Liquid biopsy: A new diagnostic concept in oncology

Video Transcript

• 0:00 - 00:09: Hello, and thank you for watching this webinar.
• 00:09 - 00:14: My name is Subham Bashu, and I’m the Director of Strategy for Immuno-Oncology at Abcam.
• 00:14 - 00:18: I’m hosting this webinar, and I’m delighted to welcome Professor Klaus Pantel, who will
• 00:18 - 00:23: present a talk entitled Liquid Biopsy, a New Diagnostic Concept in Oncology.
• 00:23 - 00:27: Klaus is the Chairman of the Institute of Tumor Biology at the University Medical Center
• 00:27 - 00:32: at Hamburg, and he graduated from Cologne University in 1986 and completed his thesis
• 00:32 - 00:37: on mathematical modeling of hemopoietidosis in 1987.
• 00:37 - 00:42: After receiving his postdoctoral period in the U.S.A. on hemopoietic stem cell regulation
• 00:42 - 00:46: at Wayne State University in Detroit, he performed research at the Institute of Immunology at
• 00:46 - 00:49: the University of Munich for 10 years.
• 00:49 - 00:54: Klaus’ pioneering work in the field of cancer micrometastasis, circulating tumor cells,
• 00:54 - 00:59: and circulating nucleic acids is reflected by over 400 publications, and he’s been awarded
• 00:59 - 01:05: the AACR Outstanding Investigator Award in 2010, the German Cancer Award in 2010, and
• 01:05 - 01:10: two ERC Advanced Investigator Grants in 2011 and 2019.
• 01:10 - 01:16: Moreover, Klaus has coordinated the European IMI Consortium Cancer ID on blood-based liquid
• 01:16 - 01:22: biopsies in lung and breast cancer, which comprises over 40 partner institutions
• 01:22 - 01:26: from academia, non-profit organizations, and industry.
• 01:26 - 01:31: And on top of that, he’s established the European Liquid Biopsy Society, the ELBS, as a successor
• 01:31 - 01:32: consortium.
• 01:32 - 01:34: I would like to now hand over to Klaus.
• 01:34 - 01:38: Well, thank you very much for this kind introduction.
• 01:38 - 01:44: It’s actually my pleasure today to give my presentation on liquid biopsy, a new diagnostic
• 01:44 - 01:46: concept in oncology.
• 01:46 - 01:49: What is the definition of liquid biopsy?
• 01:49 - 01:55: It has been extended to the analysis of tumor cells, but also immune cells or endothelial
• 01:55 - 02:02: cells or their products, and particularly DNA, microRNA, extracellular vesicles, and
• 02:02 - 02:07: proteins, of course, in the blood, but also in other body fluids.
• 02:07 - 02:13: And the vision here is to get more comprehensive and real-time information by the analysis
• 02:13 - 02:18: of these body fluids, in addition to the information that we get from the usual tissue
• 02:18 - 02:21: biopsies.
• 02:21 - 02:28: As you can see on that scheme, there are many different liquid biopsy analytes in the blood,
• 02:28 - 02:33: and they can be released either from the primary tumor, from the lymph nodes where the tumor
• 02:33 - 02:41: cells can actually travel to, or from distant metastatic sites like metastasis in the brain,
• 02:41 - 02:43: in the liver, or in the bones.
• 02:43 - 02:50: And all these different tumor sites, depending on the tumor stage, can release tumor cells
• 02:50 - 02:56: or non-tumor cells and tumor cell products into the bloodstream, and by taking a blood
• 02:56 - 03:03: sample, we can get this comprehensive picture.
• 03:03 - 03:09: How can we actually analyze and capture these liquid biopsy markers?
• 03:09 - 03:15: There are two big classes of liquid biopsy markers, the tumor DNA, which is really part
• 03:15 - 03:22: of the plasma, where we also find the circulating proteins, the extracellular vesicles, or the
• 03:22 - 03:29: microRNAs, and I won’t go into details about the circulating DNA today, but we also can
• 03:29 - 03:35: assess the circulating tumor cells, which are part of the cellular components of the
• 03:35 - 03:41: blood, and I would like to go into more detail how we do that, actually.
• 03:41 - 03:45: First of all, we need to enrich the tumor cells, and for that, we use, actually, protein
• 03:45 - 03:50: markers expressed on the cell surface of the circulating tumor cells.
• 03:50 - 03:56: These are antibodies to epithelial markers, such as EPCAM, or also antibodies to mesenchymal
• 03:56 - 04:00: markers when the tumor cells underwent an EMT.
• 04:00 - 04:06: We can also negatively select the tumor cells by pulling out the leukocytes with anti-CD45
• 04:06 - 04:13: antibodies, or we can use physical differences between tumor cells and non-tumor cells to
• 04:13 - 04:16: enrich for the CTCs.
