WEBVTT 00:00:08.049 --> 00:00:09.592 Welcome to our video lecture 00:00:09.592 --> 00:00:13.513 entitled Classification of Leukemias and Lymphomas 00:00:13.805 --> 00:00:18.393 Increasing Role of Molecular Testing, presented by Doctor Rodney Miles. 00:00:19.019 --> 00:00:21.563 Doctor Rodney Miles is the medical director 00:00:21.563 --> 00:00:26.776 of the Hematologic Flow Cytometry and Immunohistochemistry Laboratories. 00:00:27.360 --> 00:00:31.322 Additionally, he is the section chief of the Genetic Pathology 00:00:31.573 --> 00:00:35.160 at r U.P laboratories along with being an associate 00:00:35.160 --> 00:00:39.164 professor of pathology at the University of Utah School of Medicine. 00:00:39.831 --> 00:00:43.543 He graduated from the University of Nebraska medical center 00:00:43.835 --> 00:00:47.547 with a master's degree and a doctorate degree in cell biology. 00:00:47.922 --> 00:00:52.844 He completed a pathology residency at the University of Utah School of Medicine, 00:00:53.136 --> 00:00:57.432 and a fellowship in Connecticut Biology at the University of Michigan. 00:00:58.141 --> 00:01:01.770 He is the recipient of the 2015 Warren G. 00:01:01.770 --> 00:01:05.523 Sanger Award for Translation Lymphoma Research. 00:01:06.107 --> 00:01:09.569 His research interests include biological subtypes 00:01:09.569 --> 00:01:14.199 of adult and pediatric non-Hodgkin's lymphoma as doctor, 00:01:14.199 --> 00:01:17.952 Miles is board certified in anatomic and clinical pathology 00:01:18.328 --> 00:01:22.165 with a subspecialty certification in hematology. 00:01:22.624 --> 00:01:25.335 I'll now turn our lecture over to Doctor Niles. 00:01:25.335 --> 00:01:25.835 Okay. 00:01:27.003 --> 00:01:29.089 Thank everyone for joining us today. 00:01:29.089 --> 00:01:33.551 Would be nice to all be in the same room, but that's, seems no longer practical. 00:01:33.551 --> 00:01:36.971 So today I'm going to talk about classification of leukemia and lymphoma 00:01:37.180 --> 00:01:41.976 and how the role of molecular testing has has really increased in this. 00:01:41.976 --> 00:01:46.189 The objectives are to discuss how molecular and genetic findings 00:01:46.189 --> 00:01:50.693 are used to provide more precise classifications of leukemia and lymphoma, 00:01:51.111 --> 00:01:54.114 and add prognostic information 00:01:54.906 --> 00:01:59.119 and and how the classification is really evolving 00:01:59.244 --> 00:02:03.331 with the more, more and more molecular side of genetic understanding we have. 00:02:03.748 --> 00:02:06.543 I also want to talk about clonal hematopoiesis, 00:02:06.543 --> 00:02:09.504 which is related to, 00:02:09.504 --> 00:02:12.966 mild dysplastic syndrome, which is a kind of myeloid cancer. 00:02:14.092 --> 00:02:17.428 And then there's going to be cases sprinkled throughout that show 00:02:17.428 --> 00:02:21.432 how, molecular insights, genetic data are used for classification. 00:02:24.853 --> 00:02:25.186 Okay. 00:02:25.186 --> 00:02:28.982 So in terms of genetically defined hematologic malignancies, 00:02:28.982 --> 00:02:32.026 chronic myeloid leukemia or CML was really the first 00:02:32.527 --> 00:02:36.656 in this diagnosis requires demonstration of the 922 translocation 00:02:36.656 --> 00:02:40.994 which is the Philadelphia chromosome and leads to overexpression. 00:02:41.202 --> 00:02:43.872 The BCR able fusion transcript. 00:02:43.872 --> 00:02:46.457 But in the 2001 version of the W.H.O. 00:02:46.457 --> 00:02:50.461 classification, that was really the only molecularly or genetically 00:02:50.461 --> 00:02:53.464 defined, neoplasm and hematology, 00:02:53.590 --> 00:02:56.426 but that's evolved quite a bit since then. 00:02:56.426 --> 00:02:59.429 Now we have acute leukemia. 00:02:59.804 --> 00:03:02.140 That, it includes 00:03:02.140 --> 00:03:05.560 subtypes with recurrent sided genetic changes and mutations. 00:03:06.102 --> 00:03:08.980 And these are used to, in some cases, 00:03:08.980 --> 00:03:11.983 diagnosed acute myeloid leukemia, 00:03:12.025 --> 00:03:14.527 when it would not fulfill morphologic criteria 00:03:14.527 --> 00:03:17.989 because there were less than 20% blast or more often, 00:03:17.989 --> 00:03:22.493 to more precisely classify the leukemias in the area of lymphoma. 00:03:22.493 --> 00:03:25.872 The role of molecular inside of genetic testing is is a bit more limited, but it's 00:03:25.914 --> 00:03:29.709 it's expanding and it's going to continue to to become more important. 00:03:32.003 --> 00:03:34.547 So acute myeloid leukemia 00:03:34.547 --> 00:03:37.884 these are all specific classifications at our current W.H.O. 00:03:37.926 --> 00:03:41.012 classification that are defined by the presence 00:03:41.012 --> 00:03:42.889 of a recurrent genetic abnormality. 00:03:42.889 --> 00:03:44.766 Take the first one for example. 00:03:44.766 --> 00:03:49.604 If this has an 821 translocation, which forms this fusion gene 00:03:50.688 --> 00:03:54.400 and that defines this is a specific subtype of AML. 00:03:54.400 --> 00:03:59.030 So as soon as you know it has that translocation, you move it out of the AML 00:03:59.030 --> 00:04:03.576 not otherwise specified category and into this specific subtype 00:04:03.576 --> 00:04:07.789 that has a 21 and that carries prognostic and therapeutic information. 00:04:08.456 --> 00:04:11.459 The ones with the asterisks by them are actually, 00:04:11.918 --> 00:04:16.756 ones where the the translocation is the most important defining, feature. 00:04:16.756 --> 00:04:20.468 And they don't even require 20% blasts to make the diagnosis, 00:04:20.468 --> 00:04:23.471 which has been the standard, 00:04:23.930 --> 00:04:28.268 you know, morphologic criteria for acute leukemia for, for many years. 00:04:31.229 --> 00:04:31.729 So this is 00:04:31.729 --> 00:04:35.525 a picture of a case with a lot of immature myeloid cells. 00:04:35.525 --> 00:04:36.943 And some of them show 00:04:36.943 --> 00:04:40.697 a real distinctive bilo phenotype with some prominent granulation. 00:04:41.114 --> 00:04:44.158 And some of them show prominent our rods. 00:04:45.285 --> 00:04:49.914 And then this again, a lot of these hyper granular blasts in here. 00:04:49.914 --> 00:04:54.043 So immature myeloid cells with some characteristic morphologic features 00:04:54.836 --> 00:04:58.298 that are highly suggestive of acute profile acidic leukemia. 