Podcasts about RAD51

BUFFALO, NY - December 11, 2024 – A #news feature on the #research paper “Next-generation cell-penetrating antibodies for tumor targeting and RAD51 inhibition” by Rackear et al. was #published in Oncotarget's Volume 15 on November 22, 2024, titled “Advancements in cell-penetrating monoclonal antibody treatment." This new publication by Sai Pallavi Pradeep and Raman Bahal from the Department of Pharmaceutical Sciences at the University of Connecticut highlights significant advancements in monoclonal antibody (mAb) therapies. The focus is on the 3E10 antibody, originally derived from autoimmune mouse studies in systemic lupus erythematosus. Unlike traditional mAbs, which struggle to reach intracellular targets, this cell-penetrating antibody targets cancer cells by addressing a major limitation of current therapies. By targeting RAD51, a key intracellular protein involved in DNA repair, the 3E10 antibody shows great promise for cancer treatment, particularly in cancers with defective DNA repair pathways. mAbs have already changed the landscape of cancer therapy, offering treatments that are more targeted and have fewer side effects compared to chemotherapy. However, current therapies are limited since mAbs only target proteins on the surface of cancer cells. This research pushes the boundaries by demonstrating how 3E10 antibodies can penetrate cells and access their internal molecules. This unique capability expands the potential of mAb therapies and targeted cancer treatments. Different humanized versions of the 3E10 antibody were created and carefully tested. Some versions were particularly effective at blocking RAD51, while others showed promise for carrying other therapeutic molecules like genetic material into the cancer cells. This flexibility means that 3E10 could be used to treat different cancer types and deliver various therapeutic molecules directly into tumor cells. This progress offers exciting new possibilities for treating cancer tumors that are resistant to conventional therapies. In conclusion, the 3E10 antibody's dual function—targeting DNA repair pathways and delivering therapeutic molecules—positions it as a transformative tool in cancer research and targeted cancer treatments. DOI - https://doi.org/10.18632/oncotarget.28674 Correspondence to - Raman Bahal - raman.bahal@uconn.edu Video short - https://www.youtube.com/watch?v=3uMdPvThFHA Sign up for free Altmetric alerts about this article: https://oncotarget.altmetric.com/details/email_updates?id=10.18632%2Foncotarget.28674 Subscribe for free publication alerts from Oncotarget: https://www.oncotarget.com/subscribe/ Keywords - cancer, monoclonal anti-bodies, cell penetration, nucleic acid delivery, 3E10 About Oncotarget Oncotarget (a primarily oncology-focused, peer-reviewed, open access journal) aims to maximize research impact through insightful peer-review; eliminate borders between specialties by linking different fields of oncology, cancer research and biomedical sciences; and foster application of basic and clinical science. Oncotarget is indexed and archived by PubMed/Medline, PubMed Central, Scopus, EMBASE, META (Chan Zuckerberg Initiative) (2018-2022), and Dimensions (Digital Science). To learn more about Oncotarget, please visit https://www.oncotarget.com and connect with us: Facebook - https://www.facebook.com/Oncotarget/ X - https://twitter.com/oncotarget Instagram - https://www.instagram.com/oncotargetjrnl/ YouTube - https://www.youtube.com/@OncotargetJournal LinkedIn - https://www.linkedin.com/company/oncotarget Pinterest - https://www.pinterest.com/oncotarget/ Reddit - https://www.reddit.com/user/Oncotarget/ Spotify - https://open.spotify.com/show/0gRwT6BqYWJzxzmjPJwtVh Media Contact MEDIA@IMPACTJOURNALS.COM 18009220957

Cancer research has made remarkable progress in recent years, with monoclonal antibody (mAb) therapy emerging as one of the most promising advancements. These treatments are designed to precisely target cancer cells, offering a more focused approach that helps patients fight different malignancies with fewer side effects compared to traditional chemotherapy. Despite this progress, a major challenge remains: targeting cancer-related molecules inside cells rather than on the surface, which has been the main focus of available mAb therapies until now. This is where the groundbreaking research in the paper “Next-generation cell-penetrating antibodies for tumor targeting and RAD51 inhibition,” published in Volume 15 of Oncotarget on October 1, 2024, comes into play. Full blog - https://www.oncotarget.org/2024/10/24/next-generation-antibodies-for-cancer-therapy/ Paper DOI - https://doi.org/10.18632/oncotarget.28651 Correspondence to - Peter M. Glazer - peter.glazer@yale.edu Video short - https://www.youtube.com/watch?v=sTHjJ0Qq0YQ Sign up for free Altmetric alerts about this article - https://oncotarget.altmetric.com/details/email_updates?id=10.18632%2Foncotarget.28651 Subscribe for free publication alerts from Oncotarget - https://www.oncotarget.com/subscribe/ Keywords - cancer, 3E10, cell penetration, nucleic acid binding, nucleic acid delivery, RAD51 About Oncotarget Oncotarget (a primarily oncology-focused, peer-reviewed, open access journal) aims to maximize research impact through insightful peer-review; eliminate borders between specialties by linking different fields of oncology, cancer research and biomedical sciences; and foster application of basic and clinical science. Oncotarget is indexed and archived by PubMed/Medline, PubMed Central, Scopus, EMBASE, META (Chan Zuckerberg Initiative) (2018-2022), and Dimensions (Digital Science). To learn more about Oncotarget, please visit https://www.oncotarget.com and connect with us: Facebook - https://www.facebook.com/Oncotarget/ X - https://twitter.com/oncotarget Instagram - https://www.instagram.com/oncotargetjrnl/ YouTube - https://www.youtube.com/@OncotargetJournal LinkedIn - https://www.linkedin.com/company/oncotarget Pinterest - https://www.pinterest.com/oncotarget/ Reddit - https://www.reddit.com/user/Oncotarget/ Spotify - https://open.spotify.com/show/0gRwT6BqYWJzxzmjPJwtVh MEDIA@IMPACTJOURNALS.COM

BUFFALO, NY- October 21, 2024 – A new #research paper was #published in Oncotarget's Volume 15 on October 1, 2024, entitled, “Next-generation cell-penetrating antibodies for tumor targeting and RAD51 inhibition.” As highlighted in the abstract, monoclonal antibody therapies for cancer have shown extraordinary clinical success in recent years. However, these strategies are primarily limited to targeting specific cell surface antigens, despite many disease targets being located intracellularly. In their paper, researchers Madison Rackear, Elias Quijano, Zaira Ianniello, Daniel A. Colón-Ríos, Adam Krysztofiak, Rashed Abdullah, Yanfeng Liu, Faye A. Rogers, Dale L. Ludwig, Rohini Dwivedi, Franziska Bleichert, and Peter M. Glazer from the Departments of Therapeutic Radiology and Genetics at Yale University School of Medicine, Gennao Bio, and the Department of Molecular Biophysics and Biochemistry at Yale University report on the humanization of the full-length, nucleic acid-binding monoclonal lupus-derived autoantibody 3E10, which exhibits a novel mechanism for cell penetration and tumor-specific targeting. The authors compare humanized variants of 3E10 and demonstrate that cell uptake relies on the nucleoside transporter ENT2. They also find that faster cell uptake and superior in vivo tumor targeting are associated with higher affinity nucleic acid binding. “We show that one human variant retains the ability of the parental 3E10 to bind RAD51, serving as a synthetically lethal inhibitor of homology-directed repair in vitro.” DOI - https://doi.org/10.18632/oncotarget.28651 Correspondence to - Peter M. Glazer - peter.glazer@yale.edu Video short - https://www.youtube.com/watch?v=sTHjJ0Qq0YQ Sign up for free Altmetric alerts about this article - https://oncotarget.altmetric.com/details/email_updates?id=10.18632%2Foncotarget.28651 Subscribe for free publication alerts from Oncotarget - https://www.oncotarget.com/subscribe/ Keywords - cancer, 3E10, cell penetration, nucleic acid binding, nucleic acid delivery, RAD51 About Oncotarget Oncotarget (a primarily oncology-focused, peer-reviewed, open access journal) aims to maximize research impact through insightful peer-review; eliminate borders between specialties by linking different fields of oncology, cancer research and biomedical sciences; and foster application of basic and clinical science. Oncotarget is indexed and archived by PubMed/Medline, PubMed Central, Scopus, EMBASE, META (Chan Zuckerberg Initiative) (2018-2022), and Dimensions (Digital Science). To learn more about Oncotarget, please visit https://www.oncotarget.com and connect with us: Facebook - https://www.facebook.com/Oncotarget/ X - https://twitter.com/oncotarget Instagram - https://www.instagram.com/oncotargetjrnl/ YouTube - https://www.youtube.com/@OncotargetJournal LinkedIn - https://www.linkedin.com/company/oncotarget Pinterest - https://www.pinterest.com/oncotarget/ Reddit - https://www.reddit.com/user/Oncotarget/ Spotify - https://open.spotify.com/show/0gRwT6BqYWJzxzmjPJwtVh MEDIA@IMPACTJOURNALS.COM