• 04:16 - 04:21: After the enrichment, we do a detection at the single-cell level to say which of the
• 04:21 - 04:26: enriched cells are really tumor cells, and here we again use protein markers, and particularly
• 04:26 - 04:34: markers to epithelial proteins, such as the keratins, which are not expressed on the leukocytes,
• 04:34 - 04:40: but in the newer studies, we also have extended that concept with mesenchymal markers to catch
• 04:40 - 04:45: tumor cells that underwent an epithelial-mesenchymal transition.
• 04:45 - 04:50: In addition to that, we can also look at viable tumor cells, and that is an assay, the EpiSpot
• 04:50 - 04:55: Assay and EpiDrop Assay, developed by Catherine Alix-Panabières from Montpellier, and here the
• 04:55 - 05:03: tumor cells release tumor-specific proteins that can then be detected with antibodies.
• 05:03 - 05:07: And last but not least, of course, we can also look at tumor-specific transcripts using
• 05:07 - 05:10: PCR technology.
• 05:10 - 05:16: The newest development is really that besides enumeration of the tumor cells, we can also
• 05:16 - 05:21: characterize the tumor cells by picking the single cells that have been identified with
• 05:21 - 05:28: proteins, and then we can perform a genome analysis, including FISH and MGS.
• 05:28 - 05:34: We can look at the expression profiling, for example, by RNA sequencing.
• 05:34 - 05:39: We can look at the protein analysis, and we do that a lot using immunostaining and proteomics
• 05:39 - 05:44: methods, or we can do functional assays.
• 05:44 - 05:50: Why do we need actually different types of proteins to identify our tumor cells?
• 05:50 - 05:55: Because the tumor cells can have a high plasticity if they are released from an epithelial tumor,
• 05:55 - 06:02: for example, from breast cancer, tumor cells can undergo an epithelial-mesenchymal transition,
• 06:02 - 06:05: and that may help them to disseminate through the body.
• 06:05 - 06:11: But when they want to form a metastasis in a distant organ, they may have to go the revised
• 06:11 - 06:19: pathway, a mesenchymal-epithelial transition, and then their re-expressed epithelial markers.
• 06:19 - 06:24: That’s why epithelial protein markers are still very important in the CTC field, but
• 06:24 - 06:31: we also have to consider that some of the cells have changed their phenotype.
• 06:31 - 06:36: What are the clinical applications that we can use this analysis for?
• 06:36 - 06:42: Well, I think overall, we can monitor tumor burden in cancer patients from the beginning
• 06:42 - 06:49: of tumor development to the development of overt metastatic disease.
• 06:49 - 06:55: In the beginning, I think the first application is, can we use blood analysis and tissue-body-fluid
• 06:55 - 07:00: analysis to actually early detect tumors?
• 07:00 - 07:06: The second question is, can we identify those tumors after initial surgery and adjuvant
• 07:06 - 07:14: therapy where there is minimal residual disease left, and this disease may actually cause relapse?
• 07:14 - 07:20: And the third application, when the patient has metastatic full-blown relapse, we can
• 07:20 - 07:29: ask the question, what characteristics of the CTCs and other liquid biomarkers can give
• 07:29 - 07:36: us an idea of what kind of therapy is most effective?
• 07:36 - 07:40: It should also be mentioned just briefly that besides blood, there are also other body fluids
• 07:40 - 07:46: which are very interesting, and we have done quite a bit of work on the bone marrow, which
• 07:46 - 07:51: is a common reservoir for disseminating dormant tumor cells.
• 07:51 - 07:54: Let me go through the different clinical applications now.
• 07:54 - 07:59: The first one is early detection of cancer, and early detection means that the aim is
• 07:59 - 08:07: really to identify small tumors, T1 tumors, or early-stage, stage 1 disease, because we
• 08:07 - 08:12: want to detect tumors early in order to cure the patients.
• 08:12 - 08:16: Here, there are several things that have to be taken into account.
• 08:16 - 08:21: First of all, the sensitivity, we want to avoid false negative findings.
• 08:21 - 08:27: Secondly, is the specificity, we want to avoid false positive findings, and this is just
• 08:27 - 08:29: a theoretical calculation.
• 08:29 - 08:37: If we have a 99% specificity and 1% false positive findings, it sounds not much, but
• 08:37 - 08:43: it means 1 out of 100, and if we screen 10 million, there might be 100,000 false positives,
• 08:43 - 08:45: which is quite a lot.