00:04:58.756 --> 00:05:02.677 This is, an urgent diagnosis because these patients can present with, 00:05:03.553 --> 00:05:07.432 clotting disorder called DIC, where they have thrombocytopenia 00:05:07.432 --> 00:05:10.893 and not just the sites, but actually can form clots 00:05:11.686 --> 00:05:14.939 and this is really requires emergent treatment. 00:05:14.939 --> 00:05:18.818 So, diagnosis, you know, must be rendered as quickly as possible. 00:05:19.360 --> 00:05:22.822 These patients actually have a good prognosis if they're treated quickly. 00:05:23.698 --> 00:05:28.619 Especially if, the regimen includes, Astra or all trans retinoic acid. 00:05:29.078 --> 00:05:32.081 And this is distinct from other acute myeloid leukemias. 00:05:32.707 --> 00:05:35.918 And again, this this has a very good prognosis. 00:05:36.169 --> 00:05:37.712 But we don't make the diagnosis 00:05:38.963 --> 00:05:41.257 based on morphology alone. 00:05:41.257 --> 00:05:43.885 This new profile of Citic leukemia requires 00:05:43.885 --> 00:05:46.888 demonstration of the ra ra 00:05:47.263 --> 00:05:49.474 which is a 1517 translocation. 00:05:49.474 --> 00:05:53.394 So I'm showing here a fish assay where normally probes targeting 00:05:53.394 --> 00:05:57.315 the chromosome 15 and chromosome 17 loci are separated. 00:05:57.774 --> 00:06:01.402 But in this leukemia you get a fusion transcript where 00:06:01.736 --> 00:06:05.531 genes from chromosome 15 and chromosome 17 are fuzed together. 00:06:05.531 --> 00:06:09.786 And that's the abnormal finding that defines this subtype of acute leukemia. 00:06:10.536 --> 00:06:14.165 And this this can be turned around within 24 hours 00:06:14.916 --> 00:06:17.377 to, you know, quickly 00:06:17.377 --> 00:06:20.380 give this specific diagnosis and classification. 00:06:20.671 --> 00:06:23.674 And then it's possible to amplify this abnormal fusion. 00:06:23.674 --> 00:06:27.845 Transcript by PCR for disease monitoring and detection of minimal 00:06:27.845 --> 00:06:30.848 residual disease post therapy. 00:06:32.558 --> 00:06:34.685 So what about acute myeloid 00:06:34.685 --> 00:06:37.897 leukemia or AML that have normal cytogenetics. 00:06:37.897 --> 00:06:42.318 So the list I showed earlier all had specific defining translocations. 00:06:42.527 --> 00:06:46.656 But many cases of acute myeloid leukemia have a normal karyotype. 00:06:47.573 --> 00:06:51.452 So we now know that there are recurrent gene mutations. 00:06:51.452 --> 00:06:54.747 These are either point mutations or smaller mutations. 00:06:54.747 --> 00:06:57.750 And you can see in conventional cytogenetics. 00:06:58.000 --> 00:07:00.586 And these are also beginning to enter 00:07:00.586 --> 00:07:03.548 our classification that in 2017 00:07:03.840 --> 00:07:08.219 there is a unique category of AML with mutated NP one. 00:07:08.428 --> 00:07:11.472 I'll show an example of that. So these are 00:07:12.640 --> 00:07:14.016 going beyond sort 00:07:14.016 --> 00:07:18.813 of genetically defined AML and entering into molecularly defined AML. 00:07:19.730 --> 00:07:22.733 And this is really only in the past few years. 00:07:24.610 --> 00:07:26.195 So this is a case of a 54 year 00:07:26.195 --> 00:07:29.407 old woman who noticed increased bleeding while brushing her teeth. 00:07:30.283 --> 00:07:32.243 Dentist probably think of something different. 00:07:32.243 --> 00:07:35.246 But when I hear that, I think about most acidic leukemias. 00:07:35.371 --> 00:07:39.542 So she had, a normal white count, but there were circulating blasts. 00:07:40.042 --> 00:07:43.045 She's mildly anemic and thrombocytopenia 00:07:43.129 --> 00:07:46.132 and underwent a bone marrow evaluation. 00:07:46.632 --> 00:07:49.760 This is a high power view of the bone marrow aspirate smear. 00:07:49.760 --> 00:07:52.763 And you see too many immature cells here. 00:07:52.889 --> 00:07:55.892 A lot of them have sort of a folded nucleus. 00:07:56.142 --> 00:07:57.643 And you see these indentations. 00:07:57.643 --> 00:08:00.646 This is characteristic of immature monocytes. 00:08:00.771 --> 00:08:04.609 So we would morphologically diagnosis 00:08:04.609 --> 00:08:07.820 as an acute myeloid leukemia with an acidic differentiation. 00:08:08.863 --> 00:08:11.866 And there's also an abnormal red cell precursor here. 00:08:12.074 --> 00:08:16.078 So that's indicative of a background of dysplasia. 00:08:16.078 --> 00:08:19.040 These are normal red cell precursors here that have round nuclei. 00:08:19.040 --> 00:08:22.460 But this guy with the snowman look is really abnormal. 00:08:23.169 --> 00:08:26.589 So a background of dysplasia in acute myeloid leukemia 00:08:26.589 --> 00:08:29.592 can carry significance. 00:08:30.343 --> 00:08:32.261 So the amount of pathology 00:08:32.261 --> 00:08:35.598 workup starts with the morphologic differential and the aspirate smear. 00:08:35.598 --> 00:08:39.644 And there were 57% blast flow cytometry was performed 00:08:39.644 --> 00:08:43.356 and showed the phenotype of the myeloid blasts 00:08:43.689 --> 00:08:47.985 having myeloid markers CD 13 CD 33 and CD 117 00:08:49.529 --> 00:08:51.697 and then the in combination. 00:08:51.697 --> 00:08:54.700 This was used to make a diagnosis of acute myeloid leukemia 00:08:55.618 --> 00:08:57.912 and then fish came back. 00:08:57.912 --> 00:09:01.290 It usually takes between 1 and 4 days to get the full panel back. 00:09:01.832 --> 00:09:04.835 Our 1517 result comes out first within one day, 00:09:04.835 --> 00:09:07.338 and then the rest of the panel is about four days. 00:09:07.338 --> 00:09:12.969 But this AML fish panel was normal so that the most common recurrent sided 00:09:12.969 --> 00:09:15.972 genetic abnormalities were absent. 00:09:16.764 --> 00:09:20.142 And then seven days later, the conventional karyotype came back 00:09:20.142 --> 00:09:21.435 and it was also normal. 00:09:21.435 --> 00:09:23.813 So this is a normal karyotype. 00:09:23.813 --> 00:09:25.147 Acute myeloid leukemia. 00:09:26.190 --> 00:09:29.151 And then two weeks later 00:09:29.151 --> 00:09:33.155 a next generation sequencing myeloid panel came back and identified 00:09:34.073 --> 00:09:38.202 an NPM one mutation without three or BPA mutations. 