Biotechnology

Protein analyses provide new insight into cancer cells

Analyses of proteins in single cells—mass spectrometry—is a rapidly developing technology that provides new insight into the structure of cancer cells and can be used to clarify whether chemotherapy works in a cell.

Laserudskæring af celler i vævsprøver styret med mikroskop åbner nye muligheder for proteinanalyser af høj kvalitet. Fra venstre lektor på DTU Erwin Schoof og seniorforsker på Rigshospitalet Pia Klausen ser på . Foto: Marie Bentzon Sørensen. — Laser cutting of cells in a tissue sample using a microscope enables high-quality protein analyses. From left: Associate professor at DTU Erwin Schoof and senior researcher Pia Klausen, Rigshospitalet. Photo: Marie Bentzon.

Thursday 26 October 2023

Peter Aagaard Brixen

Very few people think about the billions of cells and proteins in our body in their everyday life. But in DTU’s mass spectrometry unit Proteomics Core—a platform for analysing proteins—researchers focus on the content of proteins in every single cell when analysing biological material and tissue sections from patients. Among other sources, the samples come from Rigshospitalet’s Department of Pathology, which sends samples from patients with liver, pancreatic, brain, and blood cancer to gain greater insight into the role proteins play in the individual diseases.

“A number of hospitals in Denmark and Norway order analyses of cell samples. One of the things they look at is whether patterns in the protein variation can be used to extract more knowledge from their biobanks, where millions of tissue sections and preparations containing biological material from patients are currently stored,” says Marie Vestergaard Lukassen, Facility Manager at Proteomics Core.
The mass spectrometry analyses in Proteomics Core take place in a new building at DTU, where instruments the size of photocopiers and refrigerators are lined up in a laboratory room.

Before the biological samples are placed in the machines, they must be prepared. The protein content of the sample is extracted from cells and tissue and then subjected to a process where the laboratory technicians use a digestive enzyme to break down the proteins. The enzymes act as a kind of chemical scissors that cut the proteins into smaller pieces, making it easier to see which proteins are in the sample and how much of them is present.

Cuts protein into peptides

To perform the analyses, the laboratory technicians take a sample of the proteins and place them in a tube with a digestive enzyme. For larger samples, the mixture is heated and shaken to ensure the enzyme comes into contact with the proteins and starts cutting them into smaller parts. In single-cell mass spectrometry, however, the laboratory technicians work with much smaller amounts of material, and therefore this entire process can take place in a droplet, which does not offer the same physical opportunities to heat and mix.

Proteins are extracted from cells and tissues and then subjected to a process in which the laboratory technicians use a digestive enzyme to break them down into so-called peptides, which are then ready for analysis.

Marie Vestergaard Lukassen works on an instrument that performs high-pressure liquid chromatography, which separates the peptides according to their physical properties before they are analyzed by mass spectrometry.

Once the proteins have been cut into smaller pieces, into so-called peptides, they are ready to be examined with mass spectrometry. Here, the peptides are sprayed out of a thin needle and atomized into gaseous form into the mass spectrometer, where it is possible to determine their size and composition.

By analyzing the smaller building blocks, peptides, that the proteins consist of, researchers can gain knowledge about which proteins are present and which are regulated in different disease states. This can provide insight into how the proteins function in the body and how they affect human health.

Once the enzyme has cut the proteins, the laboratory technicians now have a mixture of smaller protein pieces called peptides. These peptides are like small LEGO bricks that are now ready to be studied using mass spectrometry, which measures the different sizes and the electrical charge of the peptides. Describing the peptides requires breaking them into smaller pieces in the instruments to see the differences in their amino acid structure. By analysing the different sizes, researchers can identify the peptides and find out which proteins they come from and how much of them is present. This helps scientists understand the proteins’ function in the body and how they affect human health.

