 Hello, welcome to the OncoTarget YouTube channel. This week I'll be introducing you to Dr. Kelber. He works with the Department of Biology at California State University Northridge in Northridge, California. Please enjoy. Good morning. My name is Jonathan Kelber and I'm an assistant professor at the Cal State University Northridge campus in the Los Angeles area. I've been here for approximately five years and prior to this I did my training in San Diego at the Salk Institute and UCSD's Morris Cancer Center. My laboratory here at Cal State Northridge is an NIH funded laboratory and we study cancer progression, metastasis and therapy responsiveness or lack thereof in epithelial tumors. In particular we're interested in a better understanding of how the growth factor, transforming growth factor, TGF-beta, has its paradoxical effects on epithelial cancers such as pancreatic cancer. On this particular project published in OncoTarget in 2016 titled, A Novel Method for RNA Extraction from FFPE samples reveals significant differences in biomarker expression between orthotopic and subcutaneous pancreatic cancer patient-derived xenografts. We have the opportunity to collaborate with two industry partners, Mark Brown and Bob Dobler at Claremont BioSolutions and Robert Hoffman at Anticancer Inc. as well as an academic collaborator of ours, Michael Bouvet at the UCSD Morris Cancer Center. So as I'm sure the readers of OncoTarget can appreciate pancreatic cancer is a devastating disease. The five-year survival rate for patients is just under 7% and as such it's important for basic translational and clinical researchers to get a better molecular profile of this disease so that the patients can be parsed out into good and poor outcome categories as well as those who are responsive or less responsive to available therapies. And so to do this, one approach is to retrospectively profile the disease. And I'll talk a little bit about how that played a big part in the impetus for our current study. Additionally, pancreatic cancer has a very well-developed stromal microenvironment that's very complex and so this adds additional challenges to understanding and diagnosing or treating the disease. It also underscores the importance for looking at pancreatic cancer cells as they grow or they progress within the proper microenvironment such as the pancreas. And so we have several interests in our lab as I mentioned earlier but one of the long-term goals in our laboratory is to profile the proteomes and transcriptomes of patient samples in a retrospective fashion as I earlier mentioned. We want to compare these molecular profiles to linked clinical data and we believe that by doing so we can really understand the complexities that are presented in a disease such as pancreatic cancer. I think the importance of this is even further underscored or emphasized by the 2016 Cancer Moonshot Bluen Ribbon Panel recommendations. One of the 10 recommendations in quote is to mind past patient data to predict future patient outcomes. So it's very important that we're as a research community entering into these efforts. Now one of the challenges however with this retrospective analysis has been to really recover sufficient high quality molecular information from these banked samples. RNA proteins, these types of macromolecules are highly degraded and this is normally due to the fixation and embedding process. Formalin is used to fix and paraffin is used to embed patient samples. This is very helpful obviously in clinical practice for pathologists. This is the standard practice but for the basic science researchers who are trying to access this kind of trapped molecular information it makes it very difficult. So to tackle this issue we hypothesized that a tool known as the micro-homogenizer that had been previously developed by Claremont BioSolutions for tissue disruption could be introduced into ongoing or current methods that were available for extracting RNA from FFPE samples. And that by doing so we could recover more high quality RNA. In fact as we developed this protocol and as it's now published we see almost a three-fold increase in RNA quantity from the same starting sample size and these RNA molecules that are recovered are larger in length suggesting that we're actually accessing some of the more trapped or previously inaccessible, longer, less degraded RNA molecules and this of course has major implications for downstream next generation sequencing since we really need to get access to as much of the transcriptome as possible. So we went a step further and we validated this method with samples provided to us from our clinical collaborators in San Diego and they provided us with xenograft samples from patient or human pancreatic cancer cells and that were either grown in the subcutaneous or in orthotopic microenvironment for us to then compare with the same cell samples grown in vitro. Now we specifically chose one particular cell line to include in this study known as FG, Nakamura at all in 2007 published a study using these cells as well as some more metastatic derivatives of these cells and they profiled the gene expression of these cells grown in different microenvironments, the same microenvironments that we were able to use in our study. So we felt like this would be a very good control for us to see how the gene expression of genes in our study was matching or otherwise not matching what had been previously reported. And so what we found very strikingly was that only when we use the microhomogenizer based protocol that we developed could we faithfully reproduce the gene expression patterns for some well-known regulators of pancreatic cancer. We went a step further then and we looked at some more novel genes that have been more recently characterized to contribute to pancreatic cancer progression or onset and we now have really valuable information about how those genes are changing in the context of where these tumor cells are grown, whether they're grown and expanded in vitro or in vivo subcutaneously or orthotopically. For example, one of the genes that my lab studies, PICT1, it's a new kinase that we've been working on for about the last seven or eight years. We now know its expression is almost a hundredfold more when those cells are expanded in the subcutaneous in vivo microenvironment. And so this really drives home the importance of understanding really a context-dependent role for what we may otherwise report across the board as an important biomarker of therapeutic target in this cancer. The biggest challenge in this study was early on as we were trying to identify where within current FFPE RNA extraction protocols this micro-homogenizer was best introduced. First authors, Malakaya Hoover and Ibis Damian, they worked tirelessly to really test different permutations of what eventually became our published protocol. And while we're certainly aware that we're going to see probably some new, even improved iterations of this protocol, we believe that what we've established is certainly a great starting point. I also know that our collaborators at Claremont BioSolutions are continuing to develop their technology. Most recently they have converted the single-use kind of single-well micro-homogenizer and expanded that into a 96-well plate format. So this protocol can be applied really in a high-throughput fashion now. As I mentioned earlier, we're certainly interested in scaling up this work. And so we're working with our collaborators to now apply this on a large scale so that we can get some statistically relevant information about molecular markers in what otherwise is a very deadly malignancy. We're also taking a few other approaches using proteomics and bioinformatics to predict new diagnostic and therapeutic biomarkers. And so we're hopeful that as we've done in the past with Peak 1, showing that it regulates pancreatic cancer progression and therapy resistance, we're going to over the next year or two be able to report on several new diagnostic and therapeutic targets in this disease. Thank you. By clicking the link below, you can learn more about the research discussed in the interview from Cover Paper of Volume 8, Issue 4. Thank you for watching.