sites is conserved across the family members. The fulfillment of these objectives will provide a better understanding of Spy1 to target it in cancer treatments.

“I have nothing but great things to say about my two years in Dr. Porter’s lab. Whether it was interacting with fellow undergrads, the RA’s, or Dr. Porter herself, the entire team was welcoming from the beginning and always displayed a willingness to help. Despite the fact that Covid-19 disrupted my chances of returning to the lab, I managed to stay connected to the entire team throughout my thesis experience. Since joining Dr. Porter’s lab in my third year, I have acquired so many great skills that I will be taking with me on my future endeavors. While I am sad to be leaving the University of Windsor and such a great group of devoted researchers, I know my experiences will have a long-lasting impact on myself as a student, researcher, and future medical professional. Thank you everyone in the lab for your kindness, guidance, and encouragement.”

2021 Undergraduate Theses – Kate, Kim, and Sami

fidalgo — Mon, 03 May 2021 16:40:38 +0000

Our 2021 undergraduate students have done wonderful work presenting their research results at the Ontario Biology Day, UWill Discover, and our Undergraduate Research Colloquium. Our lab has faced many challenges during this past year, the pandemic has tested our limits in many ways, but we are keeping up with our outstanding research projects. We had 5 undergrad students, some of them worked from home, while others were able to go to the lab, in one way or another we made sure they had the best research experience possible under these circumstances. As you can see below and here, they really enjoyed their time in our group.

Katie Zuccato

The Effects of Dexamethasone on Breast Cancer and Chemotherapeutic Treatment

Breast cancer is the second leading cause of cancer deaths in Canadian women. A particular aggressive subset of breast cancer, triple-negative breast cancer (TNBC), accounts for approximately 15% of all breast cancer diagnoses. Unlike other subtypes, it does not express well-defined molecular targets, such as hormone receptors, that allow for targeted treatment. TNBCs are therefore treated with cocktails of potent but general cytotoxic chemical therapies which have been known to cause harsh side effects such as hypersensitivity, nausea, and vomiting. These side effects are combatted by the addition of the synthetic glucocorticoid, Dexamethasone (Dex) to the treatment regime. It is of clinical significance to understand the impact of Dex on breast cancer behavior and how it influences chemotherapy treatment and patient response. Through the completion of a literature review, the role of Dex in promoting breast cancer progression through cell survival and metastasis was evaluated. Understanding the effects of glucocorticoids, such as Dex, on breast cancer cell biology and their interaction with chemotherapeutic agents can lead to alternative therapeutic strategies and improved patient outcomes.

“Joining the Porter Lab in my second year was a highlight of my university experience. Not only did I get to meet such talented and inspiring people, but I got to learn several skills that are applicable to many life situations. Even though my thesis project was unconventional due to the pandemic, I had such amazing mentors that ensured I learned as much as I would have if I had been able to physically be in the lab. I am grateful for the experience and the people that helped me succeed, and I am honored to have witnessed the many successes of my fellow lab members. I am looking forward to continuing to hear about the amazing work the Porter Lab Team is doing.”

Katie will be starting Optometry School at Waterloo University next fall.

Kim Nguyen

The Tumour Suppressor Tuberin: Coordinating DNA Damage and Mitotic Onset

The cell cycle is a series of events that involve cell growth and division. It is regulated by cyclins and CDKs, as well as checkpoints that control its progression. Tuberin (TSC2 gene) is a tumour suppressor protein that controls cell growth and proliferation. Tuberin mutations are implicated in several cancers and diseases such as tuberous sclerosis complex (TSC), a multisystem genetic disorder that is characterized by the growth of hamartomas. Previous data from our lab has demonstrated tuberin’s role in controlling mitotic onset by binding to the G2/M cyclin, Cyclin B1 according to nutrient availability. My project explores the role of the tuberin in the G2/M transition during DNA damage repair. DNA damage can result from events such as radiation, X-ray, and reactive oxygen species which activates the G2/M checkpoint to restrict mitotic onset and ensure sufficient time to repair damaged DNA. In our studies, NIH3T3 p53 positive cells were transfected with TSC2-WT or clinical TSC2 mutants that present low affinity for cyclin B1 (C696Y) or lack a GAP domain. The transfected cells were treated with etoposide, a topoisomerase II drug that induces double-stranded DNA breaks during the G2/M transition. The changes in cell cycle phases were analyzed using flow cytometry. Our preliminary results have determined that following treatment with 2µM of etoposide at 8 hours, there is a significant decrease in the % of cells at the G2/M transition in C696Y transfected cells vs. those overexpressing tuberin. This project will help expand the role of tuberin during the mitotic onset and clarify mechanisms of proliferative diseases such as TSC and cancers.