La doble hélice es la estructura canónica del ADN, descubierta por Watson y Crick en 1953. Pero no es la única. Puede haber hélices triples y cuádruples, entre otras disposiciones, que han adquirido una notable relevancia en los últimos años porque parecen estar implicadas en procesos de regulación de la expresión de los genes y servir para el desarrollo de fármacos que bloqueen la expresión de un gen determinado. Hemos entrevistado a Carlos González y Miguel Garavís, investigadores del Instituto de Química Física Blas cabrera del CSIC que emplean la resonancia magnética nuclear para observar estas curiosas estructuras de nuestro material genético. investigadores del CSIC que estudian estas estructuras. Hemos informado del hallazgo un nuevo mecanismo que impide que el ADN se triplique durante la división celular. Se trata de una proteína llamada RAD51 descubierta por investigadores del CNIO. Ángela Bonachera nos ha hablado de una investigación del Instituto de Ciencias de Materiales del CSIC que ha logrado monitorizar con rayos X la temperatura de nanopartículas empleadas contra el cáncer en el interior de las células tumorales. Con testimonios de Ana Espinosa, autora principal del artículo. Jesús Puerta nos ha hablado del nacimiento de la física de altas energías y del estado actual de la tecnología de los aceleradores de partículas. Javier Ablanque nos ha llevado en su máquina del tiempo a la segunda guerra mundial para participar en la misión aérea británica que reventó dos presas en el valle del Ruhr, en Alemania, y conocer la física que lo hizo posible. El próximo jueves, 14 de marzo, se celebra el Día Internacional de las Matemáticas y Fernando Blasco nos ha comentado algunas de las muchas actividades previstas para estos días. Hemos reseñado los libros "Cuando el mundo se detiene. Cáncer: del mito a la esperanza", de Juan Fueyo (Ediciones B); “Cementos y hormigones”, de Francisca Puertas Maroto (CSIC-La Catarata); y “Las manos del tiempo. Todo lo que la historia de los relojes cuenta sobre nosotros”, de Rebecca Struthers (geoPlaneta). Escuchar audio

La doble hélice es la estructura canónica del ADN, descubierta por Watson y Crick en 1953. Pero no es la única. Puede haber hélices triples y cuádruples, entre otras disposiciones, que han adquirido una notable relevancia en los últimos años porque parecen estar implicadas en procesos de regulación de la expresión de los genes y servir para el desarrollo de fármacos que bloqueen la expresión de un gen determinado. Hemos entrevistado a Carlos González y Miguel Garavís, investigadores del Instituto de Química Física Blas cabrera del CSIC que emplean la resonancia magnética nuclear para observar estas curiosas estructuras de nuestro material genético. investigadores del CSIC que estudian estas estructuras. Hemos informado del hallazgo un nuevo mecanismo que impide que el ADN se triplique durante la división celular. Se trata de una proteína llamada RAD51 descubierta por investigadores del CNIO. Ángela Bonachera nos ha hablado de una investigación del Instituto de Ciencias de Materiales del CSIC que ha logrado monitorizar con rayos X la temperatura de nanopartículas empleadas contra el cáncer en el interior de las células tumorales. Con testimonios de Ana Espinosa, autora principal del artículo. Jesús Puerta nos ha hablado del nacimiento de la física de altas energías y del estado actual de la tecnología de los aceleradores de partículas. Javier Ablanque nos ha llevado en su máquina del tiempo a la segunda guerra mundial para participar en la misión aérea británica que reventó dos presas en el valle del Ruhr, en Alemania, y conocer la física que lo hizo posible. El próximo jueves, 14 de marzo, se celebra el Día Internacional de las Matemáticas y Fernando Blasco nos ha comentado algunas de las muchas actividades previstas para estos días. Hemos reseñado los libros "Cuando el mundo se detiene. Cáncer: del mito a la esperanza", de Juan Fueyo (Ediciones B); “Cementos y hormigones”, de Francisca Puertas Maroto (CSIC-La Catarata); y “Las manos del tiempo. Todo lo que la historia de los relojes cuenta sobre nosotros”, de Rebecca Struthers (geoPlaneta). Escuchar audio

BUFFALO, NY- January 10, 2024 – A new #editorial paper was #published in Oncotarget's Volume 14 on December 22, 2023, entitled, “One more step toward treatment of PARP inhibitor-resistant ovarian cancers.” Over 80% of ovarian cancer cases experience recurrence, resulting in roughly 12,000 annual deaths in the United States. While targeted therapies like poly (ADPribose) polymerase inhibitors (PARPis) have received FDA approval for both initial and recurrent treatments, extending median progression-free survival for individuals with homologous recombination repair (HRR) deficiency, the emergence of PARPi resistance remains a common challenge among patients. Consequently, addressing resistance to PARPi treatment in ovarian cancer has become a pressing therapeutic dilemma, necessitating innovative strategies. In this editorial, researchers Upasana Ray, Prabhu Thirusangu and Viji Shridhar from Mayo Clinic School of Medicine and Science responded to this unmet need with their current study, which unveiled promising findings related to the Pixatimod (PG545) drug, a sulfated small molecule compound. Engineered with a core structure mimicking heparan sulfate, this compound targets heparanase and heparin binding growth factor (HB-GF) signaling. “Our present study has revealed a previously unknown effect of PG545 in ovarian cancer cells, inducing DNA damage. The investigation unveiled that PG545 induces both single- and double-strand breaks in DNA while also promoting the autophagic degradation of RAD51, a critical DNA repair protein, thereby impeding the homologous recombination repair (HRR) pathway in cancer cells.” DOI - https://doi.org/10.18632/oncotarget.28545 Correspondence to - Viji Shridhar - Shridhar.Vijayalakshmi@mayo.edu Sign up for free Altmetric alerts about this article - https://oncotarget.altmetric.com/details/email_updates?id=10.18632%2Foncotarget.28545 Subscribe for free publication alerts from Oncotarget - https://www.oncotarget.com/subscribe/ Keywords - cancer, ovarian cancer, PARP inhibitors, PG545, DNA damage, cell death About Oncotarget Oncotarget (a primarily oncology-focused, peer-reviewed, open access journal) aims to maximize research impact through insightful peer-review; eliminate borders between specialties by linking different fields of oncology, cancer research and biomedical sciences; and foster application of basic and clinical science. To learn more about Oncotarget, please visit https://www.oncotarget.com and connect with us: SoundCloud - https://soundcloud.com/oncotarget Facebook - https://www.facebook.com/Oncotarget/ X - https://twitter.com/oncotarget Instagram - https://www.instagram.com/oncotargetjrnl/ YouTube - https://www.youtube.com/@OncotargetJournal LinkedIn - https://www.linkedin.com/company/oncotarget Pinterest - https://www.pinterest.com/oncotarget/ Reddit - https://www.reddit.com/user/Oncotarget/ Media Contact MEDIA@IMPACTJOURNALS.COM 18009220957