• 08:45 - 08:50: And also, I think in most of the studies, the cancer cases are compared to a control
• 08:50 - 08:55: group, but the control group needs to be age-matched to the cancer patients, and there also need
• 08:55 - 09:02: to be a control group with confounding non-cancer diseases, because many patients that are at
• 09:02 - 09:09: the age 50 or older have not only the cancer, but maybe also other diseases that may interfere
• 09:09 - 09:13: with the assay specificity.
• 09:13 - 09:19: A good example was really published a couple of years ago by the Hopkins group, Nick Papadopoulos
• 09:19 - 09:26: and his colleagues, and here they actually paved the avenue for combining markers in
• 09:26 - 09:28: the blood.
• 09:28 - 09:34: What they actually did here, they tested on the circulating tumor DNA, 16 genes, and they
• 09:34 - 09:40: looked at hundreds of mutations on these genes, and then they combined it with eight protein
• 09:40 - 09:47: biomarkers, and these were circulating proteins in the blood, and then they used a machine-based
• 09:47 - 09:53: learning approach to analyze the data and came up with highly specific tests that worked
• 09:53 - 09:55: in several tumor entities.
• 09:56 - 10:01: Also, some of the entities did not show such a high sensitivity, such as breast cancer.
• 10:01 - 10:09: I still feel that this is really the way to go to combine and include protein markers.
• 10:09 - 10:10: Why is that the case?
• 10:10 - 10:17: Well, if we only focus on genomic markers, there is also a challenge, because if people
• 10:17 - 10:23: get older, there is a background of genomic aberrations, which may not necessarily lead
• 10:23 - 10:26: to cancer in the lifetime of the patients.
• 10:26 - 10:31: This has been shown in this publication a few years ago that actually could identify
• 10:31 - 10:37: leukemia-associated mutations in non-cancer-controlled patients with increasing age, so-called CHIP
• 10:37 - 10:43: mutations, and also these patients have a slight increase to develop leukemia.
• 10:43 - 10:49: Most of these patients are not developing any cancer, so these mutations have to be
• 10:49 - 10:52: subtracted.
• 10:52 - 10:59: So our vision is really that we should develop a composite liquid biopsy marker panel for
• 10:59 - 11:04: early cancer detection, but also for other applications, and this panel can include the
• 11:04 - 11:10: different markers that are present in the circulation that I mentioned before, and definitely
• 11:10 - 11:20: also include proteins that can be found in the plasma or serum of cancer patients.
• 11:20 - 11:27: In Europe, we have actually two networks that are funded by the European Union.
• 11:27 - 11:33: One is the ELBA, the European Liquid Biopsy Academy Network, coordinated by Tom Wurdinger,
• 11:33 - 11:39: with a focus on detection of lung cancer, and the second consortium is the Prolipsy Consortium
• 11:39 - 11:46: and ARINET Transcan Project, coordinated by myself, and here the focus is on early detection
• 11:46 - 11:49: of prostate cancer.
• 11:49 - 11:55: In Hamburg, we also have started the Hamburg City Health Study in 2015, which is a large
• 11:55 - 12:04: cohort study where we get biomaterial from 45,000 individuals between age 45 and 74,
• 12:04 - 12:12: which is a risk for many diseases, including cancer, and we collect 120 ml of blood plasma
• 12:12 - 12:21: together with other tissues and, of course, a large network of data.
• 12:21 - 12:26: The second application is, can we improve with liquid biopsy cancer staging?
• 12:26 - 12:33: And here, CTCs detected by protein markers have shown remarkable prognostic relevance.
• 12:33 - 12:40: This is shown on this slide, which is a summary of the studies that we performed, and it shows
• 12:40 - 12:47: that the CTC counts at initial cancer diagnosis are associated with unfavorable prognosis,
• 12:47 - 12:51: and these are CTCs identified by protein markers.
• 12:51 - 12:57: And as you can see here, particularly in breast cancer, there is quite some evidence that
• 12:57 - 13:05: the CTCs actually are inherited with an increased risk of developing metastatic relapse, and
• 13:05 - 13:12: that is shown really on the next slide, but this is just to summarize that the CTCs can
• 13:12 - 13:18: be used as an enrichment tool to study high-risk populations, and there has been even a certain
• 13:18 - 13:22: classification in the AJCC Cancer Staging Manual.
• 13:22 - 13:30: It’s called CM0, which means no signs of overt metastasis, but Iplus identifies the presence
• 13:30 - 13:35: of individual tumor cells in the blood or other tumor tissues.
• 13:35 - 13:40: This is just one piece of data from this list of clinical studies.
• 13:40 - 13:46: It’s a large-scale study on thousands of breast cancer patients together with our European
• 13:46 - 13:52: and American colleagues, and here we actually looked at CTCs before neoadjuvant cancer therapy
• 13:52 - 13:58: in breast cancer and showed the connection to overall survival.