00:09:38.202 --> 00:09:41.872 So this is one of the weaknesses of NGS 00:09:41.872 --> 00:09:44.917 testing right now is this long turnaround time. 00:09:45.084 --> 00:09:48.045 This can be compressed to to 10 to 12 days. 00:09:48.045 --> 00:09:51.465 But it's hard to get it get results much sooner than that. 00:09:54.260 --> 00:09:55.428 So based on, 00:09:55.428 --> 00:09:59.765 the morphology for cytometry, cytogenetics and molecular testing, 00:09:59.765 --> 00:10:03.769 this AML is classified as AML with mutated NPM one. 00:10:04.312 --> 00:10:07.732 We go back and put an amendment on our original pathology report 00:10:07.732 --> 00:10:09.775 that includes this precise classification. 00:10:09.775 --> 00:10:13.029 Because the initial diagnosis just went out as acute myeloid the chemo. 00:10:14.196 --> 00:10:17.158 Now in this case we did see significant dysplasia. 00:10:17.158 --> 00:10:19.869 And that's often associated with a worse prognosis. 00:10:19.869 --> 00:10:22.538 But now that we know there's an NPM one mutation, 00:10:23.497 --> 00:10:24.915 it's actually good news 00:10:24.915 --> 00:10:29.045 because displays in that setting is not associated with a worse prognosis. 00:10:29.837 --> 00:10:32.381 The other thing this mutation allows is 00:10:32.381 --> 00:10:36.218 it can be detected by our rt-PCR in quantitative. 00:10:36.218 --> 00:10:39.847 So we have a way to follow this patient for 00:10:41.265 --> 00:10:44.226 early disease recurrence or minimal residual disease 00:10:44.226 --> 00:10:47.229 after therapy because this will detect lower level 00:10:48.189 --> 00:10:50.650 then the NGS panel can detect. 00:10:50.650 --> 00:10:53.653 And finally it provides prognostic information. 00:10:55.571 --> 00:10:57.406 This is Kaplan-Meier plot 00:10:57.406 --> 00:11:00.368 for those who may not be very familiar with them, 00:11:00.660 --> 00:11:04.830 you basically start at time zero with 100% of the patients. 00:11:04.830 --> 00:11:07.833 In this case, we're looking at overall survival. 00:11:08.167 --> 00:11:12.296 And as you go out to say 60 months or five years, 00:11:13.005 --> 00:11:16.300 you can see that in this, these groups down here have less 00:11:16.300 --> 00:11:20.388 than 40% survival, where this group here has greater than 60% survival. 00:11:20.388 --> 00:11:23.724 So each one of these marks is basically a patient 00:11:23.724 --> 00:11:28.437 that has either died or been, far the center. 00:11:28.437 --> 00:11:31.273 But that's that's getting into too much detail there. 00:11:31.273 --> 00:11:35.319 So the important point here is that the characteristics, 00:11:35.528 --> 00:11:37.071 you know, as with this patient 00:11:37.071 --> 00:11:40.449 I described here, that is NP one positive and three negative, 00:11:41.033 --> 00:11:45.204 this is a better prognosis than if they do not have an NPM 00:11:45.204 --> 00:11:48.666 one mutation or if they had an NPM one mutation combined with the flip 00:11:48.708 --> 00:11:49.542 three mutation. 00:11:49.542 --> 00:11:54.463 So this is the prognostic information that we've gotten out of this classification. 00:11:54.505 --> 00:11:59.385 We're able to put this patient into a different strata prognostic. 00:11:59.760 --> 00:12:01.387 And this patient would not be considered 00:12:01.387 --> 00:12:05.433 for a bone marrow transplant whereas patients in this category would. 00:12:05.433 --> 00:12:09.645 So this is really guiding, therapy as well. 00:12:13.357 --> 00:12:16.569 So for acute leukemias, 00:12:16.569 --> 00:12:19.572 morphology is really still the first and most important step. 00:12:20.448 --> 00:12:23.993 Most cases are still diagnosed based on the blast percentage. 00:12:23.993 --> 00:12:27.079 So having more than 20% blast in the blood or bone marrow, 00:12:27.788 --> 00:12:30.166 besides genetics are really critical. Now. 00:12:30.166 --> 00:12:35.421 They define a few instances where AML can be diagnosed without increased blast. 00:12:35.671 --> 00:12:38.674 But more importantly, they provide prognostic information. 00:12:39.592 --> 00:12:42.553 Molecular testing has an emerging role 00:12:42.553 --> 00:12:47.892 where we now have specific classifications of AML that have normal cytogenetics, 00:12:47.892 --> 00:12:52.021 but then are put into a particular box based on their mutation profiling. 00:12:53.606 --> 00:12:56.484 And as always, there's a lot of ancillary testing 00:12:56.484 --> 00:12:59.862 that can be performed, but it really depends on the clinical situation. 00:13:00.112 --> 00:13:03.699 Is the patient Elbo eligible, physically 00:13:03.699 --> 00:13:07.286 fit enough to undergo curative intent therapy? 00:13:07.536 --> 00:13:12.583 If not, then really a lot of this testing could be considered, extraneous. 00:13:15.461 --> 00:13:16.962 Okay, so the next 00:13:16.962 --> 00:13:19.965 topic I want to jump in to is clonal hematopoiesis 00:13:20.758 --> 00:13:23.761 with and without side opinions. 00:13:23.844 --> 00:13:28.057 So with the proliferation 00:13:28.057 --> 00:13:33.395 of NGS panels and myeloid panels, there are now a lot of test results 00:13:33.395 --> 00:13:36.857 showing that some patients have, mutations 00:13:36.857 --> 00:13:39.860 in leukemia associated driver genes 00:13:40.152 --> 00:13:43.739 in, in patients that are otherwise apparently healthy. 00:13:43.739 --> 00:13:46.992 In other words, they have, you know, normal blood counts 00:13:46.992 --> 00:13:50.746 and wouldn't have been considered in any risk for a hematologic disease. 00:13:51.163 --> 00:13:53.958 But these patients were assessed for leukemia 00:13:53.958 --> 00:13:56.961 associated mutations as part of population studies. 