Advancing Bladder Cancer Research with Mass Spectrometry: A FeMS Interview with Marta Relvas-Santos

Marta Relvas-Santos, of the Portuguese Oncology Institute of Porto in Portugal, is a winner of the Females in Mass Spectrometry (FeMS) Empowerment Award, which is presented to members of the group that demonstrate excellence both in their work and their support of women in science.

FeMS is a community-led group that has developed a network of support for women working in mass spectrometry. For years, FeMS has supported events all over the world where women in mass spectrometry can gather and discuss their findings and career experiences.

Relvas-Santos graduated in Biochemistry from the University of Porto in 2016 and is now pursuing a PhD in Biomedical Sciences there. Her research focuses on using mass spectrometry to study cancer glycosylation dynamics and identify glycoproteins for potential clinical applications as biomarkers or therapies. Her interest in glycomics and glycoproteomics started during her master's degree, and since then, she has applied mass spectrometry to advance understanding of glycans' roles in health and disease. LCGC International recently sat down with Relvas-Santos to discuss her career, how mass spectrometry relates to her field, the award, and her work with FeMS.

What is the focus of your Ph.D. research?

I have been focusing on the study of bladder cancer, which faces limited therapeutic options. It is well known that solid tumors, namely bladder tumors, experience remodeling in cell surface glycosylation, starting to express glycosignatures linked to cancer progression and poor prognosis. Hence, the focus on glycobiology. Even so, in oncology, we search for cancer specificity. Therefore, it’s advantageous to focus on the glycoproteome instead of zooming in only on glycosidic structures, due to their expression in some healthy tracts or non-tumoral conditions. With this context in mind, my objective has been to discover molecular candidates that may serve as improved biomarkers or therapeutic targets. I don’t know a better technique than MS to address this analytical problem. In this case, we have been applying nano-liquid chromatography (nanoLC) coupled to HRMS, to comprehensively perceive the glycoproteome and the dynamics throughout cancer development. Hopefully, during the next year, the work I have been developing will be published, so stay tuned.

What are examples of work you have helped with during your time at the Instituto Português de Oncologia do Porto (IPO-Porto)?

The Portuguese Oncology Institute of Porto is the largest cancer institute in Portugal, and it is devoted to patient care, teaching in oncology, and cancer research through its research center, which I am part of. The research center brings together academic researchers, clinicians, patients, medical associations, supported by several scientific platforms, a Clinical Research Unit, and a biobank. This has been an opportunity to grow in such a multidisciplinary institution that addresses basic, translational, and clinical research. Our research aim is to respond to the patients’ needs, and the proximity to the hospital reminds us of the responsibility we have in the research we do every day.

I have collaborated on the glycoproteogenomics characterization of the glycoprotein CD44 in bladder cancer. This is a protein with several isoforms resulting from alternative splicing, which together with glycosylation, gives origin to several glycoproteoforms. I will share the links for those interested in learning more. In the end, we have identified the signatures with clinal relevance in bladder cancer (1). The group I belong has also been developing vaccine candidates for cancer immunotherapy (2,3). I have collaborated in the optimization of glycopeptide synthesis and separation, and the characterization of different glycoconjugates. It has been an interesting challenge, that has also brought me enthusiasm. Finally, let me mention a project addressing a triad in bladder cancer: tumor microenvironment, specifically the effect of oxygen and glucose, abnormal glycosylation, and aggressiveness (4). We have performed several omics on a wide array of cell models derived from different cancer types, glycoengineered cell models, healthy and tumor tissues.

Could you elaborate a bit more on the techniques you use and the application areas?

For glycomics analysis, we perform nanoLC coupled to orbitrap mass spectrometer, using a C18 reverse-phase and either collision-induced dissociation (CID) or higher energy collisional dissociation (HCD) fragmentation methods. For a quick and general view, matrix assisted laser desorption ionization-time of flight-mass spectrometry (MALDI-TOF-MS) is also an on-demand option. Usually, we derivatize the glycans by permethylation. I am a fan of the advantages of this labelling. We also receive good separations and analysis results with a standard C18 column for nanoLC, for both neutral and sialylated glycans. In the field of glycoproteomics, we have established lectin-affinity chromatography for the enrichment of proteins or peptides carrying specific glycan signatures and subcellular fractioning. The cell membrane is our focus in biomarker/target discovery, so that is important. We also often use formalin-fixed paraffin-embedded tissues, as these are the most available samples in cancer research, and we have also all the protocols optimized for this type of sample. In terms of analysis, we employ nanoLC-MS/MS, using Orbitrap mass spectrometers. We have mainly used data-dependent analysis and depending on the study’s objectives, we use HCD, CID, or EThcD fragmentation. Using mass spectrometry to address glycoproteins is a magical, but tough, technique.

When did you become a member of FeMS? How has the group impacted your career?

I became a member of the Females in Mass Spectrometry community earlier this year. I came across this network on LinkedIn, and I became an avid follower of the group publications and the stage it provided for the fascinating work developed by my colleagues working with mass spectrometry. Firstly, since I was a master’s student, I have been inspired by other women working in mass spectrometry-based glycobiology. So, having found an international network of women working in mass spectrometry both in my field and other applications is cool. Specifically, the different career mentorship initiatives and panels of discussion where we can share experiences got my attention.