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.04.12.536536v1?rss=1 Authors: Di Biagi, L., Malacaria, E., Aiello, F. A., Valenzisi, P., Marozzi, G., Franchitto, A., Pichierri, P. Abstract: Replication gaps can arise as a consequence of perturbed DNA replication and their accumulation might undermine the stability of the genome. Loss of RAD52, a protein involved in the regulation of fork reversal, promotes accumulation of parental ssDNA gaps during replication perturbation. Here, we demonstrate that this is due to the engagement of Polalpha downstream of the extensive degradation of perturbed replication forks after their reversal, and is not dependent on PrimPol. Polalpha is hyper-recruited at parental ssDNA in the absence of RAD52, and this recruitment is dependent on fork reversal enzymes and RAD51. Of note, we report that the interaction between Polalpha and RAD51 is stimulated by RAD52 inhibition, and Polalpha-dependent gap accumulation requires nucleation of RAD51 suggesting that it occurs downstream strand invasion. Altogether, our data indicate that RAD51-Polalpha-dependent repriming is essential to promote fork restart and limit DNA damage accumulation when RAD52 function is disabled. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2023.03.03.531058v1?rss=1 Authors: Miller, J. M., Prange, S., Ji, H., Rau, A. R., Butova, N. L., Lutter, A., Chung, H., Merigliano, C., Rawal, C. C., McVey, M., Chiolo, I. Abstract: Pericentromeric heterochromatin is highly enriched for repetitive sequences prone to aberrant recombination. Previous studies showed that homologous recombination (HR) repair is uniquely regulated in this domain to enable 'safe' repair while preventing aberrant recombination. In Drosophila cells, DNA double-strand breaks (DSBs) relocalize to the nuclear periphery through nuclear actin-driven directed motions before recruiting the strand invasion protein Rad51 and completing HR. End-joining (EJ) repair also occurs with high frequency in heterochromatin of fly tissues, but how different EJ pathways operate in heterochromatin remains uncharacterized. Here, we induce DSBs in single euchromatic and heterochromatic sites using the DR-white reporter and I-SceI expression in spermatogonia. We detect higher frequency of HR repair in heterochromatic insertions, relative to euchromatin. Sequencing of repair outcomes reveals the use of distinct EJ pathways across different euchromatic and heterochromatic sites. Interestingly, synthesis-dependent michrohomology-mediated end joining (SD-MMEJ) appears differentially regulated in the two domains, with a preferential use of motifs close to the cut site in heterochromatin relative to euchromatin, resulting in smaller deletions. Together, these studies establish a new approach to study repair outcomes in fly tissues, and support the conclusion that heterochromatin uses more HR and less mutagenic EJ repair relative to euchromatin. Copy rights belong to original authors. Visit the link for more info Podcast created by Paper Player, LLC