• 13:58 - 14:03: In the Kaplan-Meier curves and the graph as you see here, you can see that the number
• 14:03 - 14:11: of CTCs are a risk factor for survival, and in the table below that Kaplan-Meier curves,
• 14:11 - 14:18: you can see actually the results of the multivariate analysis, and if we found five or more CTCs
• 14:18 - 14:26: in 7.5 mL blood, the risk to develop metastasis was more than six times increased, so quite
• 14:26 - 14:30: a remarkable effect.
• 14:30 - 14:35: The next application is, of course, after surgery and after adjuvant therapy, can we
• 14:35 - 14:42: follow up the blood samples and actually analyze and detect relapse much earlier than with
• 14:42 - 14:45: imaging procedures?
• 14:45 - 14:51: And this is one data set, again, in breast cancer from our large clinical trials, a success
• 14:51 - 14:57: study group in Germany led by Wolfgang Janni from Ulm, and here we were evaluating the
• 14:57 - 15:03: blood two years after adjuvant chemotherapy, and we could detect circulating tumor cells
• 15:03 - 15:08: two years after adjuvant chemotherapy, and these cells are not derived from the primary
• 15:08 - 15:13: tumor that was taken out by surgery, but they are actually derived from occult metastatic
• 15:13 - 15:15: disease.
• 15:15 - 15:22: And if we detect these tumor cells, we can see that it was connected to decreased overall
• 15:22 - 15:27: survival and decreased disease-free survival.
• 15:27 - 15:34: So if we have now the chance to detect minimal disease years after initial treatment, there
• 15:34 - 15:39: is, of course, now the next step required, which is the design of so-called post-adjuvant
• 15:39 - 15:45: clinical trials that find out whether an early treatment of these metastatic
• 15:45 - 15:52: patients will actually benefit the patients in terms of disease-free or overall survival.
• 15:52 - 16:00: The next application is that we can use liquid biopsy markers to detect therapeutic targets
• 16:00 - 16:06: and resistant mechanisms, and there are two types of analysis that can be done.
• 16:06 - 16:12: There are analysis of genomic aberrations, which looks for druggable mutations and resistant
• 16:12 - 16:18: mechanisms engraved in the DNA, and this can be done on the circulating tumor cells or
• 16:18 - 16:24: on the circulating DNA, and it’s one of the key applications of circulating DNA, but there’s
• 16:24 - 16:31: also another world where the DNA is not giving us any information so far, which is the transcriptional
• 16:31 - 16:37: plasticity, which is very important for resistant therapy, and here we have done a lot of work
• 16:37 - 16:44: looking at proteins expressed on circulating tumor cells that are involved as therapeutic
• 16:44 - 16:45: targets.
• 16:45 - 16:50: For example, the androgen receptor, the estrogen receptor in breast cancer, the HER2 in breast
• 16:50 - 16:58: cancer, the PD-L1 as a target for immunotherapy, or the PSMA antigen in prostate cancer.
• 16:58 - 17:03: Just to show you one example, which I think is probably in the focus of many clinical
• 17:03 - 17:09: oncologists today, it’s immunotherapy, immune checkpoint inhibition therapy, that is based
• 17:09 - 17:15: on the Nobel Prize-winning knowledge that there is an interaction between the tumor
• 17:15 - 17:18: cells and the lymphocytes.
• 17:18 - 17:25: The tumor cells are recognized by the lymphocytes through the MHC system, but this is not enough.
• 17:25 - 17:30: They need to be also activated, and that actual activation can be blocked by the
• 17:30 - 17:33: interaction between PD-L1 and PD-1.
• 17:33 - 17:40: So these two molecules have become targets of antibody-based therapy in many types of
• 17:40 - 17:46: solid tumors, and of course, there is a big need now to find out which patients are the
• 17:46 - 17:52: ones that respond best to these therapies, because these therapies also have side effects,
• 17:52 - 17:59: and can we also monitor the therapy effect by continuous blood analysis.
• 17:59 - 18:06: This is just out of a paper published together with Catherine Aix-Panabières a few years ago,
• 18:06 - 18:12: and we could actually show that we are able, with antibodies to PD-L1, to characterize the
• 18:12 - 18:18: circulating tumor cells and the leukocytes, and it shows that this type of analysis gives
• 18:18 - 18:24: you the information whether the CTCs are PD-L1 positive in the upper panel or negative in
• 18:24 - 18:30: the lower panel, but also whether the surrounding leukocytes express these markers.
• 18:30 - 18:36: Well, we can also go one step further beside descriptive analysis.