“My experience in the Porter Lab has been one that has been truly life-changing! By deciding to embark on this journey, I had the opportunity to really see science in action! Throughout our undergraduate career, we learn about these concepts and different experiments from listening to lectures and reading textbooks. Being able to go into the lab and perform my own experiments has shed light to truly how hard and rewarding science can be. The feeling of finally seeing the hard work pay off from repeating and troubleshooting an experiment countless times is one that is so fulfilling! While starting my 420 and performing my own research had its challenging moments, I was constantly supported and reassured by the rest of our lab every step of the way. My experience as a Porter Lab Rat has given me the opportunity to meet and work with so many amazing people. From Dr. Porter to the RAs, grad students, and other undergrads, everyone has been so passionate and dedicated in our pursuit of scientific advancement and it has been an honor being able to work with everyone. Joining the Porter Lab and doing a 420 has been such a gratifying learning experience for me and I highly encourage others to do the same!”

Kim is the recipient of the 1st place 10 Min Presentation for Cell and Genetics at the Ontario Biology Day. She will continue her research in our lab while coursing the 5th year undergrad studies at the University of Windsor.

Sami Alrashed

The impact of HA-labeled nanoparticles on CD44+ Tumour Initiating Cells in Glioblastoma

Glioblastoma (GBM) is the most aggressive brain tumor with a median survival of only 15 months. Despite decades of research, GBM continues to pose a great therapy challenge due to its extreme genetic and phenotypic heterogeneity. The pools of stem-like tumour initiating cells (TICs) maintain and recapitulate the heterogeneity of GBM; their ability to self-renew fuels resistance to treatment and tumour recurrence. Furthermore, successful treatment of GBM is hindered by the poor penetration of the available therapeutics through a tightly regulated blood-brain barrier (BBB). Conjugated polymer nanoparticles (CPNs) are a class of nanotechnology of potential novel application for GBM treatment. CPNs can penetrate the BBB and can also be encapsulated with drugs for cargo delivery, in addition to functionalizing their surface with ligands complementary to protein receptors expressed on the glioma cell surface. In collaboration with the Rondeau- Gagné group, my project focuses on a diketopyrrolopyrrole (DPP)-based CPNs labeled with a fluorescently tagged hyaluronic acid (HA). HA is the primary ligand of the CD44 receptor present on the surface of TICs. CD44 overexpression is implicated in GBM progression and correlates with poor patient survival. Using the human U-251 MG GBM cells and patient-derived primary lines, I will investigate the effects of CPNs on CD44 signaling and stem cell properties of TICs in vitro as well as in vivo, using a Zebrafish model. Collectively, these results will not only further validate the CPN system as a potential future anti-GBM therapy but also contribute to a better understanding of CD44 biology in GBM.

“When I joined the Porter Lab at the start of my second year, I had no idea that it would end up being both the highlight of my undergraduate journey and the most rewarding experience I’ve had to date. Before joining the lab, I wasn’t familiar with the research at all, but by the end of my fourth year, it’s safe to say that research has become a genuine, lifelong passion of mine. My time in the lab has made it clear that I want to pursue a career involving research in the future. It’s been an opportunity for me to both learn and develop many valuable skills and traits like working as a team, problem-solving, and patience, as well as responding to failure and keeping determination. I’ve learned an incredible amount of information during my time in the lab and the desire to learn only continues to grow. Research can be very time-consuming, and it does require a lot of dedication and effort, but I can confidently say that I’d repeat my experience in the Porter Lab over and over again with no complaints. The time I spent here has felt like my family away from home. A big part of my research experience revolved around building relationships with my mentors, graduate students, and fellow lab mates, and it has been a surreal experience. These are relationships that I’ll be beyond grateful for forever; I’ll always remember being a Porter Lab Rat. I’ve caught myself thinking about how amazing it is to be able to contribute to cancer research and work towards elevating the health of our community. Even if my actions seem minuscule, I recognize now that these little steps are all vital in understanding the bigger picture. The Porter Lab is in good hands – I know every individual member is going to make some very important contributions to the field of research, and I hope I can be there to celebrate these accomplishments with them all!”

Sami is the recipient of 1st place undergraduate speaker at UWill Discover conference, NSERC USRA 2020 summer, and 2021 summer. He will continue his research in our lab during the summer.

Happy Holidays 2020

fidalgo — Tue, 22 Dec 2020 21:16:51 +0000

Producer and Director – Our fabulous graduate student Isabelle Hinch.

Porter Lab in quarantine – part 5

fidalgo — Thu, 21 May 2020 14:00:06 +0000

All about barns and pigeons! A DIY by Emily, a Master student in our lab.

Check out:

Part 1 Part 2 Part 3 Part 4 videos!