Dr. Shannon Westin discusses germline genetic testing in gastrointestinal cancer with Heather Hampel and Dr. Matthew B. Yurgelun.   TRANSCRIPT The guest on this podcast episode has no disclosures to declare. Shannon Westin: Hello, everyone, and welcome to another episode of JCO After Hours. This is our podcast where we get down in the nitty-gritty of articles that are published in the Journal of Clinical Oncology. I am your fearless leader and host, Shannon Westin, the Social Media Editor of the Journal of Clinical Oncology, as well as Professor of Gynecologic Oncology at The University of Texas, MD Anderson Cancer Center. And I am very excited to bring two guests in today to discuss a review article that was published in a special series on, 'Precision Medicine and Immunotherapy in GI Malignancies,' back in June of 2022, and this is, 'Point/Counterpoint: Is It Time for Universal Germline Genetic Testing for All GI Cancers?' And please note that our participants have noted no Conflict of Interest. So, without further ado, let me welcome our guests. First is Heather Hampel. She is a Cancer Genetic Counselor, and the Associate Director in the Division of Clinical Cancer Genomics, and a professor in the Department of Medical Oncology and Therapeutics Research at the City of Hope National Medical Center. Welcome, Heather. Heather Hampel: Thanks so much for having me. Shannon Westin: We're also accompanied by Dr. Matthew Yurgelun, he is a Senior Physician in Medical Oncology at the Dana-Farber Cancer Institute, the Director of the Lynch Syndrome Center, and Assistant Professor of Medicine at Harvard Medical School. Welcome. Dr. Matthew Yurgelun: Thanks for having me. Shannon Westin: And we have all decided we were going by first names. So, audience, don't be alarmed. Okay, let's get right into it. So, this is a really great review. I learned a ton and I think, you know, just to kind of get back to basics, I think we've been seeing an increase in the use of germline genetic testing across a number of different cancer types. As I mentioned, I'm a gynecologic oncologist, certainly this is something we're doing for patients with ovarian cancer. What are the reasons this has become so widespread across all cancer types? Heather Hampel: Matt and I probably agree on this one. I will, but I'll say you a couple of reasons, and see if Matt has any to add. I think that 2013 marked a major turning point in the field of cancer genetics for a couple of reasons. One was; the advent of next-generation sequencing, so that we could do multiple genes at the same time for a lower cost. The other was that that was the year the Supreme Court struck down the patent on BRCA1 and BRCA2, which allowed lots of different competitors into the market to offer sort of these pan-cancer panels, including, BRCA1 and BRCA2, among other genes. And the price has dropped precipitously since then, giving better access for patients. The competition, I think, has been good, so that a lot of the laboratories now will offer out-of-pocket maximums of $250. And then, we've seen a lot of research. Because of that, I think, where we've just done pan-cancer panels on different solid tumor cancers, just to determine what the prevalence of mutations is, all of this is sort of leading to, I think, just greater use of germline genetic testing across the board. I don't know. Matt, what do you think? Dr. Matthew Yurgelun: No, I fully agree. This is an example of the more you look, the more you find, and I think we've seen that both in the studies that have been done looking at multi-gene panel testing in virtually any setting across different cancer types and then I think people who use these in clinical practice, whether they are genetic counselors, oncologists, gastroenterologists, gynecologists, primary care physicians, I think as people have become more experienced and more comfortable using them in routine practice—I think it's not an uncommon phenomenon for those of us who use these to find things that were somewhat unexpected, which kind of naturally leads to the question, "Well, what else might I be missing if I'm not doing these tests further and wider?" What's made it a little bit difficult is that this is an example of testing that's become available commercially before we really understood how to use it. And so, we've been figuring a lot of this out kind of on the fly a little bit. Shannon Westin: Yeah. I think it brings up, and not to get too nitty-gritty right from the beginning, but to me, it brings up the whole idea around variance of uncertain significance, right? I think we've really struggled with this on the GYN space, and I don't know how common that is for you all in the colorectal space, but we get answers, we don't know how to tell a patient what to do with that information. And in fact, we've personally seen people get risk-reducing surgeries, probably not appropriately in response to these variants. Dr. Matthew Yurgelun: It's a real phenomenon, and it's the other side of the more you look, the more you find. You know, you end up finding a lot of these variants of uncertain significance. I think we've become a lot more comfortable and maybe even cavalier about them as panel testing has become so widespread. But there are data out there, and not to mention just anecdotes of people who are potentially being harmed by these variants of uncertain significance, as you said, whether it's through unnecessary surgery, whether it's even just psychological burdens and harms that come from the angst of those uncertainties. So, it is important that we be thoughtful about just how to use this technology. Heather Hampel: And really it is, "the more you look, the more you'll find." So, on a panel of about 50 genes, there's about a 30% rate of finding a variant of uncertain significance, increase that panel to 80 genes, you're up to probably a 40 to 50% chance of finding a variant of uncertain significance. A panel of 150 genes, maybe an 80% chance of finding a variant of uncertain significance, and it becomes almost the rule and not the exception. So, this is where genetic counseling becomes really important in terms of having people understand that these are sort of common, and usually, not anything, and setting expectations so that people don't over screen, or overreact, or get in a situation where they're mismanaged. And this is one of the things that Matt and I go back and forth a lot about when you start to think about testing all-comers because if you're testing all-comers, you kind of have to give up pre-test genetic counseling and kind of move to a post-test genetic counseling scenario more for the positives, or people with a strong family history or concerns. And I know Matt worries, and I do too, that that's where we've risked these variants of uncertain significance getting mismanaged, particularly in centers that aren't as used to dealing with cancer genetics. Dr. Matthew Yurgelun: I would just add one more concept on that. We're probably also, in the case of some of these larger panels, dealing even with genes of uncertain significance. At the end of the day, it's the commercial laboratories in many cases that are really setting the agenda on some of these panels as far as choosing which genes to include or not to include, and a lot of these genes are genes where the link to cancer risk is sometimes very preliminary. Shannon Westin: Those are some great points. I think just to kind of take a step back, since the paper's in GI cancers, and I want to make sure we have—we have a mixed audience out there, so I want to make sure we level set. So, can you tell us the current standard of practice for germline testing in GI cancers? What are you looking for specifically? What are some of the things that you know are of certain significance? Heather Hampel: Currently, the NCCN guidelines recommend that all pancreatic cancer patients be offered germline genetic testing. And what is very new in 2022 is that there's now a consideration recommendation that you could consider offering germline genetic testing to all colorectal cancer patients. That is logistically much more challenging than offering germline genetic testing to pancreatic cancer patients because there are so many more of them. And it comes with a page of caveats of things that you need to think about before you would consider offering testing to all colorectal cancer patients. And then, I'd say among the rest of the GI cancers, you're going to be offering testing in cases of early onset, multiple primaries, maybe three affected's on the same side of the family with cancers that could go together in a family, and raise a red flag that there could be hereditary, diffuse gastric cancer comes to mind when you think of the stomach, certainly, polyposis is an indication for testing as well. But for most of the GI organs, you're going to need early age, multiple primaries or some family history. The one clear exception being pancreas, and now, a lot of debate about colorectal. Dr. Matthew Yurgelun: And I would just add to that from some of Heather's own seminal work. Tumor testing is often used to drive a lot of this in day-to-day practice. Certainly, the presence of microsatellite instability, and/or mismatch repair deficiency. This isn't limited to GI cancers, obviously, but it is where we often think about it the most. But any finding of mismatch repair deficiency in microsatellite instability should really strongly trigger strong consideration for germline testing, at least for Lynch syndrome, which is often, at least a way in the door for germline testing a bit further and wider. Shannon Westin: Yeah, that's kind of what we've done in endometrial cancer, and it's definitely a less expensive way of kind of getting at that, and we use those same Amsterdam criteria for the full germline outside of that. But that's really how we've gotten to universal testing for endometrial is using the less expensive protein testing, you know, as a trigger to break down the door. Heather Hampel: But it's not necessarily less expensive anymore. So, that's where things are getting a little challenging. You know, at most hospitals, if you're going to do four immunohistochemistry stains, you're going to take whatever they charge for IHC times four, then you're going to have a reading fee for the pathologist times four, and believe it or not, that adds up pretty quickly, and can become a test that's $1,500+, compared to potentially a $250 germline panel. I find it an odd situation for me to be in, who I've spent 20 years of my career advocating for universal tumor screening for Lynch syndrome. But I do feel that we really need to relook at the cost-effectiveness analysis now that the cost of germline genetic testing has gotten so low, and we need to think about what we're missing with universal tumor screening. So, yes, it will detect most of the cases of Lynch syndrome, and it should detect anyone who could benefit from immune checkpoint blockade therapy. And those are very important points, and I think that's the reason we're never going to go away from tumor screening. But it's not going to detect mutations in any other cancer susceptibility gene. And that's what you risk missing if that's your only approach. Shannon Westin: Those are some great points. Again, bringing those of us that aren't GI experts, up to speed, what's the overall incidence of these germline genetic abnormalities in GI cancers? What are we looking for? Dr. Matthew Yurgelun: I think it depends on which cancer you look at, and it also depends a little bit on even just how you define the prevalence here. Where it's been a little bit difficult, or where it's been kind of moving goal posts is that the panels that we're using in day-to-day practice are getting bigger and bigger, and certainly, the panels that are being used in a lot of the studies that are examining this are getting larger and larger. And as far as the number of genes being tested-- and not surprisingly, as you test more genes, you find more stuff. We make a point in the paper that some of the older studies, these are all still relatively new studies, but some of the older studies that have looked at gene panels of say, 30 genes or fewer, you actually find germline prevalence rates that are maybe 10% or lower across most of the GI tract malignancies. But as you start getting into panels that are 50 genes, 60 genes, 100+ genes, that's when you start getting these prevalence rates that are 15%, 20%, almost across the board. But the prevalence is only part of the story, in my opinion, it's a matter of what you're finding, in addition to how many people you're finding stuff in. Because you know, finding a diagnosis of Lynch syndrome, finding a BRCA1 or BRCA2 abnormality, things that are high penetrance, clearly actionable that we understand reasonably well, I would argue is much more impactful than finding something like a monoallelic MUTYH pathogenic germline variant, which arguably has very little clinical significance for the person themselves, and is honestly, much more common than some of these other things too, and drives up some of these prevalence rates. Heather Hampel: I agree, Matt, completely. And I struggle myself, sometimes, with how people should report out the incidence of mutations in series like this because when you include those MUTYH heterozygotes, you include your APCI 1307Ks, you're padding the numbers, but are you finding something that's going to really make a major management change for that family? I don't know. The one thing though that has stuck with me as I look at this is that it appears that people who meet the criteria for testing aren't much more likely to test positive than people who don't meet criteria for testing. And as a genetic counselor who, you know, loves to take family histories, and for years, liked to believe that if I took a good family history, and assessed it, I could pick who was going to test positive appropriately, it's been a little bit of a dash to the confidence to see rates of positivity that are pretty similar in the patients who don't meet criteria. And I think that's been a challenge for all of us. So, if we could pick these cases well, it would be one thing, but I don't think we can all the time, outside of Lynch syndrome. Lynch syndrome, I think we can pick, to a large degree, through tumor testing more than family history. But the non-lynch genes are much harder to predict based on age of diagnosis, family history, or any other clinical criteria. Dr. Matthew Yurgelun: I totally agree, and for me, that's what moved the needle a handful of years ago with pancreatic cancer in particular, in my own mind, is that it was becoming quite clear that you could find some of these with clinical criteria or even just clinical intuition, but there were a lot that were just being missed. We were looking at age of diagnosis, we were looking at family histories of BRCA-related cancers, and family structures being what they are, in many cases, you know, the fact that pancreatic cancer, even in the setting of some of these germline variants is often diagnosed well after age 50, and often after age 60, or even after age 70, you know, our usual clinical criteria just weren't working. Shannon Westin: I think you kind of already started touching on this, but I'd love to get a little bit more, you know, what would be the additional benefits to doing this universal testing? I think one of the things you just mentioned, like, not being able to completely pick the right people to test. I mean, this is exactly why we started doing this in ovarian cancer is because, more than half of the women that were testing positive BRCA, did not meet the age criteria, they had no family history to note, you know. So, we were missing tons of people. What are the other things we can gain from universal testing? Heather Hampel: I think that's a key one. I think another one that Matt and I agree on is that from that proband there, are all those at-risk family members who get to benefit because of the cascade testing that begins from that first person who gets diagnosed in the family. And those are often unaffected at-risk people who you can really keep from getting cancer in the first place, and make a major impact in their health outcomes. So, not wanting to miss a potential hereditary family, and that ability to get to those at-risk family members, I think is a major benefit. I think the one that hasn't really panned out yet in GI, and I'll leave this to the oncologists in the room, is a change in treatment. But I think it's coming. I hope it's coming; we'll see what research all you oncologists can do. But I think that what moved the needle on ovarian and pancreatic was the fact that we had mutations in the homologous repair deficiency pathway, leading to a change in treatment, and the use of PARP inhibitors that just hasn't born out in GI cancers yet, outside of pancreas. And the one thing that you do see a treatment change for is mismatch repair deficiency, which you can find by doing IHC for the mismatch repair proteins or MSI testing, so, you don't really need the germline panel. In fact, some people with Lynch syndrome don't have an MSI-high tumor, and won't benefit from immune checkpoint blockade therapy. And so, I feel like that's the one benefit that maybe hasn't been fully realized in GI cancer, but I hope it will one day. I don't know. Matt? Dr. Matthew Yurgelun: No, I agree. I think the therapeutic actionability has been one that we've been hoping for more than what we've actually seen in real-world practice, the big exception being PARP inhibitors for pancreatic cancer. But even there, I think so far, the benefits have been maybe a bit more modest than people would've originally hoped for. I agree, I think the therapeutic benefits are ones that we're still trying to work towards. Shannon Westin: Yeah, and you kind of got around this before, and I think this is what we're experiencing in some of the gynecologic malignancies is like, not every gene is created equally. You know, we originally thought, "Oh, any homologous or combination gene will do. We could do PARP for all," and then realized, "Actually, no, it's probably just BRCA, maybe PALB2, RAD51." So, I think it's exactly like what we're seeing in honestly, frankly, across precision medicine, right? Where it's like, not every PI-3-kinase mutation will lead to benefit from a PI-3-kinase inhibitor. So, I think the science got ahead of us, or we got ahead of this science, and so, I do think that that's where the struggle is. Because I think once you've got therapeutic actionability, it becomes a no-brainer. And then, you've already hinted at this, but I just want to be really clear for everyone listening, why shouldn't we do it right now? What's holding us back from universal germline testing for everybody in GI cancers? Dr. Matthew Yurgelun: You know, I think if it were a perfect world, then it would be a no-brainer - test everybody. The finances, as Heather alluded to, are in some ways kind of the least compelling argument against universal testing, that the cost of the germline testing itself has come down tremendously. But it's more than just the cost of the test itself, at least in my opinion. I mean, first of all, I think we've got massive work to do just to figure out the care delivery here. As it stands right now, roughly half of colorectal cancer patients meet criteria for germline testing, putting aside the recent update to the NCCN, which says, "consider testing for all individuals." But even if you just look at colorectal cancer alone, if you expanded germline testing to all-comers with colorectal cancer in the US, that's another 80,000+ new diagnoses per year in the US, who are all over age 50, have mismatch repair proficient tumors, have no striking family history, you know, where the odds of finding something truly actionable in my mind, is exceedingly low. Then you add in all of the other GI cancer types. You add in the literal millions of GI cancer survivors that are out there, and you're talking about bringing this testing to a whole lot of people. On top of that, there's really all of the uncertainties and nuances that come from the testing itself, as we were talking about at the beginning of the chat here. Whether it's variance of uncertain significance, whether it's genes where there's really no link between the gene in which they have a germline variant and the cancer that they were ultimately diagnosed with, or whether they're genes we don't understand or don't have clear-cut management guidelines for, it's really all the unintended consequences in some ways of a lot of what we're doing. And I think too often out there, whether it's from the patients themselves, or us healthcare providers, or combination of the two, there's this misperception that genetic testing is giving black and white answers to what's going on. There's a whole lot of gray here, as far as understanding what needs to be done with this. Yes, if we could test everybody and get them clear-cut answers, and then get their at-risk family members in for testing, the benefits would supersede all of these concerns, but I don't think the real world is as simple and straightforward as that. Heather Hampel: And I find myself just in the struggle between-- when I get in a room with people who discuss this, most people feel like we should be doing it, and the thing that's stopping us is that it's difficult. And that doesn't seem like a good reason to not do this. If it's the right thing to do, I think we have to figure out how to do it. And you think of, you know, Mary Claire King's Lasker Award talk when she talked about offering BRCA1 and BRCA2 testing to all women at age 35, population-based screening. And one of the things she recommended there was, not reporting out variants of uncertain significance. I realize it's a different situation when you're talking about population testing and healthy people, but are we doing more harm than good with reporting out variants of uncertain significance even in cancer patients? Whereas, you know, we could just ask the lab to let us know if anything ever got upgraded and avoid risking mismanagement of patients based on a variant that you know is likely benign. So, I think there are things we can do. We've been working on some of them, I know Matt has done a little work with mainstreaming in pancreatic cancer patients. I did as well, my former job, because that was sort of the first new tumor outside of ovarian where we needed to recommend all patients get seen by cancer genetics. And the wait time often in cancer genetics is too long, given the prognosis for pancreatic cancer patients, we had to kind of come up with new service delivery models. And there's some great data out there, people are using genetic testing kiosks in the waiting room, videos at the oncology clinic. We can get creative, and the trouble is, I think we're learning while we're doing, which is coming full circle in our discussion here, that's a hard way to do things. Dr. Matthew Yurgelun: I would just add on top of that, in my mind, the reason why not to do this, is really why not to do it. I think we can recognize that more germline testing is going to pick up more people with inherited risk to cancer. There's the unintended consequences, and we need to figure out how to deal with them. And as Heather said, just because it's hard, that shouldn't be a reason not to do it. At the same time, I think it's okay for us to recognize that this is where the field is heading, but to also recognize that we don't yet have all the answers, and to say, "Well, let's be thoughtful about it. Let's figure out how to implement these types of things, how to study them," because it's not going to be one size fits all. What works in a major tertiary care academic medical center is going to be very different from what works in other settings and for other populations. What works for English-speaking patients versus non-English-speaking patients is probably going to be very different. And I think there's all sorts of permutations when you start breaking it down like that. And I think it's okay for us to say, "Well, this is where the field seems to be going, but let's really be thoughtful about it and make sure that we're not doing harm in the short term just because we think it makes more sense to just test everybody in a shotgun approach." Shannon Westin: This has been great; I've learned so much. I was like trying to frantically take notes for thoughts of what we're doing in our clinic right now. I just want to thank my two guests. I think that I remain hopeful that we will get here. I think that you all outlined some really clear steps that we need to take to get there. And audience, I just want to thank you all for being here with us. Again, this was a discussion of, ‘Point/Counterpoint: Is It Time for Universal Germline Genetic Testing for All GI Cancers?' Thanks again for joining us on JCO After Hours, and we will see you next time.   The purpose of this podcast is to educate and to inform. This is not a substitute for professional medical care, and is not intended for use in the diagnosis or treatment of individual conditions. Guests on this podcast express their own opinions, experience, and conclusions. Guest statements on the podcast do not express the opinions of ASCO. The mention of any product, service, organization, activity, or therapy, should not be construed as an ASCO endorsement.  