00:13:57.169 --> 00:14:00.589 And found that, surprisingly, they do harbor 00:14:00.589 --> 00:14:02.758 a subset of these patients do harbored mutations, 00:14:02.758 --> 00:14:05.761 particularly in these three genes. Here, 00:14:06.011 --> 00:14:09.223 we know that these mutations are present in a clonal population 00:14:09.223 --> 00:14:12.977 of cells in these patients, because if it was only in a single cell, 00:14:12.977 --> 00:14:16.689 our assay would not be, sensitive enough to detect it. 00:14:16.689 --> 00:14:17.273 It has to, 00:14:19.066 --> 00:14:21.944 you know, it starts in a single cell, but that cell has to grow 00:14:21.944 --> 00:14:25.406 as a clonal population to be large enough to, to be detectable 00:14:26.824 --> 00:14:29.285 in the presence of such a, clonal 00:14:29.285 --> 00:14:32.746 population is associated with the risk of, of developing, 00:14:33.789 --> 00:14:35.833 usually myeloid neoplasms, 00:14:35.833 --> 00:14:38.836 sometimes lymphoid neoplasms. 00:14:38.878 --> 00:14:42.798 And interestingly, there's an association with cardiovascular events. 00:14:42.798 --> 00:14:45.801 We'll also talk about. 00:14:45.968 --> 00:14:48.554 So this graph just shows you the frequency 00:14:48.554 --> 00:14:52.516 of detecting a mutation over patient age. 00:14:52.516 --> 00:14:55.185 So take someone who is in their 50s. 00:14:55.185 --> 00:14:58.188 There is a less than 5% chance that you're going to find, 00:14:59.106 --> 00:15:02.109 a mutation in a patient in this age range. 00:15:02.526 --> 00:15:07.156 But once you get up into a patient that's in their 90s, this is now over 00:15:07.156 --> 00:15:11.118 ten to over 15% of the patients you would expect to find mutations in. 00:15:11.118 --> 00:15:15.581 So these mutations are accumulating, with age. 00:15:19.418 --> 00:15:20.461 So there's 00:15:20.461 --> 00:15:23.464 some important terminology to talk about here. 00:15:24.173 --> 00:15:26.592 In this clonal hematopoiesis world. 00:15:26.592 --> 00:15:32.139 So Chip is an acronym for clonal hematopoiesis of indeterminate potential. 00:15:32.264 --> 00:15:35.768 So the key here is that these patients are not site Apnic. 00:15:35.809 --> 00:15:40.064 So these are normal patients that were entered into a population study 00:15:40.606 --> 00:15:44.985 and assessed for myeloid mutations as a part of a research study. 00:15:44.985 --> 00:15:47.738 But they didn't present to a hematologist because anyone thought 00:15:47.738 --> 00:15:51.325 they had a hematologic disorder, or because they had a side of PDL. 00:15:52.910 --> 00:15:54.495 C course is a clonal site. 00:15:54.495 --> 00:15:56.497 Opinions of undetermined significance. 00:15:56.497 --> 00:16:00.417 So these patients have mutations, but they also have side opinions. 00:16:00.918 --> 00:16:03.545 So this is a most important group here. 00:16:03.545 --> 00:16:06.548 But we also know that there are patients that have side opinions 00:16:06.840 --> 00:16:08.801 that don't have mutations. 00:16:08.801 --> 00:16:12.346 So this is the idiopathic side opinions of undetermined significance. 00:16:13.180 --> 00:16:16.266 So it's kind of a lot of alphabet soup here. 00:16:16.475 --> 00:16:19.269 But you're basically looking at two factors. 00:16:19.269 --> 00:16:22.898 Does a patient have mutations and does a patient have side opinions. 00:16:22.898 --> 00:16:26.568 And the ones that have both mutations and side opinions, as you'll see 00:16:26.568 --> 00:16:31.365 in a little bit, are kind of the most interesting, in terms of, 00:16:32.408 --> 00:16:35.202 hematologic disorders. 00:16:35.202 --> 00:16:40.040 Now, just finding a clonal population and a mutation does not equal malignancy, 00:16:40.040 --> 00:16:45.546 because we know the incidence of, of finding mutations in healthy 00:16:45.546 --> 00:16:48.590 adults is a lot higher than the incidence of hematologic disorders. 00:16:49.508 --> 00:16:53.429 And finally, none of these qualify as a myelodysplastic syndrome 00:16:53.429 --> 00:16:57.182 because those require, morphologic or solid genetic evidence 00:16:57.182 --> 00:17:01.186 and by definition, all of these patients we're talking about here lack 00:17:01.186 --> 00:17:04.189 the criteria to be called a myelodysplastic syndrome. 00:17:06.567 --> 00:17:07.776 Okay, 00:17:07.776 --> 00:17:10.654 so this is a little bit dense, 00:17:10.654 --> 00:17:13.657 but we're really looking at 00:17:13.782 --> 00:17:17.411 two different population studies here the J.s and the MSI. 00:17:17.411 --> 00:17:19.288 The acronyms aren't important, but those are two different 00:17:19.288 --> 00:17:20.914 population studies. 00:17:20.914 --> 00:17:23.459 And if you look up here this is a group with no mutations. 00:17:23.459 --> 00:17:26.462 So they're set at a hazard ratio of one. 00:17:26.795 --> 00:17:31.759 And then the patients who had a mutation have a hazard ratio of developing 00:17:32.926 --> 00:17:36.263 a hematologic cancer in this case is 11 fold 00:17:36.263 --> 00:17:40.184 higher than these patients that don't have a mutation. 00:17:40.893 --> 00:17:45.272 So this is maybe easier to see down here when you're following the patients 00:17:45.272 --> 00:17:49.943 from time zero, forward for up to ten or more years. 00:17:50.235 --> 00:17:54.323 You can see that those patients that had a mutation at time zero 00:17:54.698 --> 00:17:58.994 have a nearly 50% chance of developing a hematologic cancer. 00:18:00.204 --> 00:18:05.459 This would be out about seven years or so versus those without a mutation. 00:18:05.459 --> 00:18:06.919 Have a have a low risk. 00:18:06.919 --> 00:18:10.631 There's still some risk, but it's a low risk of developing a hematologic cancer. 00:18:11.465 --> 00:18:14.718 So this is really telling us that the presence 00:18:14.718 --> 00:18:18.806 of these myeloid mutations in otherwise healthy people 00:18:19.431 --> 00:18:23.143 does not equal diagnosis of hematologic cancer, 00:18:23.143 --> 00:18:26.230 but it predicts the development of a hematologic cancer. 00:18:27.314 --> 00:18:29.566 And if the 00:18:29.566 --> 00:18:32.319 this is getting into the variant allele frequency. 00:18:32.319 --> 00:18:36.323 So, which is basically the percent estimate percent 00:18:36.615 --> 00:18:40.202 of the the cells that are mutated, if there is a higher proportion 00:18:40.202 --> 00:18:43.205 of the cells in the sample that are mutated, these have, 00:18:44.289 --> 00:18:45.999 rather I should say it the other way. 00:18:45.999 --> 00:18:49.795 Patients with the developing hematologic cancer had a higher variant 00:18:49.795 --> 00:18:52.798 allele frequency or a higher proportion of mutated cells 00:18:52.965 --> 00:18:56.051 than those who did not go on to develop a hematologic cancer. 