• 18:36 - 18:42: We can also have entered now the age of functional analysis that can be done, and for this functional
• 18:42 - 18:47: analysis, we can either use xenografts, and we have published a paper together with a
• 18:47 - 18:53: group in Heidelberg in 2013 that xenografts can be established from circulating tumor
• 18:53 - 18:59: cells in breast cancer, and together with Catherine Alix-Panabières in Montpellier, we have
• 18:59 - 19:05: also established the first cell lines from circulating tumor cells for functional analysis,
• 19:05 - 19:10: and the last publication just came out in EMBO Molecular Medicine on a new cell line
• 19:10 - 19:17: established from an ER-positive breast cancer patient, and this cell line also mimics what
• 19:17 - 19:19: happens in the cancer patients.
• 19:19 - 19:26: It shows growth in immunodeficient mice, but it also metastasizes to organs that are relevant
• 19:26 - 19:31: in breast cancer, and it’s one of the few estrogen receptor-positive cell lines, and
• 19:31 - 19:37: ER-positive tumors are the main type of breast cancers.
• 19:37 - 19:43: Of course, we can also learn something about the biology of tumors in cancer patients,
• 19:44 - 19:49: and one example we are very much interested in is the biology of brain metastasis and
• 19:49 - 19:51: how that develops.
• 19:51 - 19:58: So here we performed actually single-cell exome sequencing of CTCs that were isolated
• 19:58 - 20:04: from the blood of breast cancer patients with brain metastasis, and we could actually perform
• 20:04 - 20:11: a whole genome analysis and look for gains here in green or losses of genomic material
• 20:11 - 20:19: in red, and you can see that quite a few of those aberrations were seen in 80-100%
• 20:19 - 20:25: of the CTCs isolated, so there was quite a high ploidy, and maybe these regions can
• 20:25 - 20:31: be used to determine whether the circulating cells in the blood are derived from brain
• 20:31 - 20:33: metastasis.
• 20:33 - 20:40: Another example is we looked at the expression of proteins on the tumor cells, and this adhesion
• 20:40 - 20:47: protein L1CAM was found in brain metastasis from non-small cell lung cancer patients.
• 20:47 - 20:52: We also could show that it had a prognostic relevance, and we could also show a functional
• 20:52 - 20:59: relevance in this publication that just came out in Neuro-Oncology this year, and interestingly,
• 20:59 - 21:04: in a very small subset of patients, we looked whether the expression of this protein on
• 21:04 - 21:10: the CTCs resembled the expression on the metastasis, and that was actually the case,
• 21:10 - 21:20: so maybe the CTCs can be also used to get information on the biology of brain metastasis.
• 21:20 - 21:27: So in conclusion, we believe that the CTCs, the DNA, but also definitely the proteins
• 21:27 - 21:31: provide complementary information for liquid biopsies.
• 21:31 - 21:36: The assays need to be validated by independent expert groups, that is very important, and
• 21:36 - 21:41: also we need more interventional clinical studies, which are required to demonstrate
• 21:41 - 21:47: the clinical utility of liquid biopsy, which actually means, can the liquid biopsy test
• 21:47 - 21:53: help the patients to have a better life or to live longer?
• 21:53 - 22:00: Well, one of the problems is that we have many great publications on liquid biopsy markers,
• 22:00 - 22:06: but there is very little in the clinic, and there is this translation from publication
• 22:06 - 22:10: to clinical routine, which is usually a bottleneck.
• 22:10 - 22:16: To overcome this bottleneck, we have founded a few years ago a European consortium, which
• 22:16 - 22:21: was actually supported by the European Union, and I had the pleasure to do the scientific
• 22:21 - 22:29: management together with the coordinator from the FPR side, Thomas Schlange, and Leon Terstappen
• 22:29 - 22:35: from the academic side, and we were all together at the end, 40 partners from the European
• 22:35 - 22:41: Union, academic institutions, non-profit organizations, and companies, and I think it’s very important
• 22:41 - 22:46: to bring both sides together and be focused on blood-based diagnostics in lung and breast
• 22:46 - 22:47: cancer.
• 22:47 - 22:56: This funding has been, it’s not extendable, so we actually did something interesting,
• 22:56 - 23:02: we founded our own network, which is called the European Liquid Biopsy Society in 2019
• 23:02 - 23:08: to sustain our activities and go on towards bringing these markers into clinical practice.
• 23:08 - 23:15: And we had our kickoff meeting last year already with 40 institutions, 26 from academia, 14
• 23:15 - 23:22: from companies, and the main goal here is to support translational liquid biopsy research.
• 23:22 - 23:27: We also have reached out to other areas of the world, particularly to the USA, but also
• 23:27 - 23:33: to Asia, and I think this is very important that we actually have a worldwide network
• 23:33 - 23:34: and teamwork.