Biomarkers are any measurement in the body that can indicate disease, infection, or effects of the environment. These indicators are commonly used to reveal disease, but they can also be used to support ​​personalized cancer therapy. “In recent years, there has been an increased focus on personalized cancer therapy. One important aspect is the identification of key biomarkers that support a given treatment plan.” Multiple biomarkers have been identified in breast cancers as helpful tools to guide targeted therapies, including the expression of HER2 and estrogen and progesterone receptors. In triple-negative breast cancer (TNBC), a number of distinct biomarkers have given rise to new targeted therapies, such as EGFR antibodies, AKT inhibitors, and PARP inhibitors. In a well-read paper from Immunomedics, Inc. (recently acquired by Gilead Sciences, Inc.), researchers analyzed sacituzumab govitecan (SG; Trodelvy™) in TNBC. The team authored a research paper that was published by Oncotarget in 2020, and entitled, “Predictive biomarkers for sacituzumab govitecan efficacy in Trop-2-expressing triple-negative breast cancer.” To date, this paper has scored an Altmetric Attention score of 48. “Here we examined the potential role of biomarkers in predicting the efficacy of SG.” Sacituzumab govitecan is an antibody-drug conjugate that targets human trophoblast cell-surface antigen-2 (Trop-2)ーwhich is a glycoprotein that is commonly overexpressed in many solid tumor types. “Trop-2 is a 46 KDa transmembrane glycoprotein that is overexpressed on many solid tumor types and is correlated with an overall poor prognosis in patients, making it an attractive target for therapy [7, 9].” The researchers examined a highly invasive and aggressive TNBC cell line (MDA-MB-231), which is not responsive to SG, and compared their findings to TNBC cell lines that are responsive to SG. Their goal was to determine how Trop-2 expression and the homologous recombination repair (HRR) pathway (through Rad51 expression) play roles in protecting the MDA-MB-231 cell line from SG mediated DNA damage. Trop-2 expression in transfected MDA-MB-231 and tumor xenografts were assessed, in vitro and in vivo Rad51 expression and DNA damagewere assessed via western blot, and statistical analysis was carried out for data from in vivo therapy studies. “Trop-2 expression levels as a positive, primary biomarker and HRR proficiency as a secondary, negative biomarker were assessed in vitro and in vivo.” Full blog: https://www.impactjournals.com/journals/blog/oncotarget/biomarker-predicts-treatment-efficacy-in-triple-negative-breast-cancer/ Press release - https://www.oncotarget.com/news/pr/predictive-biomarkers-in-trop-2-expressing-triple-negative-breast-cancer/ Sign up for free Altmetric alerts about this article - https://oncotarget.altmetric.com/details/email_updates?id=10.18632%2Foncotarget.27766 DOI - https://doi.org/10.18632/oncotarget.27766 Full text - https://www.oncotarget.com/article/27766/text/ Correspondence to - Thomas M. Cardillo - tcardillo@immunomedics.com Keywords - sacituzumab govitecan, Trop-2, biomarker, RAD51, triple-negative breast cancer About Oncotarget Oncotarget is a bi-weekly, peer-reviewed, open access biomedical journal covering research on all aspects of oncology. To learn more about Oncotarget, please visit https://www.oncotarget.com or connect with: SoundCloud - https://soundcloud.com/oncotarget Facebook - https://www.facebook.com/Oncotarget/ Twitter - https://twitter.com/oncotarget YouTube - https://www.youtube.com/c/OncotargetYouTube/ LinkedIn - https://www.linkedin.com/company/oncotarget Pinterest - https://www.pinterest.com/oncotarget/ Reddit - https://www.reddit.com/user/Oncotarget/ Oncotarget is published by Impact Journals, LLC please visit https://www.ImpactJournals.com or connect with @ImpactJrnls Media Contact MEDIA@IMPACTJOURNALS.COM 18009220957