00:19:01.098 --> 00:19:02.766 So this is just 00:19:02.766 --> 00:19:06.145 if we look, at all patients with side opinions. 00:19:06.353 --> 00:19:09.481 So these are patients that come with anemia or low platelets. 00:19:09.731 --> 00:19:13.527 And if you watch them over years, many of them about half 00:19:13.527 --> 00:19:16.530 are going to develop a hematologic cancer. 00:19:17.156 --> 00:19:18.824 But this is the interesting part. 00:19:18.824 --> 00:19:20.909 We can split those into two different groups. 00:19:20.909 --> 00:19:26.373 The red line that goes up like this is those patients that had side opinions 00:19:26.373 --> 00:19:29.501 but also had a mutation or more than one mutation. 00:19:29.501 --> 00:19:32.504 So these are the clonal side opinions of uncertain significance. 00:19:33.463 --> 00:19:34.798 Contrast those with the 00:19:34.798 --> 00:19:38.594 ICOs which is the idiopathic side opinions of uncertain significance. 00:19:38.594 --> 00:19:41.597 So these people have side opinions but they don't have a mutation. 00:19:42.431 --> 00:19:46.602 So over ten years they have a 10% risk of evolving 00:19:46.602 --> 00:19:51.231 to a hematologic cancer where it's over 90% for those with mutated. 00:19:51.231 --> 00:19:54.234 So it's clear that when we look in patients 00:19:54.276 --> 00:19:58.155 that are in the middle part of that Venn diagram with both side opinions 00:19:58.572 --> 00:20:04.036 and mutations, they're at a high risk for progressing to a hematologic cancer. 00:20:05.037 --> 00:20:07.372 And what this is showing you up here is that those 00:20:07.372 --> 00:20:10.375 with greater than two mutations have a very high risk. 00:20:10.542 --> 00:20:12.336 And then those with, 00:20:12.336 --> 00:20:15.339 you know, a higher risk than those with either 2 or 1 mutation. 00:20:15.631 --> 00:20:18.634 And then these patients with no mutations have the lowest risk. 00:20:19.301 --> 00:20:22.930 And this fourth panel is really they just split the mutations 00:20:22.930 --> 00:20:26.725 into ones that they classified as higher risk and lower risk. 00:20:26.725 --> 00:20:30.520 So the presence of any mutation as a carries a significant 00:20:30.520 --> 00:20:31.897 risk of a hematologic cancer. 00:20:31.897 --> 00:20:34.149 But some mutations are more significant than others. 00:20:37.236 --> 00:20:38.862 Okay. 00:20:38.862 --> 00:20:41.865 So the last couple slides we've all been talking about, 00:20:42.908 --> 00:20:47.537 mutations and whether or not patients are going to develop hematologic cancer. 00:20:48.121 --> 00:20:52.417 But what's really interesting, and I think a little bit, 00:20:52.960 --> 00:20:55.963 I would actually say quite surprising the first time I learned about this 00:20:56.421 --> 00:21:00.217 is that patients that have clonal hematopoiesis. 00:21:00.217 --> 00:21:01.134 So these are patients 00:21:01.134 --> 00:21:04.346 that have mutations detected, but they don't have side opinions. 00:21:05.138 --> 00:21:08.100 These patients are at a much higher risk to develop 00:21:08.100 --> 00:21:10.936 coronary heart disease and myocardial infarction. 00:21:10.936 --> 00:21:14.815 So let's look at this meta analysis 00:21:14.815 --> 00:21:18.443 just combines all of the data from two different studies. 00:21:18.944 --> 00:21:21.947 Those that have a mutation are two fold more likely 00:21:21.947 --> 00:21:24.950 to have coronary heart disease than those patients without. 00:21:26.201 --> 00:21:31.039 And again, down here in these these are two different studies. 00:21:31.039 --> 00:21:33.166 But if we look at those patients that are mutated, 00:21:33.166 --> 00:21:36.712 their four fold higher risk of having an early onset heart attack 00:21:36.712 --> 00:21:39.923 or myocardial infarction than those patients without a mutation. 00:21:40.841 --> 00:21:45.137 So these are not, you know, maybe the highest hazard ratios. 00:21:45.137 --> 00:21:49.308 But when we remember that cardiovascular disease is very common 00:21:49.516 --> 00:21:53.937 in North America, doubling your risk is actually really significant. 00:21:57.024 --> 00:22:00.444 So how do we detect these mutations? 00:22:00.444 --> 00:22:03.447 Well, most NGS sequencing panels 00:22:03.697 --> 00:22:07.242 will include these genes because these are the most common ones 00:22:07.242 --> 00:22:12.456 associated with clonal hematopoiesis as1 DNA three a and tattoo. 00:22:13.332 --> 00:22:15.751 This is a erupts myeloid NGS panel. 00:22:15.751 --> 00:22:18.754 But these will be found at any myeloid panel. 00:22:20.881 --> 00:22:24.801 Okay so here's really the key points to thinking 00:22:24.801 --> 00:22:29.056 about clonal hematopoiesis and clonal cytokine site opinions. 00:22:29.389 --> 00:22:33.435 So Chip is what you get when you run NGS on normal older patients. 00:22:33.727 --> 00:22:36.605 So we generally don't do that. 00:22:36.605 --> 00:22:39.608 But when it's done as a part of a research study, 00:22:39.983 --> 00:22:43.028 we did identify that there are patients that have 00:22:44.112 --> 00:22:47.908 mutated clones, even though they there otherwise appear healthy. 00:22:49.242 --> 00:22:54.039 And then the ICOs are those patients who have side opinions without mutations. 00:22:54.039 --> 00:22:55.248 And then these, I think, are 00:22:55.248 --> 00:22:58.251 the ones that are most interesting in the hematology world. 00:22:58.460 --> 00:22:59.669 Clonal cytokine is, 00:23:00.670 --> 00:23:02.506 of undetermined significance. 00:23:02.506 --> 00:23:07.344 So these are this basically ass acts like an early myelodysplastic syndrome. 00:23:07.344 --> 00:23:10.680 We don't see any dysplasia and we don't classify it as MDS. 00:23:10.680 --> 00:23:12.849 So there's no sign of genetic abnormalities. 00:23:12.849 --> 00:23:15.936 But they have side opinions and single gene mutations. 00:23:16.895 --> 00:23:21.316 And these patients progressed to other hematologic neoplasms. 00:23:21.566 --> 00:23:24.569 So this is really, 00:23:24.778 --> 00:23:28.657 similar to an early or low risk MDS. 00:23:29.157 --> 00:23:32.786 And I think in the future classification side opinions 00:23:32.786 --> 00:23:36.456 and a mutation will earn a specific classification. 