The latest research project in the area focuses on cells from patients suffering from acute myeloid leukaemia, AML, and is a collaboration between Erwin Schoof, Associate Professor at DTU, and Rigshospitalet. Erwin Schoof has received DKK 10 million from the Lundbeck Foundation to establish a research team that will identify what determines the viability of individual blood cells and whether it is possible to specifically develop cancer stem cells for the treatment of patients with leukaemia. The work is based on a combination of cell samples from patients, big data, and advanced computational calculation methods.

Analysing single cells

Since launching the mass spectrometry platform in 2018, DTU has expanded its activities and seen a user increase of 30 per cent. This corresponds to around 80 users in 2022, spread across 140 different projects. It can take one to three weeks before the analysis results are ready.

The good results seen when performing analyses on small cell samples have, in particular, attracted new partners and customers. Until 2018, researchers could only perform analyses if they had larger cell samples, which could consist of both healthy tissue and cells from a tumour. Today, they can perform analyses at single-cell level, enabling specific insight into smaller tissue samples.

Laser-cutting

The latest development in the field is a technique where researchers and pathologists use a laser and a microscope to cut out a small group of cells.

“By using laser dissection, we can cut out suspicious or diseased cells and get very precise answers via mass spectrometry. In this way, we can find out what happens in the cancer cells, but also read their interaction with surrounding cells, and perhaps find new treatments,” says Pia Klausen, Senior Researcher at Rigshospitalet’s Department of Pathology.

Proteins warn about cancer

Rigshospitalet’s Department of Pathology is the largest of its kind in Denmark and performs a number of analyses using DTU’s mass spectrometry platform. In an ongoing research project, researchers are examining preserved samples from patients who have had pancreatic cancer. By using a laser to cut out cancer cells from tissue samples and the surrounding cells in a separate sample, the researchers are examining whether they can use mass spectrometry to identify proteins that can provide new insight into the development of cancer.

“We hope to be able to use laser dissection and mass spectrometry to find special proteins that can be used, e.g., to predict how patients will respond to a certain treatment. It is currently not possible to find these signatures by examining a person’s complete set of genes using genomic analysis. Cancer is a disease in which mutations play a major role. And although it is possible to map mutations using genomic analysis, they will only give an indication of what can happen. The proteins will tell directly what has happened. In addition, it is the proteins that doctors are trying to target with cancer drugs, and we hope to find new therapeutic targets with our research,” explains Pia Klausen.

In collaboration with DTU, the researchers are currently analysing samples from patients with pancreatic, lung and brain cancer. All three are types of cancer with a low survival rate and few treatment options.

Facts

Proteomics platform

DTU Bioengineering’s platform Proteomics Core is the only mass spectrometry platform of its kind in the Øresund Region which is open to all customers.

Today, mass spectrometry has become a standard method in the proteomics field and is widely accepted as a tool for identifying proteins and for quantification in complex biological samples. The method is also used for disease diagnosis, food analysis, environmental monitoring, and pharmaceutical research.

DTU is currently performing mass spectrometry analyses for projects coming in from the majority of Danish universities. The tasks often arise from research done in hospital departments at Rigshospitalet, Herlev Hospital, and Bispebjerg Hospital.

The proteomics platform also performs a number of analyses for foreign universities and for primarily Danish companies, both established companies and small start-ups.

Topic

Biotechnology

DTU has a strong focus on the use of biotechnology within pharma and health, where biology and technology are utilized for early diagnosis of diseases as well as the development of new medicines and treatment methods.

Biotechnology is part of the life science area and research into this area is so extensive that it alone accounts for a third of the university's scientific publications. The latest national figures show that the biotech industry's total annual growth in this area is 6 %. This is three times higher than the average growth in the private business community.

Statistics Denmark's latest figures show a 30 % increase in the number of graduates with a long scientific education within technical sciences who are employed in the sector.

Read more in our special topic on biotechnology.

Contact

Marie Vestergaard Lukassen Proteomics Core Facility Manager

Erwin Schoof Associate Professor Phone: +45 45252059 Mobile: 93 51 06 77