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.11.05.369629v1?rss=1 Authors: Belan, O., Barroso, C., Kaczmarczyk, A., Anand, R., Federico, S., O'Reilly, N., Newton, M. D., Maeots, E., Enchev, R. I., Martinez-Perez, E., Rueda, D. S., Boulton, S. J. Abstract: Homologous recombination (HR) is an essential DNA double-strand break (DSBs) repair mechanism frequently inactivated in cancer. During HR, RAD51 forms nucleoprotein filaments on RPA-coated resected DNA and catalyses strand invasion into homologous duplex DNA. How RAD51 displaces RPA and assembles into long HR-proficient filaments remains uncertain. Here, we employ single-molecule imaging to investigate the mechanism of nematode RAD-51 filament growth in the presence of BRC-2 (BRCA2) and RAD-51 paralogs, RFS-1/RIP-1. BRC-2 nucleates RAD-51 on RPA-coated DNA, while RFS-1/RIP-1 acts as a "chaperone" to promote 3' to 5' filament growth via highly dynamic engagement with 5' filament ends. Inhibiting ATPase or mutation in RFS-1 Walker box leads to RFS-1/RIP-1 retention on RAD-51 filaments and hinders growth. rfs-1 Walker box mutants display sensitivity to DNA damage and accumulate RAD-51 complexes non-functional for HR in vivo. Our work reveals the mechanism of RAD-51 nucleation and filament growth in the presence of recombination mediators. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.11.04.367136v1?rss=1 Authors: Roy, U., Kwon, Y., Marie, L., Symington, L., Sung, P., Lisby, M., Greene, E. C. Abstract: Homologous recombination (HR) is essential for the maintenance of genome integrity. Rad51 paralogs fulfill a conserved, but undefined role in HR, and their mutations are associated with increased cancer risk in humans. Here, we use single-molecule imaging to reveal that the Saccharomyces cerevisiae Rad51 paralog complex Rad55-Rad57 promotes the assembly of Rad51 recombinase filaments through transient interactions, providing evidence that it acts as a classical molecular chaperone. Srs2 is an ATP-dependent anti-recombinase that downregulates HR by actively dismantling Rad51 filaments. Contrary to the current model, we find that Rad55-Rad57 does not physically block the movement of Srs2. Instead, Rad55-Rad57 promotes rapid re-assembly of Rad51 filaments after their disruption by Srs2. Our findings support a model in which Rad51 is in flux between free and ssDNA-bound states, the rate of which is dynamically controlled though the opposing actions of Rad55-Rad57 and Srs2. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.08.24.265033v1?rss=1 Authors: Silva-Garcia, M., Bolgi, O., Ross, B., Pilla, E., Vijayalakshmi, K., Killisch, M., Stark, N., Lenz, C., Spitzner, M., Gorrell, M. D., Grade, M., Urlaub, H., Dobbelstein, M., Huber, R., Geiss-Friedlander, R. Abstract: Dipeptidyl peptidase 9 (DPP9) is a serine protease cleaving N-terminal dipeptides preferentially post-proline with (patho)physiological roles in the immune system and cancer. Only few DPP9 substrates are known. Here we identify an association of human DPP9 with the tumour suppressor BRCA2, a key player in repair of DNA double-strand breaks that promotes the formation of RAD51 filaments. This interaction is triggered by DNA-damage and requires access to the DPP9 active-site. We present crystallographic structures documenting the N-terminal Met1-Pro2 of a BRCA21-40 peptide captured in the DPP9 active-site. Mechanistically, DPP9 targets BRCA2 for degradation by the N-degron pathway, and promotes RAD51 foci formation. Both processes are phenocopied by BRCA2 N-terminal truncation mutants, indicating that DPP9 regulates both stability and the cellular stoichiometric interactome of BRCA2. Consistently, DPP9-deprived cells are hypersensitive to DNA-damage. Together, we identify DPP9 as a regulator of BRCA2, providing a possible explanation for DPP9 involvement in cancer development. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.08.05.237172v1?rss=1 Authors: Paschini, M., Reyes, C. M., Gillespie, A. E., Lewis, K. A., Glustrom, L. W., Sharpee, T. O., Wuttke, D. S., Lundblad, V. Abstract: Telomeres present unique challenges for genomes with linear chromosomes, including the inability of the semi-conservative DNA replication machinery to fully duplicate the ends of linear molecules. This is solved in virtually all eukaryotes by the enzyme telomerase, through the addition of telomeric repeats onto chromosome ends. It is widely assumed that the primary site of action for telomerase is the single-stranded G-rich overhang at the ends of chromosomes, formed after DNA replication is complete. We show here that the preferred substrate for telomerase in wild type yeast is instead a collapsed fork generated during replication of duplex telomeric DNA. Furthermore, newly collapsed forks are extensively elongated by telomerase by as much as ~200 nucleotides in a single cell division, indicating that a major source of newly synthesized telomeric repeats in wild type cells occurs at collapsed forks. Fork collapse and the subsequent response by telomerase are coordinated by the dual activities of a telomere-dedicated RPA-like complex, which facilitates replication of duplex telomeric DNA and also recruits telomerase to the fork, thereby ensuring a high probability of re-elongation if DNA replication fails. We further show that the ability of telomerase to elongate newly collapsed forks is dependent on the Rad51 protein, indicating that telomerase activity in response to fork collapse proceeds through a regulatory pathway distinct from how telomerase engages fully replicated chromosome termini. We propose a new model in which spontaneous replication fork collapse and the subsequent response by telomerase is a major determinant of telomere length homeostasis. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.07.29.227678v1?rss=1 Authors: McKinzey, D. R., Gomanthinayagam, S., Griffin, W. C., Klinzing, K. N., Jeffries, E. P., Rajkovic, A., TRAKSELIS, M. Abstract: The MCM8/9 complex is implicated in aiding fork progression and facilitating homologous recombination (HR) in response to several DNA damage agents. MCM9 itself is an outlier within the MCM family containing a long C-terminal extension (CTE) comprising 42% of the total length, but with no known functional components and high predicted disorder. In this report, we identify and characterize two unique motifs within the primarily unstructured CTE that are required for localization of MCM8/9 to sites of mitomycin C (MMC) induced DNA damage. First, an unconventional bipartite-like nuclear localization (NLS) motif consisting of two positively charged amino acid stretches separated by a long intervening sequence is required for the nuclear import of both MCM8 and MCM9. Second, a variant of the BRC motif (BRCv), similar to that found in other HR helicases, is necessary for localization to sites of MMC damage. The MCM9-BRCv directly interacts with and recruits RAD51 downstream to MMC-induced damage to aid in DNA repair. Patient lymphocytes devoid of functional MCM9 and discrete MCM9 knockout cells have a significantly impaired ability to form RAD51 foci after MMC treatment. Therefore, the disordered CTE in MCM9 is functionally important in promoting MCM8/9 activity and in recruiting downstream interactors; thus, requiring full length MCM9 for proper DNA repair. Copy rights belong to original authors. Visit the link for more info