00:23:37.332 --> 00:23:41.253 As a, as a form of mild dysplastic syndrome. 00:23:43.088 --> 00:23:46.091 And the other point to take away is that cardiovascular disease 00:23:46.174 --> 00:23:50.137 is much more common than MDS or other hematologic cancers. 00:23:50.137 --> 00:23:54.307 So this is really the biggest risk for patients with clonal hematopoiesis. 00:23:54.808 --> 00:23:58.895 And there are some theories as to why this is true. 00:23:59.146 --> 00:24:03.483 It probably involves abnormal monocytes and macrophage cells 00:24:03.483 --> 00:24:09.114 that are derived from these mutated clones and then are, somehow, 00:24:10.365 --> 00:24:11.575 contribute to an increase 00:24:11.575 --> 00:24:14.578 in inflammation in plaques in arteries. 00:24:14.619 --> 00:24:20.500 And there is some evidence that anti-inflammatory therapy can decrease 00:24:20.500 --> 00:24:25.088 the risk of a second heart attack in patients with, clonal hematopoiesis. 00:24:25.380 --> 00:24:28.925 So this tends to lend some, some support to this theory. 00:24:29.259 --> 00:24:34.222 And in the future there there may be merit to treating 00:24:35.348 --> 00:24:39.936 as a preventative therapy therapy, giving patients anti-inflammatory medications 00:24:40.270 --> 00:24:43.356 when they're identified as having clonal hematopoiesis. 00:24:43.356 --> 00:24:45.567 So this is sort of tongue in cheek. 00:24:45.567 --> 00:24:49.571 But right now we get our fasting lipid panel for cardiovascular risk. 00:24:49.696 --> 00:24:52.866 But will we in the future also get a myeloid NGS 00:24:52.866 --> 00:24:55.869 panel for cardiovascular risk. 00:24:57.871 --> 00:25:00.248 Okay I want to switch gears 00:25:00.248 --> 00:25:03.251 and talk about lymphoma. Now. 00:25:04.669 --> 00:25:07.380 So this is a diffuse large B-cell lymphoma. 00:25:07.380 --> 00:25:11.510 This is one of the most common types of the former that we encounter 00:25:11.510 --> 00:25:12.344 in North America. 00:25:13.637 --> 00:25:16.890 And it is defined really based on morphology. 00:25:16.890 --> 00:25:21.186 So this is diffuse and that we don't see any architecture or nodular to it. 00:25:21.186 --> 00:25:23.188 It's just a sheet of cells. 00:25:23.188 --> 00:25:26.024 And I've put an arrow on a small lymphocyte here. 00:25:26.024 --> 00:25:29.027 All of the neoplastic cells are 2 to 3 times 00:25:29.027 --> 00:25:31.071 at least the size of a small lymphocyte. 00:25:31.071 --> 00:25:33.657 So we have diffuse and we have large. 00:25:33.657 --> 00:25:37.118 So we've made the diagnosis of diffuse large lymphoma. 00:25:37.452 --> 00:25:41.331 And then if we add in a B-cell marker we usually use CD 20. 00:25:41.331 --> 00:25:44.709 This is immunohistochemistry that shows that these are all B cells. 00:25:44.960 --> 00:25:48.213 We've made the diagnosis of diffuse large B-cell lymphoma. 00:25:48.672 --> 00:25:51.550 So it's really just that simple. 00:25:51.550 --> 00:25:54.553 But the complexity all comes in here. 00:25:54.678 --> 00:25:58.807 All of these other ancillary tests are often 00:25:58.807 --> 00:26:02.686 performed in the diagnosis of diffuse large B-cell lymphoma. 00:26:02.686 --> 00:26:05.814 But you have to remember that it's really not for the diagnosis. 00:26:05.814 --> 00:26:10.235 All of these are providing either precise sub classification information 00:26:10.235 --> 00:26:14.030 or more importantly, prognostic information that will guide therapy. 00:26:14.823 --> 00:26:19.995 So this IBR is an in situ hybridization assay for Epstein-Barr virus. 00:26:21.037 --> 00:26:21.830 We're going to talk more 00:26:21.830 --> 00:26:25.709 about the fish assays for specific translocations a marker of proliferation. 00:26:25.709 --> 00:26:30.338 And then germinal center subtyping and MC and BCL two protein expression. 00:26:30.338 --> 00:26:34.926 But again I hammer this point home to the fellows 00:26:34.926 --> 00:26:37.929 is that the diagnosis is made here. 00:26:38.388 --> 00:26:41.391 And everything else you do after that is simply, 00:26:41.933 --> 00:26:44.561 additional prognostic information, information 00:26:44.561 --> 00:26:47.564 which may not always be indicated. 00:26:48.064 --> 00:26:48.857 Okay. 00:26:48.857 --> 00:26:51.526 So I want to start with the MC and BCL 00:26:51.526 --> 00:26:54.529 two rearrangements. 00:26:54.904 --> 00:26:57.907 First, well, I guess I have my intro slide here. 00:26:57.907 --> 00:26:59.743 I'm talking about both rearrangements. 00:26:59.743 --> 00:27:02.621 So these are gene rearrangements detected by fish. 00:27:02.621 --> 00:27:04.623 And then protein expression. 00:27:04.623 --> 00:27:07.626 And both of these contribute to two 00:27:07.917 --> 00:27:11.796 prognostic stratification of diffuse large B-cell lymphoma. 00:27:11.963 --> 00:27:16.635 So we if if both of the proteins are positive. 00:27:16.635 --> 00:27:20.555 So this is my immunohistochemistry for MC and for BCL two 00:27:20.555 --> 00:27:22.724 that we call it a double express or type. 00:27:22.724 --> 00:27:24.059 And I'll show examples of this. 00:27:24.059 --> 00:27:27.979 And then if both if both genes are translocated 00:27:28.313 --> 00:27:31.316 MC and BCL two or sometimes MC and BCL six, 00:27:31.524 --> 00:27:34.944 then we actually rename it a high grade B-cell lymphoma. 00:27:34.944 --> 00:27:35.945 That's a double hit. 00:27:37.197 --> 00:27:38.323 And these are uncommon. 00:27:38.323 --> 00:27:41.409 It's around 5% of all, diffuse large 00:27:41.451 --> 00:27:44.579 B-cell lymphoma that will fulfill these criteria. 00:27:44.579 --> 00:27:47.582 But I think this will get more clear as we go forward. 00:27:48.583 --> 00:27:51.711 So this is a specific entity 00:27:51.711 --> 00:27:54.756 in the WHL 2017, 00:27:54.756 --> 00:27:57.759 which is the most recent classification that we adhere to. 00:27:58.301 --> 00:28:01.513 And again, it's called high grade B-cell lymphoma with MC and BCL 00:28:01.513 --> 00:28:03.682 two and or BCL six rearrangements. 00:28:03.682 --> 00:28:07.602 So it's quite a mouthful, but these tend to present more aggressively 00:28:07.894 --> 00:28:09.396 with more disseminated disease. 00:28:09.396 --> 00:28:11.064 In the peripheral blood or bone marrow. 