• 23:34 - 23:39: So what are the goals of the Liquid Biopsy Society?
• 23:39 - 23:44: We want to foster the introduction of liquid biopsy into clinical practice, that is really
• 23:44 - 23:50: our key goal, encourage interaction between academia and industry, provide partners, provide
• 23:50 - 23:56: a partner for regulatory agencies, healthcare providers and patient advocacy groups, implement
• 23:56 - 24:02: clinical trials that we actually use liquid biopsy tests into clinical trials, develop
• 24:02 - 24:09: guidelines and provide training, disseminate knowledge, increase the visibility of Europe
• 24:09 - 24:13: as hub for liquid biopsy research, and outreach, of course, to other areas.
• 24:13 - 24:19: And if you’re interested in this society to become a member, and here we only have institutional
• 24:19 - 24:24: memberships, not individual memberships, this will be fantastic, and just write me
• 24:24 - 24:32: a mail and let me know if you’re interested in our mission concept paper.
• 24:32 - 24:39: Well at the end I would like to thank my colleagues also at the University Medical Center in Hamburg.
• 24:39 - 24:45: We have established a liquid biopsy research network for the past 20 years, and this is
• 24:45 - 24:51: just to show you that this network includes institutes and clinical departments, and I
• 24:51 - 24:57: think this translational interaction between the people who develop assays and the people
• 24:57 - 25:02: who need these assays in clinical trials is incredibly important.
• 25:02 - 25:06: I would also like to thank my team at the Institute of Tumor Biology and the Center
• 25:06 - 25:11: of Experimental Medicine and at our Comprehensive Cancer Center.
• 25:11 - 25:20: These are the people that really do the work, and I’m very proud to be the leader of this group.
• 25:20 - 25:25: We have also received sustained funding from third parties, and particularly from the European
• 25:25 - 25:30: Union to ERC grants and advanced investigator grants.
• 25:30 - 25:35: We have also supported ERC grants on proof of concept grants to bring this technology
• 25:35 - 25:43: into clinical practice, but also from other EU consortium funding bodies.
• 25:43 - 25:48: We also have received sustained funding from German funding bodies, such as the Deutsche
• 25:48 - 25:53: Forschungsgemeinschaft and the Minister of Research and Science, and particularly also
• 25:53 - 26:00: a nice postgraduate center that’s funded by German Cancer and the Deutsche Krebsdiabetik.
• 26:00 - 26:03: Thank you very much for this funding.
• 26:03 - 26:08: At the end, I would like to point your attention to a meeting that will be held now online,
• 26:08 - 26:12: not in real, on the 24th and 25th of October.
• 26:12 - 26:19: It’s our 12th International Symposium on Liquid Biopsy for Cancer, which will cover all aspects of
• 26:19 - 26:24: liquid biopsy, and if you’re interested, please have a look at our website.
• 26:24 - 26:30: This will be actually co-hosted together with Dr. Lu from Shanghai, Dr. Catherine Alix-Panabières
• 26:30 - 26:35: from Montpellier, and Dr. Qiu Shi from Shanghai.
• 26:35 - 26:37: Thank you very much for your attention.
• 26:37 - 26:39: Thank you, Klaus, for a great talk.
• 26:39 - 26:43: That was really wonderful to hear all the different aspects of liquid biopsy, from ctDNA
• 26:43 - 26:48: to protein, and also to, most importantly, see some of the translational and clinical
• 26:48 - 26:49: work that will be done on that.
• 26:49 - 26:54: If you don’t mind, I would like to ask you some questions, which I know that people attending
• 26:54 - 26:58: this webinar would probably be quite interested in hearing your insights from.
• 26:58 - 26:59: Sure.
• 26:59 - 27:01: I’m happy to answer that.
• 27:01 - 27:02: Great.
• 27:02 - 27:06: You’ve answered this, but perhaps if you could just really delineate what you think that
• 27:06 - 27:13: liquid biopsy brings to both clinical and diagnostics that regular tissue analysis doesn’t.
• 27:13 - 27:19: Yeah, I think that regular tissue analysis is, of course, a cornerstone of our diagnostics
• 27:19 - 27:24: in oncology, but there are certainly some limitations.
• 27:24 - 27:28: For example, some locations are very difficult to biopsy.
• 27:28 - 27:35: For example, tumors in the lung or tumors in the brain, and in particular, metastases
• 27:35 - 27:40: are very difficult, actually, to biopsy and do not undergo many times biopsies.
• 27:40 - 27:46: For example, a metastasis in the bone is not easy to biopsy, or a metastasis in the lung,
• 27:46 - 27:48: or a metastasis in the brain.