A roundtable discussion about prostate cancer featuring perspectives from Drs Neeraj Agarwal, Emmanuel S Antonarakis and A Oliver Sartor. Integrating PARP inhibitors into the treatment algorithm for patients with prostate cancer (PC) (00:00) Role of PARP enzymes in DNA damage repair and mechanism of action of PARP inhibitors (04:25) Genetic testing for patients with PC (06:31) Correlation between Gleason score and incidence of DNA repair gene mutations in patients with localized PC (11:24) Role of platinum-based chemotherapy for patients with PC and DNA repair gene mutations (15:34) Sequencing PARP inhibitors and chemotherapy for patients with PC and DNA repair gene mutations(19:40) Case (Dr Antonarakis): A man in his mid-40s with castration-resistant PC (CRPC), a BRCA2 mutation and lung metastases attains an excellent response to olaparib on a clinical trial (23:28) Tolerability and side effects of PARP inhibitors (27:59) Case (Dr Antonarakis): A man in his late 40s with metastatic hormone-sensitive PC and biallelic BRCA2 mutations receives PARP inhibitor monotherapy on a clinical trial (35:21) Perspective on the potential use of PARP inhibitor monotherapy in the first-line setting for patients with hormone-sensitive PC and DNA repair gene mutations (39:49) Tolerability of PARP-inhibitor monotherapy in the first-line setting (45:11) Case (Dr Sartor): A man in his late 60s with metastatic CRPC (mCRPC) and a germline BRCA2 mutation receives olaparib (49:08) Dosing and PARP trapping activity of PARP inhibitors (51:39) Case (Dr Agarwal): A man in his late 60s with progressive mCRPC is enrolled on the TALAPRO-2 trial of enzalutamide with or without talazoparib (55:07) Efficacy and tolerability of PARP inhibitors alone or in combination with endocrine therapy (59:32) Response to PARP inhibitors in patients with PALB2, FANCA and RAD51 mutations; clinical care of patients with PC during the COVID-19 pandemic (1:03:48) CME information and select publications

In patients with lung cancer whose tumors harbor activating mutations in the EGF receptor (EGFR), increased responses to platinum-based chemotherapies are seen compared with wild-type cancers. However, the mechanisms underlying this association have remained elusive. Here, we describe a cellular phenotype of crosslinker sensitivity in a subset of EGFR-mutant lung cancer cell lines that is reminiscent of the defects seen in cells impaired in the Fanconi anemia pathway, including a pronounced G2–M cell-cycle arrest and chromosomal radial formation. We identified a defect downstream of FANCD2 at the level of recruitment of FAN1 nuclease and DNA interstrand crosslink (ICL) unhooking. The effect of EGFR mutation was epistatic with FANCD2. Consistent with the known role of FANCD2 in promoting RAD51 foci formation and homologous recombination repair (HRR), EGFR-mutant cells also exhibited an impaired RAD51 foci response to ICLs, but not to DNA double-strand breaks. EGFR kinase inhibition affected RAD51 foci formation neither in EGFR-mutant nor wild-type cells. In contrast, EGFR depletion or overexpression of mutant EGFR in wild-type cells suppressed RAD51 foci, suggesting an EGFR kinase-independent regulation of DNA repair. Interestingly, EGFR-mutant cells treated with the PARP inhibitor olaparib also displayed decreased FAN1 foci induction, coupled with a putative block in a late HRR step. As a result, EGFR-mutant lung cancer cells exhibited olaparib sensitivity in vitro and in vivo. Our findings provide insight into the mechanisms of cisplatin and PARP inhibitor sensitivity of EGFR-mutant cells, yielding potential therapeutic opportunities for further treatment individualization in this genetically defined subset of lung cancer.

WORD of HOPE Ovarian Cancer Podcast. Episode 02, Topic 2 of 10.This is the second episode in the series of "10 Exciting Ovarian Research Topics from 2010". This episode focuses on PARP Inhibitors. Be sure to look for the remaining topics of this series in subsequent episodes of WORD of HOPE Ovarian Cancer Podcast.For show notes, links, info, and how to subscribe, visit: www.wordofhopepodcast.com

Background: Inactivation of the Fanconi anemia (FA) pathway through defects in one of 13 FA genes occurs at low frequency in various solid cancer entities among the general population. As FA pathway inactivation confers a distinct hypersensitivity towards DNA interstrand-crosslinking (ICL)-agents, FA defects represent rational targets for individualized therapeutic strategies. Except for pancreatic cancer, however, the prevalence of FA defects in gastrointestinal (GI) tumors has not yet been systematically explored. Results: A panel of GI cancer cell lines was screened for FA pathway inactivation applying FANCD2 monoubiquitination and FANCD2/RAD51 nuclear focus formation and a newly identified FA pathway-deficient cell line was functionally characterized. The hepatocellular carcinoma (HCC) line HuH-7 was defective in FANCD2 monoubiquitination and FANCD2 nuclear focus formation but proficient in RAD51 focus formation. Gene complementation studies revealed that this proximal FA pathway inactivation was attributable to defective FANCC function in HuH-7 cells. Accordingly, a homozygous inactivating FANCC nonsense mutation (c.553C > T, p.R185X) was identified in HuH-7, resulting in partial transcriptional skipping of exon 6 and leading to the classic cellular FA hypersensitivity phenotype; HuH-7 cells exhibited a strongly reduced proliferation rate and a pronounced G2 cell cycle arrest at distinctly lower concentrations of ICL-agents than a panel of non-isogenic, FA pathway-proficient HCC cell lines. Upon retroviral transduction of HuH-7 cells with FANCC cDNA, FA pathway functions were restored and ICL-hypersensitivity abrogated. Analyses of 18 surgical HCC specimens yielded no further examples for genetic or epigenetic inactivation of FANCC, FANCF, or FANCG in HCC, suggesting a low prevalence of proximal FA pathway inactivation in this tumor type. Conclusions: As the majority of HCC are chemoresistant, assessment of FA pathway function in HCC could identify small subpopulations of patients expected to predictably benefit from individualized treatment protocols using ICL-agents.

FLT3 (fms-like tyrosine kinase 3) ist bei ca. 30% aller Patienten mit akuter myeloischer Leukämie konstitutiv aktiviert und repräsentiert einen krankheitsspezifischen molekularen Marker. Zwei verschiedene Arten von Mutationen sind hierbei vorherrschend: ITD (internal tandem duplications) im Bereich der juxtamembranösen Region sind bei ungefähr 20-25% der Patienten nachweisbar sowie Mutationen im Bereich der Tyrosinkinasedomäne (TKD), die bei bis zu 8% der AML-Fälle auftreten. Es wurde bisher davon ausgegangen, dass die beiden Mutationstypen nicht im gleichen Patienten auftreten, da sie eine funktionelle Redundanz aufweisen. Neuere Daten zeigen jedoch, dass 1-2% aller AML-Patienten beide Mutationen (FLT3-ITD-TKD, duale Mutanten) tragen. Die Signifikanz dieser Beobachtung ist noch unklar, erste Studien haben jedoch gezeigt, dass diese dualen Mutationen mit einer besonders schlechten Prognose assoziiert sind. In dieser Arbeit konnte gezeigt werden, dass FLT3 duale Mutanten in in vitro Modellsystemen nicht nur Resistenzen gegenüber PTK Inhibitoren, sondern auch gegenüber dem Zytostatikum Daunorubicin induzieren können. Als molekularer Mechanismus hierfür konnte eine verstärkte Aktivierung von STAT5 (signal transducer and activator of transcription 5) sowie eine erhöhte Expression der STAT5-Zielproteine Bcl-x(L) und RAD51 identifiziert werden. Darüber hinaus konnte ein Arrest in der G2/M Phase des Zellzyklus beobachtet werden. Dieser Zellzyklusarrest erlaubt, DNA-Schäden zu reparieren und eine Apoptose zu vermeiden. Dass die Überexpression von Bcl-x(L) den wohl kritischen Punkt bei der Resistenzentstehung von FLT3-ITD-TKD Mutationen ausmacht, konnte dadurch bewiesen werden, dass Bcl-x(L)- und ITD- koexprimierende Zellen das Resistenzbild der dualen Mutanten imitieren können. Interessanterweise konnte der selektive mTOR-Inhibitor Rapamycin in Kombination mit PTK Inhibitoren die Sensitivität der dualen Mutanten wiederherstellen. Diese Arbeit beschreibt die molekularen Grundlagen von Resistenzbildungen gegenüber FLT3 PTK Inhibitoren und zeigt gleichzeitig therapeutische Ansätze auf, um Resistenzen zu verhindern oder zu überkommen.