Tell us a little bit about the Empowerment Award and what it means to you.

I am delighted to be one of the awardees of the Empowerment Award. It will financially support me to attend a meeting in my field of research, enabling me to interact and exchange ideas with other researchers and expand my knowledge in mass spectrometry. I am very thankful for having the opportunity to study and do a postgraduation in what I love. Most importantly, receiving this award has brought me more confidence and encouraged me to pursue the field of mass spectrometry, especially to study glycosylation.

Who is a mentor that has impacted your career? How did you meet them, and what made them a great mentor?

I must mention two people, my supervisors José Alexandre Ferreira and André Silva. Two men who have always demonstrated their support for women’s scientific growth. Ferreira is an enthusiast for the study of sugars, and I owe him my passion for glycobiology and the application of mass spectrometry methods to address the glycosylation dynamics. He is also an inspiration because of his scientific career, hard-working example, and scientific vision. Silva introduced me to proteomics and mass spectrometry. His capacity to tackle different analytical challenges is admirable. To conclude, they are great mentors for also providing me with a variety of skills, mentoring my scientific career from the beginning, and trusting in my capabilities.

What are pieces of career advice that your mentors gave you that helped grow your career?

I can start by mentioning something that Ferreira said to me some years ago: a Ph.D. is not only a period for professional growth, but also for personal growth. Not only a period to learn bench work, but also a myriad of other skills. If we can develop different skills, that is important, as it complements our background as Ph.D. students. I keep that message with me because doing a Ph.D. is a tough challenge. I went through a period where I had more anxiety, and now looking back, I can observe my personal and professional growth. To be calmer and less anxious, do not complicate too much, and address challenges one step at a time. I think these are the pieces of advice I have heard more often from my mentors. They also used to give me other advice: not to focus only on the problems, but instead on the solutions and have a more positive perspective.

What advice would you give to other women looking to advance their careers in analytical science?

My advice is to work hard and be always up to date with what our peers are publishing and how science is evolving. Besides literature, I suggest attending seminars, courses, and meetings, which are great for exchanging ideas with other researchers and getting the latest research developments first-hand. I would say that understanding the fundamentals will always be relevant and necessary to plan research ahead and realize novel avenues. S o do not devalue that. For women, in particular, I would say that it is necessary to believe in our abilities, pursue our dreams, and trust that we can be anything we want to be and do! I also think that having good supervisors or mentors who give value to usindependently of our sex or age and help us progress and succeed is important. Find the right people.

How can the scientific community better support women in science?

We have some good initiatives in Portugal to support women in science. “L'Oréal Portugal Medals of Honor for Women in Science” supports the most promising scientists at the beginning of their career after PhD to pursue their studies. Another initiative is promoted by the Portuguese Foundation for Science and Technology, the “RESTART program,” which envisages supporting researchers who have taken parental leave, including adoption. Despite not being completely aware, I believe initiatives like this happen in other countries. I would suggest more initiatives that recognize and finance the research of young women. I would say that around 30-40 years old is probably the more delicate phase for work-life balance and when all the support that enables women to continue doing science is great. Additionally, initiatives like the “International Day of Women and Girls in Science” that also engage with the public, including children and teenagers, are necessary to better prepare the next generation for gender equality and to raise awareness that scientific research is a possibility for women and that there are great women’s minds achieving great things.

About the Interviewee

Marta Relvas-Santos graduated in Biochemistry from the University of Porto in 2016. Currently, she is a PhD student in Biomedical Sciences at the School of Biomedical Sciences, also at the University of Porto, and she is interested in applying mass spectrometry techniques to study the dynamics of cancer glycosylation and discovering glycoprotein candidates for biomarker/therapeutic use in clinical settings. Her enthusiasm for the disciplines of glycomics and glycoproteomics began during her master’s degree. And since then, she has been exploring several applications of mass spectrometry, that contribute to deepening our knowledge of the role played by glycans in health and disease.

References

(1) Gaiteiro, C.; Soares, J.; Relvas-Santos, M.; et al. Glycoproteogenomics Characterizes the CD44 Splicing Code Associated with Bladder Cancer Invasion. Theranostics 2022, 12 (7), 3150–3177. DOI: 10.7150/thno.67409

(2) Freitas, R.; Miranda, A.; Ferreira, D.; Relvas-Santos, M.; et al. A Multivalent CD44 Glycoconjugate Vaccine Candidate for Cancer Immunotherapy. J. Control. Release 2024, 367, 540–556. DOI: 10.1016/j.jconrel.2024.01.065

(3) Freitas, R.; Ferreira, E.; Miranda, A.; et al. Targeted and Self-Adjuvated Nanoglycovaccine Candidate for Cancer Immunotherapy. ACS Nano 2024, 18 (14), 10088–10103. DOI: 10.1021/acsnano.3c12487

(4) Peixoto, A.; Ferreira, D.; Miranda, A.; Relvas-Santos, M.; et al. Multilevel Plasticity and Altered Glycosylation Drive Aggressiveness in Hypoxic and Glucose-Deprived Bladder Cancer Cells. BioRxiv 2023. DOI: 10.1101/2023.10.21.561355