00:28:11.064 --> 00:28:14.609 They can look like bark at lymphoma, which is another highly aggressive type 00:28:14.609 --> 00:28:15.735 of lymphoma. 00:28:15.735 --> 00:28:18.738 And they often have a very complex karyotype. 00:28:19.197 --> 00:28:22.158 So it really looks like a disease that's different 00:28:22.158 --> 00:28:25.704 than the rest of the diffuse large B-cell lymphoma that we encounter. 00:28:27.706 --> 00:28:31.042 And the prognostic, significance is is the most important. 00:28:31.042 --> 00:28:32.711 This is progression free survival. 00:28:32.711 --> 00:28:36.005 But we'll just focus on the overall survival on the side. 00:28:36.756 --> 00:28:41.094 So this middle line here is the red line is a double hit. 00:28:41.386 --> 00:28:44.264 So this double or triple hit. 00:28:44.264 --> 00:28:47.434 So MC is translocated as well 00:28:47.434 --> 00:28:50.437 as BCL two BCL six or both. 00:28:50.520 --> 00:28:52.605 And that's this bottom line here. 00:28:52.605 --> 00:28:56.860 So again looking at a Kaplan-Meier curve fewer of these patients 00:28:56.860 --> 00:29:02.574 are alive at 24 months than patients that don't have a Mic translocation 00:29:03.074 --> 00:29:06.327 or that only have a Mic translocation in this middle line. 00:29:06.327 --> 00:29:09.330 So they don't have BCL six or BCL two. 00:29:10.206 --> 00:29:14.919 So what we're really trying to do when we perform fish on these lymphomas 00:29:14.919 --> 00:29:19.924 is to identify these patients here that are dying rapidly in the first year. 00:29:20.425 --> 00:29:22.719 Compare that to the patients without translocation. 00:29:22.719 --> 00:29:25.430 So this is really the aggressive disease behavior. 00:29:26.556 --> 00:29:27.766 And these patients should 00:29:27.766 --> 00:29:30.769 be treated differently. 00:29:31.770 --> 00:29:33.062 Okay. 00:29:33.062 --> 00:29:38.485 So all we're showing here again would ignore the progression free survival. 00:29:38.485 --> 00:29:42.280 And look at overall survival is that it doesn't really matter 00:29:42.280 --> 00:29:47.869 if you have a double hit with BCL two, a double hit with BCL six or a triple hit. 00:29:48.203 --> 00:29:51.122 And remember, MC is always one of the hits, so 00:29:51.122 --> 00:29:54.125 the prognosis is bad for any of those. 00:29:54.125 --> 00:29:57.128 So we are lumping them together. 00:30:00.256 --> 00:30:03.259 Now this is newer, 00:30:03.426 --> 00:30:06.429 data that I think is making things more complicated. 00:30:07.055 --> 00:30:09.557 And so it turns out that just whether or not 00:30:09.557 --> 00:30:13.353 you have a MC translocation may not be enough information. 00:30:13.978 --> 00:30:17.315 MC can translate with a variety of different partners 00:30:17.857 --> 00:30:20.652 when it translocated with an immunoglobulin gene. 00:30:20.652 --> 00:30:22.362 So these are the genes that make up, 00:30:23.363 --> 00:30:24.447 B-cell receptors. 00:30:24.447 --> 00:30:27.784 So you have a heavy chain and a kappa or lambda light chain. 00:30:28.117 --> 00:30:33.414 So this is the these are all lumped together as immunoglobulin genes. 00:30:33.414 --> 00:30:34.833 And when MC is translocated 00:30:34.833 --> 00:30:38.044 to an immunoglobulin gene, these patients do very poorly 00:30:38.503 --> 00:30:42.799 when you have an MC translocation, even if it's a double hit or triple hit. 00:30:43.216 --> 00:30:45.760 But the MC partner is not an immunoglobulin. 00:30:45.760 --> 00:30:50.098 These patients don't do any worse than those that lack an MC translocation. 00:30:50.682 --> 00:30:53.685 So this is going to make life more complicated for us. 00:30:54.602 --> 00:30:57.105 If it's borne out in other studies. 00:30:57.105 --> 00:31:00.441 We use a MC break apart assay which detects 00:31:00.733 --> 00:31:03.736 MC translocations, but it doesn't tell us who it's partnered with, 00:31:04.195 --> 00:31:07.240 and we may need to look in more detail on the future 00:31:07.240 --> 00:31:10.702 and demonstrate whether or not MC is partnered with IG, 00:31:11.661 --> 00:31:12.370 immunoglobulin 00:31:12.370 --> 00:31:15.456 kappa, or immunoglobulin Lambda to really get 00:31:15.665 --> 00:31:18.668 at this subset of patients. 00:31:19.335 --> 00:31:20.920 And the reason that makes biologic 00:31:20.920 --> 00:31:24.465 sense is immunoglobulins are very highly expressed in B cells. 00:31:24.465 --> 00:31:28.887 So if you put the MC team behind the promoter or enhancer 00:31:29.596 --> 00:31:32.473 of an immunoglobulin gene, you're going to get very high 00:31:32.473 --> 00:31:35.476 MC expression. 00:31:36.144 --> 00:31:37.312 Okay. 00:31:37.312 --> 00:31:42.859 So now we're going to turn away from the translocations that are detected by fish. 00:31:42.859 --> 00:31:43.526 And we're going to look 00:31:43.526 --> 00:31:47.739 for protein expression which is detected by immunohistochemistry. 00:31:48.031 --> 00:31:50.325 We're still talking about the same two genes. 00:31:50.325 --> 00:31:53.286 But now we're looking at protein expression. 00:31:53.369 --> 00:31:57.123 And the reason this is interesting is this if you look at these patients here 00:31:57.123 --> 00:32:00.376 that express both MEK and BCL two protein, they do quite a bit 00:32:00.376 --> 00:32:05.506 worse than those patients that express one or the other, or neither. 00:32:06.341 --> 00:32:08.801 And this is overall survival. 00:32:08.801 --> 00:32:12.680 And it looks like if you only look at MC expression 00:32:12.680 --> 00:32:16.017 for example, and ignore BCL two, that these patients do worse. 00:32:16.601 --> 00:32:20.480 But here is when we look at the BCL two negative cases. 00:32:20.480 --> 00:32:23.107 So this excludes the double express. 00:32:23.107 --> 00:32:27.153 Then single expression of MEK really doesn't do any worse than MC negative. 00:32:27.570 --> 00:32:29.739 The same is true for BCL two. 00:32:29.739 --> 00:32:32.825 So again the important inclusion here is that 00:32:32.825 --> 00:32:36.704 when patients express both MEK and BCL two in their lymphoma, 00:32:36.704 --> 00:32:41.000 they do worse if they have one or the other or neither, they do better. 