• 27:48 - 27:56: In order to get information from these different sites and tumors, I think that liquid biopsy
• 27:56 - 28:05: in terms of a blood analysis will contribute to that information, because I don’t think
• 28:05 - 28:09: that we can really put a needle into all of these metastatic sites.
• 28:09 - 28:16: Also, if we want to follow up tumor evolution, we have to take an analysis maybe every two
• 28:16 - 28:23: months or every six weeks, and I think it’s quite difficult to put a needle into a tissue
• 28:23 - 28:27: sequentially many times in an individual patient.
• 28:27 - 28:33: So I’m very optimistic that the concept of liquid biopsy, by getting tumor information
• 28:33 - 28:40: relevant to patient treatment from a blood sample, will actually succeed.
• 28:40 - 28:41: Great.
• 28:41 - 28:42: Thank you.
• 28:42 - 28:47: You mentioned and you showed how many groups are using both sort of nucleic acid analysis
• 28:47 - 28:49: and protein analysis.
• 28:49 - 28:55: What do you think is the real role of looking at protein analysis in liquid biopsy, and
• 28:55 - 28:59: what do you think are the major methodologies being used now and soon to be used?
• 28:59 - 29:07: Yeah, I think the major role are really the targets that are expressed on the tumor cells,
• 29:07 - 29:09: and we have many examples.
• 29:09 - 29:15: If you look, for example, in breast cancer, the oldest therapeutic target is the estrogen
• 29:15 - 29:20: receptor, and I think it’s still interesting to know whether the estrogen receptor is still
• 29:20 - 29:26: expressed, because we know in a certain fraction of patients, the original status from the
• 29:26 - 29:30: primary tumor may change toward the development of metastasis.
• 29:30 - 29:36: So we need to know, for example, whether this protein is expressed on the tumor cells.
• 29:36 - 29:43: Another example, I think, of course, in prostate cancer, the androgen receptor or the variants
• 29:43 - 29:49: of the androgen receptor or the PSMA that is also used as therapeutic targets in clinical
• 29:49 - 29:55: trials, you need to know whether this protein is really there when we add antibodies targeting
• 29:55 - 29:56: these proteins.
• 29:56 - 30:05: So I think that there is still a clear role of protein analysis in this field.
• 30:05 - 30:10: We’re also asked, of course, about what methods, and I say methods, the future methodology
• 30:10 - 30:18: or the challenges now to develop highly sensitive methods for multiplex analysis of minimal
• 30:18 - 30:20: amounts of protein.
• 30:20 - 30:22: We want the best from all worlds.
• 30:22 - 30:30: We want high sensitivity, we want multiplex information, and we want the technology developer
• 30:30 - 30:36: to do that from very small amounts of protein so we can do single cell analysis or analysis
• 30:36 - 30:41: of extracellular vesicles, for example.
• 30:41 - 30:46: And how are you going about sort of either discovering or further validating, perhaps
• 30:46 - 30:52: first discovering, then validating targets for protein in liquid biopsy?
• 30:52 - 30:55: Yeah, we have actually a very interesting line.
• 30:55 - 31:02: We use our cell lines developed from circulating tumor cells and disseminated tumor cells and
• 31:02 - 31:09: also xenografts to identify and discover new targets.
• 31:09 - 31:16: We do that, of course, with protein screening, mass spec, and other methods, and this is
• 31:16 - 31:21: quite interesting, and then we can see which of those targets are not expressed on the
• 31:21 - 31:25: normal blood cells and not on other normal tissues.
• 31:25 - 31:33: And from that analysis, we came actually up to a series of new protein candidates that
• 31:33 - 31:36: can be then validated, of course, further.
• 31:36 - 31:43: So we actually feel that these models that we have developed from like cell lines and
• 31:43 - 31:50: xenografts from real CTCs from real cancer patients can have some very interesting information
• 31:50 - 31:54: on future therapeutic and diagnostic targets.
• 31:54 - 31:59: And are you able to share, if not, actual individual targets that you think are most
• 31:59 - 32:04: promising, even classes of targets, classes of proteins, that you think are most promising
• 32:04 - 32:08: and do those differ from cancer type to cancer type?
• 32:08 - 32:13: Yes, we have definitely targets that differ from cancer type to cancer type.
• 32:13 - 32:19: For example, the PSMA, you know, the prostate-specific membrane antigen, of course, is just
• 32:19 - 32:23: interesting, most interesting in prostate cancer.
• 32:23 - 32:30: I mean, definitely, but we also have targets, new targets, that show an overlapping expression
• 32:30 - 32:37: between different tumor entities, and, of course, these are also very, very interesting
• 32:37 - 32:39: to us, definitely.
• 32:39 - 32:46: And of course, we are very happy to collaborate in that field because we can only, of course,
• 32:46 - 32:51: do the validation up to a certain point in an academic setting.
