Arun Parmar – 2020 Studentship Recipient

Arun Parmar is entering his fourth year of Health Sciences at McMaster University.

Arun Parmar

What does receiving this award mean to you?

I am sincerely grateful and honoured to receive the Brain Tumour Foundation of Canada Studentship. As I have progressed in my academic career, I have realized the importance of placing myself in an environment that helps satisfy my curiosity, peaks my interests, and facilitates my learning. Not only would this studentship help fund a project that I proudly call my own, it will also help me pursue a career in brain tumor research and fulfill my desire to improve our understanding of brain metastasis and the lives of those facing it. In the Singh laboratory, I am lucky to be immersed in a nurturing, team-oriented environment and surrounded by equally passionate peers who support and encourage my personal development.

I would like to thank the Brain Tumour Foundation of Canada, the Taite Boomer Foundation, and the generosity of patients and donors for making this possible.

How did you learn about the studentship program?

For the past several years, the Singh Laboratory has been actively involved with the Brain Tumour Foundation of Canada to raise awareness and improve the lives of those suffering from brain tumours. We strongly believe in the values and commitment of this foundation and strive to support in as many ways as we can. This year we will be attending the #VirtualBrainTumourWalk and taking part in the 27 Canadians Challenge to #EndBrainTumours. I strongly encourage anyone interested to register and take part in this challenge with us.

What led you to apply?

Being part of the Singh Laboratory for the past year, I have grown to admire and adopt a translational research approach. As a young student, this studentship helps alleviate some of the financial barriers with my project to allow me to focus on answering my research question and get the most out of this experience. With this support, I hope to further develop my knowledge of brain cancer and contribute important research towards informing and developing viable therapy options for patients with aggressive tumours.

What is your project, and why did you choose it?

For my project, I am establishing an organotypic brain slice culture system to explore the behaviour of brain tissue and its interaction with metastatic tumour cells. Pilot experiments have revealed an essential role for a gene called human leukocyte antigen (HLA-G) in promoting cancer stem cell self-renewal and proliferation. Using this model, I aim to validate the role of HLA-G knockout on the survival of cancer stem cells in a three-dimensional system that maintains the structure and architecture of the brain. We believe knocking out HLA-G will attenuate cancer stem cell survival and ultimately their ability to establish brain metastasis (BM), which will provide new therapeutic opportunities of targeting HLA-G in human BM in the clinic.

This project allows me to combine my interests in visualization techniques and cellular biology to better understand the key molecular mechanisms responsible for brain metastasis development.

What do you hope to learn through it?

While I aim to increase my knowledge of brain metastasis, I am equally motivated to develop and improve upon my research and interpersonal skills over these next two terms. I am excited to continue working in a field I am incredibly interested in and with similarly passionate scientists. I know that further development of such skills and my experience in the Singh Laboratory, as provided by this studentship, will not only follow me as I progress in my academic career, but also in my personal life.

Why do you feel it’s important to advance study of brain tumours?

Brain metastases are the most common brain tumour in adults and are ten times more likely to develop than primary tumours. The brain is widely believed to be the most unique aspect of every individual: its multiplex structural design of neural connectivity is what shapes our reality. There are so many questions that have yet to be asked and there is still so much that is currently unknown. While this may be overwhelming to some, to me this is very exciting. Having seen the impact that this devastating diagnosis has on patients and families, I am committed to help make a difference in those affected by brain tumours.

What personal experiences led you to focus on brain tumours?

As someone with a family member affected by brain cancer, I have seen the toll it can have on someone and how complex its onset can be. In first year of university, I joined a McMaster club called Student’s Advancing Brain Cancer Research (SABCR). As part of this club, my goal is to offer students exciting new events and opportunities to learn and get involved with brain cancer research. After inviting members from the Singh Laboratory to come present at a research event, I was blown away by all their research and dedication. I knew an opportunity in this lab would be an ideal foundation for a passionate student such as myself and I am grateful to have received it.

