3rd EurocanPlatform Translational Research Course
Post-translational modification (PTM) profiling – a novel tool for mapping the PTM landscape in cancer
Dr Yifat Merbl – Weizmann Institute of Science, Rehovot, Israel
During my talk I basically described the system that we now have for profiling changes in post-translational modifications which are changes that occur in proteins after they are being translated. This system allows for looking at different stages of cancer, different cancer subtypes and so on, and characterise and analyse them in the way that was done previously for gene expression changes but at the level of the protein modification. So the main message, and this is why people really reacted to it was the fact that we really need to look at that level of regulation because these changes in protein modifications and the resulting outcome of how they change the function of proteins is really determining how a protein behaves and functions in the tissue.
With regards to cancer and the ability to get molecular profiling and understanding of different stages and conditions it is really important to look at these changes, at protein modifications, because they dictate the effect of the proteins and their function in the malfunctioning tissue. Now this is an area of investigation which is largely limited by current proteomic analysis, so the standard way of looking at changes in protein modifications is done by mass spectrometry which is mainly focussed at changes at the level of phosphorylation but there are plenty of other types of protein modifications like ubiquitin and other ubiquitin-like modifications, glycosylation and so on, that would also affect the activity of proteins, their localisation in the cell, their binding partners and so on and in order to really analyse changes at that level and these types of modifications we need to think of additional tools.
What significance does this have for changing clinical practice?
The PTM profiling, PTM stands for post-translational modification and this PTM profiling approach, combined with the ability to analyse clinical samples offer a new dimension for analysing the state of tissues, of tumour biopsies, of other liquid biopsies, really trying to look at what is different at the different clinical manifestations such as disease states, response to treatment and so on based on the signature of the protein modifications. This is very important because in the current day and age we are trying to do so just by looking at the gene expression pattern which is very indirect to what actually translates into protein activity in the tissues. Specifically I can give one example of multiple myeloma, for example we have a drug which is called Velcade, bortezomib, which is the treatment for these patients and in some cases patients do not respond to it. Now if you try to address this question, since this drug is inhibiting the proteasome, the major machinery which is responsible for degradation of proteins, you would want to focus at the level of protein degradation and, for example, there is a protein modification which is called polyubiquitin which targets protein to this degradation. So by having a way to probe the activities of polyubiquitination you are more likely to identify or at least you should also look at that level and not just only at the level of gene expression in order to identify the subpopulation that may respond well to the drug.
What can you tell us about PTM profiling in lung adenocarcinoma?
This is something that we really started very recently and I’m happy to say, very excited to say, that we actually can get tumour samples, very small tumour biopsies, from patients that have different underlying mutations. Specifically, as you probably know, lung cancer is the leading cause for mortality in both men and women worldwide and among these the adenocarcinoma is the subpopulation that is most prevalent. The type of underlying mutations basically right now determines the type of treatment that one can get. So there is really a need to identify other sources and other types of pathway that are involved in KRAS driven adenocarcinoma and the reason is that currently there is no targeted therapy that these patients that have a KRAS mutation respond to. So what we have tried to do using this PTM profiling approach is to identify the signature of the downstream protein modifications and specifically differentiate between the EGFR mutated population to KRAS mutated population and see whether we can see a signature that is unique to each of them. I’m very happy to say that even though it’s on a very small subset right now we’re able to identify significant differences between them which may enable us to decipher not only underlying pathways that are involved in this disease but also in the future stratify patients in order to give them specific drugs and also maybe probe the response to different drugs.
What are the next steps for this area of work?
I really hope that this area of work would really open up, will be opened up, for investigation by many different groups and many different types of cancer. One can think of looking and studying the level of protein modification in the same manner that people have done so far for gene expression type of analysis and once you understand the potential of that you understand that it’s more than just one lab that needs to do that. But I do believe that looking at that level of regulation which is essentially where the activity happens, the level of protein modification is where we actually see the effects of the drugs, of the changes in gene expression, of the changes in the mutation and so on, so you really want to incorporate that level of regulation together with the proteomics, together with the genomics, in order to generate a comprehensive view of what happens in cancer and, by the way, it’s not just in cancer, it’s in any type of disease and also in homeostasis.
What are the eventual benefits of this work?
Looking primarily at patients, we’re talking about capabilities in biomarker discoveries, companion diagnostics to follow the response to drugs. This actually can be utilised for drug development purposes because you can actually look ahead of time and test what is the effect of a specific drug on the enzymatic activity of the tissue. You can potentially identify novel targets for therapeutics that were not discovered before but I think there are also very basic underlying questions in fundamental biology like what is the natural variability of PTMs in different people? How is the genetic make-up affecting this natural variability? How much of the population if you compare population level to single cell analysis, we don’t have even basic understanding of whether the post-translational modifications really need to occur in every cell or whether there can be major effects only by affecting subpopulations. There is a whole field of research which is the cross-talk between different post-translational modifications because we have more than 200 different types of these protein modifications in cells and we’re really in our infancy in terms of our ability to analyse the cross-talk between different modifications and how they affect each other and whether they are really communicating with one another in terms of protein regulation.
What is your take home message?
What I would like to really make people understand, and I think we are taught that at the very early stages of our career and any basic course in biology, which is that the protein can be modified in many different ways and although we usually regard the mRNA, for example, as the template to what can occur, I would actually argue that the protein itself, the sequence of amino acids is just the backbone and once a protein is modified in one way or another this is what will ultimately dictate its function in the cell and its cellular localisation and so on and so forth. So we should bear that in mind when we even conceptually talk about a protein like p53, for example, and remember that there are many different subpopulations, some of them are functional, some of them are not and this inevitably in the coming years will light our understanding and go deeper in the ability to understand regulatory mechanisms in the cell by looking at the proteins, not as proteins that are uniquely defined by their name but rather as having many different subpopulations that are functionally different.