Während der T-Zell-abhängigen Immunantwort bilden die aktivierten B-Lymphozyten das Keimzentrum, in dem Klassenwechselrekombination und somatische Hypermutation stattfinden. Der Mechanismus dieser Prozesse ist besonders im Falle der somatischen Hypermutation noch weitgehend unbekannt. Als essentieller Faktor konnte bisher die Aktivierungsinduzierte Cytidindeaminase AID identifiziert werden, die über Läsionen Mutationen und eventuell auch DNA-Doppelstrangbrüche in die DNA einführen kann. Da DNA-Brüche für die Zelle potentiell gefährlich sind, ist eine sehr stringente Regulation der DNA-Reparatur besonders in B-Zellen notwendig, denn eine fehlgeleitete Reparatur in B-Zellen kann zur Tumorentstehung beitragen. Prinzipiell haben die Zellen zwei Möglichkeiten DNA-Doppelstrangbrüche zu reparieren: die nicht homologen Endverknüpfung und die homologe Rekombination. Letztere unterteilt sich in die fehlerfreie konservative und die potentiell aberrante nicht konservative Rekombination. In dieser Arbeit wurde die Regulation und Aktivität der Doppelstrangbruchreparatur, im Besonderen der homologen Rekombination, in B-Zellen untersucht. Diese Analysen sollten Aufschluss über eine mögliche Beteiligung dieses Prozesses an der somatischen Hypermutation geben und ergründen, wie B-Zellen auf die AID-abhängigen DNA-Läsionen reagieren. Eine Analyse der Expression von Doppelstrangbruchreparaturfaktoren in primären B-Zellen und B-Lymphomzelllinien ergab eine erhöhte Expression dieser Faktoren in Keimzentrums-B-Zellen, die unter Anderem auf eine proliferationsgekoppelten Regulation der Reparaturgene zurückzuführen ist. Auffällig war besonders die differentielle Expression des Rekombinationsfaktors Rad51 in den Zelllinien. Die Bestimmung der Aktivität der homologen Rekombination in diesen Zellen mit Hilfe eines extrachromosomalen Reporters, der auf zwei hintereinander liegenden nicht funktionellen GFP-Genen basiert, zeigte eine Hyperrekombinationsaktivität für einige Zelllinien. In einem Tetracyclin regulierbaren System konnte transkriptionsgekoppelte Hyperrekombination gezeigt werden, die mit AID-Expressionsmengen korreliert. Es handelt sich dabei um fast ausschließlich nicht konservative Rekombination. Eine Nutzung der konservativen Rekombination konnte mit Rad51-Expressionsmengen korreliert werden. Eine Überexpression von AID bewirkte eine Erhöhung der Rekombinations- und der Hypermutationsaktivität. Folglich führt AID wahrscheinlich eine erhöhte Anzahl von Brüchen in die DNA ein, während Rad51 Einfluss auf die Wahl des Rekombinationsweges nehmen kann. In den reparierten GFP-Genen konnten keine Mutationen gefunden werden. Somit ist die Rekombinationsaktivität wohl eher eine Folge der AID-induzierten Brüche, als ein an der somatischen Hypermutation direkt beteiligter Weg. Die präferentielle Nutzung nicht konservativer Rekombination in den B-Lymphomzelllinien weist auf mögliche Folgen oder auch Ursachen der Lymphomentstehung hin oder ist eine von den Keimzentrumsprozessen geforderte physiologische Bedingung in Keimzentrums-B-Zellen, um einen Beitrag zur Antikörper-Diversifizieung zu leisten. Diese Fragen könnten durch weitere Untersuchungen geklärt werden, die die Rolle von AID in der Induktion der Rekombination betreffen und den Einfluss von Rad51 auf die Rekombinationswege.

Modern light microscopical techniques were employed to follow dynamical nuclear processes during the cell cycle and during DNA-repair. Laser-UVA-microirradiation The protein Rad51 is essential for the repair of double-strand breaks (DSBs) via the conservative homologous recombination repair pathway. To test the hypothesis that Rad51 localizes to damaged sites during DSB repair, a laser-UVA-microirradiation system was established. With this system spots with sizes around 1 µm in nuclei of living cells can be irradiated with UVA-light. After sensitization of cells by incorporation of BrdU into nuclear DNA and staining with the live cell dye Hoechst 33258, the system can be used to introduce double-strand breaks and single-strand breaks in the irradiated spots. The response of Rad51 to microirradiation By use of laser-UVA microirradiation the localization of Rad51 at damaged sites containing DNA double-strand breaks could be demonstrated. The accumulation of Rad51 at microirradiated sites was followed in cells fixed at increasing times after microirradiation. First Rad51 accumulations were visible 5 - 10 minutes after irradiation, and the number of cells with Rad51 accumulations increased until a plateau was reached 20 - 30 minutes after irradiation. In contrast, the majority of irradiated cells had accumulations of Mre11 protein already 5 - 10 minutes after irradiation. This is consistent with reports that nuclear Mre11 foci appeared early in the response to ionizing radiation, but absolute response times were faster after microirradiation than after ionizing radiation. Large-scale nuclear patterns were microirradiated, and Rad51 accumulations that reflected the shape of the irradiated patterns were found up to eight hours after irradiation. This conservation of the pattern of Rad51 accumulations, which reflect sites containing the damaged DNA, indicated that the chromatin in the irradiated cells performs no large scale reordering in response to DNA damage. The dynamics of chromosomes and chromosome territories In 1909 Theodor Boveri forwarded the hypothesis that arrangements of chromosome territories (CTs) are stably maintained during interphase, but subject to changes during mitosis. In the last decade several groups reported evidence for the stability of CT arrangements, but considerable movements of chromosomal subregions were also observed. The data concerning the maintenance or reordering of CTs during mitosis have been contradictory. Live cell imaging To follow the movements of chromosomes and CTs, a novel experimental approach was taken. Cells expressing a fusion protein of the core histone H2B with GFP (H2B-GFP) stably incorporate H2B-GFP into nucleosomes. In these cells chromatin regions were selectively marked by laser-photobleaching and followed by live cell microscopy. To this end, a live cell imaging system was established at a confocal laser-scanning microscope, which allows the observation of living cells for several days. Chromatin movements visualized by photobleached H2B-GFP To track possible movements in interphase cell nuclei, stripe patterns were bleached into nuclei at several stages of interphase. These patterns were retained for up to two hours, until they became invisible due to the replacement of bleached H2B-GFP by unbleached H2B-GFP, supporting the hypothesis that CT order is stably maintained during interphase. Nuclei, in which all chromatin except for a contiguous zone at one nuclear pole was bleached, were followed through mitosis. At prophase a number of unbleached chromosomal segments became visible. The segments showed a variable degree of clustering in metaphase. When daughter nuclei were formed, the segments locally decondensed into patches of unbleached chromatin. In all daughter cells the patches were separated by bleached chromatin, and clustered to a variable extent. These observations support the hypothesis that changes of chromosome neighborhoods occur during mitosis and that CT neighborhoods can profoundly vary from one cell cycle to the next.