• 32:51 - 32:58: And are you looking at these by ELISA, by sort of regular Western blot, by some sort
• 32:58 - 32:59: of flow cytometry?
• 32:59 - 33:03: What are some of the methodologies that you are using here?
• 33:03 - 33:09: Yeah, when we do our cell lines, of course, we do first the protein screening approaches
• 33:09 - 33:15: that are well-developed, and then we confirm the expression by Western blot analysis, but
• 33:16 - 33:22: then we go, if we go to the clinical samples, we want to develop, we have started to develop
• 33:22 - 33:27: specific ELISAs because they are much more sensitive, because in the clinical samples
• 33:27 - 33:33: we deal with very small amounts of proteins, so, and for the analysis of the circulating
• 33:33 - 33:38: single cells, we use immunohistochemical methods.
• 33:38 - 33:41: Okay, so IHC is also a part of it.
• 33:41 - 33:47: So, a wide variety depending on how many cells are available for analysis, right?
• 33:47 - 33:54: And what are some, you mentioned earlier multiplex analysis, what are some of the multiplex methodologies
• 33:54 - 33:55: you’re using?
• 33:55 - 34:05: Well, we actually use multiplex immunocytochemistry with up to eight, nine markers, you know.
• 34:05 - 34:11: We have also, of course, for multiplexing, if we have our cell lines, you know, we can
• 34:11 - 34:20: go really in mass spec and SILAC analysis, so we, but we have also started collaboration
• 34:20 - 34:28: with new methods where people use, for example, immuno-PCR technologies for multiplexing, so
• 34:28 - 34:32: we are really very much into that.
• 34:32 - 34:39: But I still think that there is a need to develop more of these methods, and we are
• 34:39 - 34:42: very happy for collaboration in that field.
• 34:42 - 34:47: Great, and finally, obviously a lot of the methodology that’s been used has been for
• 34:47 - 34:54: research or for use in, with patient samples in a clinical trial, retrospective, sometimes
• 34:54 - 35:00: a prospective analysis, but what do you think it would take for protein analysis of, by
• 35:00 - 35:06: whatever methodology, of clinical material to make it to the routine pathology?
• 35:06 - 35:12: Well, I think what we really need to do, and that was also the reason why we founded this
• 35:12 - 35:19: European Liquid Biopsy Society consortium, I think we need to do a thorough technical
• 35:19 - 35:21: and clinical validation of new technologies.
• 35:21 - 35:27: There are many assays that are published or even commercialized, but in our cancer ID
• 35:27 - 35:32: consortium, we could also see that doing ring experiments, doing quality proficiency
• 35:32 - 35:39: testing, you know, really helped also the companies to see how robust their assays are,
• 35:39 - 35:44: and this will also help, actually, to bring these tests into clinical trials and eventually
• 35:44 - 35:46: into clinical practice.
• 35:46 - 35:53: And I think what we really need, actually, to adopt, actually, are the barriers that
• 35:53 - 35:59: we have today is, we need more interventional clinical trials where we use these assays,
• 35:59 - 36:04: and depending on these assays, we make a therapeutic decision, and this can be only done in
• 36:04 - 36:08: controlled clinical trials, of course, but we need more of that.
• 36:08 - 36:13: We also need to disseminate the knowledge about liquid biopsy into the medical community.
• 36:13 - 36:19: I think we need to integrate, then, the tests into national or international guidelines,
• 36:19 - 36:20: that’s very important.
• 36:20 - 36:25: And, of course, at the end of all of this development, there’s always a key question,
• 36:25 - 36:29: will the test be reimbursed by the healthcare providers or not?
• 36:29 - 36:35: But I guess we need to do our homework before we can ask for the reimbursement, but also
• 36:35 - 36:42: the reimbursement is, of course, needs a good strategy and a clear goal.
• 36:42 - 36:43: Thank you.
• 36:43 - 36:47: Well, that was a fantastic talk and really great to get your insights afterwards.
• 36:47 - 36:52: It just leaves me to thank you again for participating in this webinar, and I really hope to hear
• 36:52 - 36:57: more from you and the group at ELBS and all your international collaborators in either
• 36:57 - 37:03: publications or future conferences, and I agree with you that I think liquid biopsy
• 37:03 - 37:09: is a very bright future in both research diagnostics and for routine pathology diagnostics.
• 37:09 - 37:10: Thank you.
• 37:10 - 37:15: Well, thank you, Olu, very much, and also for your interesting questions and giving
• 37:15 - 37:18: me the opportunity to present this work here.
• 37:18 - 37:19: Thank you.
• 37:19 - 37:20: Great.
• 37:20 - 37:21: Bye-bye.