December 2020 midterm update:

I am happy to report on the progress that I have made over the summer term for my project. The first component of my project was to establish a viable organotypic mouse brain slice culture system. After successfully optimizing our slicing technique and co-culture procedure, I was able to successfully seed patient-derived lung tumour cells in the mouse brain tissue. After staining the brain tissue for haematoxylin and eosin, this assay allowed me to visualize the migration capacity of the human tumour cells and their interactions with the surrounding tissue. After evaluation of binding partners of HLA-G and reviewing the literature for the mechanism behind HLAG expression and tumor cell survival, we identified several different proteins and possible pathways: SPAG9, CRK, AKT, B-Actin, ERK1/2 and STAT3 signaling. For these pathways, I assisted in the completion of western blot analysis to determine the effect of knockout of HLA-G on the protein expression of these genes. The result of this investigation showed that SPAG9 and CRK expression was increased following the knockout of HLA-G. These promising results has prompted further inquiry to determine the role of these two genes as a therapeutic option for patients with brain metastasis. At the end of this term, I was also able to successfully clone three guide RNAs for CRK and SPAG9 onto CRISPR-Cas9 lentiviral vectors. The next steps for this project will be to use these knockout vectors (KO) to generate stable KO lines. I can also use my established organotypic brain slice culture system to visualize the effect of this KO on tumour cell survival in the brain. Some of the data I have collected over the summer will be used for a publication that is currently in progress, where I am a co-author. This manuscript is titled, “HLA-G regulates the pre-metastatic stage of brain metastasis formation”.

Final Report- October 2021

During the first summer of my project, I was successfully able to establish a viable organotypic mouse brain slice culture system. By seeding patient-derived lung tumour cells on the mouse brain tissue, I was able to visualize and evaluate the migration and survival capacity of the human tumour cells with the surrounding brain tissue. The results from this system prompted me to review the literature for potential protein binding partners of a gene called HLA-G and downstream signalling pathways. For these pathways, I assisted in the completion of western blot analysis to determine the effect of knockout of HLA-G on these binding partners. These results showed promising novel evidence for the increased expression of two proteins (SPAG9 and CRK) following the knockout of HLA-G. To validate that the tumour cells use HLA-G through this signalling pathway to survive in the brain, I successfully cloned guide RNAs for these proteins, which were used to generate knockout vectors using CRISPR-Cas9 technology. Furthermore, this past summer I was also able to work on an existing mouse model in the Singh Lab. In this model, human patient-derived non-small cell lung cancer cells were injected into the mouse lungs. Using CRISPR activation technology, we were able to activate genes in a lung tumor cell pool, which resulted in the lung tumor cells migrating to the brain. After scanning and quantifying tumour bioluminescence, I assisted in the extraction of mouse lung and brain tissue, and subsequent DNA isolation. Currently, the DNA from these tissues is being sequenced. The results of this experiment will help to identify genes that drive lung to brain metastasis in humans. The next steps for this project would be to validate these genes as they may provide evidence for promising new therapies for lung to brain metastasis patients.

I am sincerely grateful and honoured to receive the Brain Tumour Foundation of Canada studentship. Before I received this prestigious award, I had so many questions regarding brain tumours and brain metastasis formation. Although it appears I have many more questions now, I am excited to continue to answer these questions as I pursue a career as a clinician scientist. By alleviating many of the financial restraints, this studentship allowed me to carry out a project that I proudly called my own. Due to restraints such as this, I recognize that unfortunately not many students are given the same opportunity and for this reason, I still quite humbled to be a recipient of this studentship.

Despite the numerous obstacles I faced completing my project, the idea that someday this research I conducted may help improve the lives of brain tumour patients kept me motivated. Although I was able to further develop my knowledge of brain cancer and research techniques, I am more proud of the skills I have learned as a young, student researcher along the way. I look forward to continuing to grow as a person and develop these skills (e.g., critical thinking, observation, planning, etc.) in my future career. As part of the Singh Laboratory, I learned to adopt a translational research approach. Working on my own project alongside equally passionate and dedicated scientists was an experience I will never forget. I would like to express my sincerest appreciation to the Brain Tumour Foundation of Canada and the Taite Boomer Foundation for your generosity and contribution to brain tumour research.

Some of the data I have collected throughout my project will be used for a publication that is currently in progress, where I am a co-author. This manuscript is titled, “Identification of an HLA-G/SPAG9/STAT3 axis presents a novel targetable liability in brain metastases”.