00:32:45.463 --> 00:32:47.090 So this is 00:32:47.090 --> 00:32:50.718 this is another thing that's that's interesting, at least to us, 00:32:50.927 --> 00:32:53.930 lymphoma oriented people, 00:32:53.972 --> 00:32:57.016 is that there are two different classifications of 00:32:58.184 --> 00:33:00.853 diffuse large B-cell lymphoma I haven't really talked about yet, 00:33:00.853 --> 00:33:05.191 and I don't want to go into too much detail on, but they're called activated 00:33:05.191 --> 00:33:08.194 B cell type in the germinal center B cell type. 00:33:08.444 --> 00:33:11.447 And these have been known about for about 20 years, 00:33:11.823 --> 00:33:14.826 but we still don't really have any different therapies. 00:33:15.368 --> 00:33:18.246 And that this prognosis differences 00:33:18.246 --> 00:33:21.249 has been important for that long. 00:33:21.374 --> 00:33:23.167 And we've been focused on this 00:33:23.167 --> 00:33:26.170 activated B cell type, which has a worse prognosis. 00:33:26.170 --> 00:33:30.133 But it turns out that if you look at 00:33:30.133 --> 00:33:33.928 if you exclude the double express or lymphomas, the activated B cell ones 00:33:34.929 --> 00:33:37.807 actually do, as well as the germinal center B cell type. 00:33:37.807 --> 00:33:39.600 So why is this important? 00:33:39.600 --> 00:33:44.230 Well, for 20 years we've been trying to split these apart. 00:33:45.106 --> 00:33:48.943 And it's challenging sometimes because 00:33:50.069 --> 00:33:53.031 it's a gene expression profiling based assay. 00:33:53.031 --> 00:33:56.325 And we have to use surrogate immunohistochemistry which doesn't work 00:33:56.576 --> 00:33:57.994 as accurately. 00:33:57.994 --> 00:34:01.873 So we've been trying to classify these and separate them 00:34:01.873 --> 00:34:03.249 for prognostic significance. 00:34:03.249 --> 00:34:07.128 But it turns out that probably looking at the double expression of MC 00:34:07.128 --> 00:34:10.548 and BCL two is a more effective way to identify the negative 00:34:10.548 --> 00:34:13.551 prognosis in this, subtype of lymphoma. 00:34:14.052 --> 00:34:17.305 So I recognize that's getting a little bit into the lymphoma weeds. 00:34:17.305 --> 00:34:20.308 So we'll we'll move forward. 00:34:21.350 --> 00:34:23.394 This is, case. 00:34:23.394 --> 00:34:25.188 It's a 79 year old woman. 00:34:25.188 --> 00:34:28.566 She had a recent breast cancer diagnosis and enlarged lymph nodes. 00:34:29.025 --> 00:34:31.986 This happens to be from the right neck. 00:34:31.986 --> 00:34:35.281 Actually, the assumption or concern is that this would be metastatic 00:34:35.281 --> 00:34:37.283 breast cancer. 00:34:37.283 --> 00:34:39.702 We see diffuse architecture here. 00:34:39.702 --> 00:34:43.456 And when we look on high power, we see large atypical lymphoid cells. 00:34:43.456 --> 00:34:46.876 And large is objective based on them 00:34:46.876 --> 00:34:50.004 being considerably larger than this small lymphocyte here 00:34:50.171 --> 00:34:54.342 at least three times the size in the case of the majority of these cells. 00:34:54.342 --> 00:34:57.345 So we have diffuse and large 00:34:57.470 --> 00:35:00.098 and we do a CD 20 B cell marker. 00:35:00.098 --> 00:35:04.018 And we can make the diagnosis of a diffuse large B-cell lymphoma. 00:35:05.436 --> 00:35:06.145 This is 00:35:06.145 --> 00:35:09.565 one which just shows that the cells are rapidly dividing. 00:35:09.565 --> 00:35:12.944 So it's a that correlates with that being an aggressive lymphoma. 00:35:13.820 --> 00:35:16.030 And then this is BCL two and MC 00:35:16.030 --> 00:35:19.617 immunohistochemistry again looking at protein expression. 00:35:19.617 --> 00:35:24.330 And you can see that this lymphoma is positive for both BCL two and MC. 00:35:24.789 --> 00:35:27.792 So that puts it into that double express or 00:35:28.334 --> 00:35:29.293 category. 00:35:29.293 --> 00:35:31.921 We also looked for MC translocations. 00:35:31.921 --> 00:35:34.465 We did a break apart probe assay and it was negative. 00:35:34.465 --> 00:35:37.468 So that means there's no MC rearrangement. 00:35:38.136 --> 00:35:41.806 And you can't have a double hit if you don't have a MC rearrangement 00:35:41.806 --> 00:35:42.473 to start with. 00:35:42.473 --> 00:35:46.644 So we didn't look for BCL two or BCL six because they were already excluded 00:35:46.644 --> 00:35:48.646 by this negative result. 00:35:48.646 --> 00:35:51.691 So we classified this as a diffuse large B-cell lymphoma. 00:35:51.941 --> 00:35:54.485 It had an activated B-cell or ABC 00:35:55.987 --> 00:35:58.197 immuno profile that I didn't show you, 00:35:58.197 --> 00:36:01.534 and it has co-expression of MC and BCL two. 00:36:02.952 --> 00:36:06.706 So this next slide will show the significance. 00:36:06.706 --> 00:36:07.915 There. 00:36:07.915 --> 00:36:10.918 This patient would be would fit on the yellow line here 00:36:10.918 --> 00:36:13.963 in the middle where they have MC and BCL two protein expression. 00:36:14.380 --> 00:36:16.299 But they're not on this line 00:36:16.299 --> 00:36:19.802 which is where they have MC and BCL two translocations. 00:36:20.136 --> 00:36:22.680 So these are the worst prognosis patients. 00:36:22.680 --> 00:36:24.807 This patient would be in the middle. 00:36:24.807 --> 00:36:29.312 And the patients who do best lack translocations or double expression. 00:36:29.854 --> 00:36:32.857 But this guides therapy because 00:36:34.734 --> 00:36:38.070 Our clinicians now will University of Utah, 00:36:38.070 --> 00:36:41.741 they'll use a different regimen called dose adjusted epoch R, 00:36:41.866 --> 00:36:44.827 which is this one, instead of our Chop, which is a more traditional, 00:36:45.703 --> 00:36:47.955 diffuse large B-cell regimen. 00:36:47.955 --> 00:36:51.125 And what that means is, based on the information 00:36:51.125 --> 00:36:53.169 we've provided, that this is a double express, 00:36:53.169 --> 00:36:56.172 or these patients will actually get a different therapy. 00:36:56.172 --> 00:36:57.965 And this curve actually moves up. 00:36:57.965 --> 00:37:00.593 So this patient will have a better chance based on the 00:37:00.593 --> 00:37:02.595 the information we've provided here. 00:37:06.557 --> 00:37:08.184 So this is 00:37:08.184 --> 00:37:13.272 looks complicated because it kind of is and this is the strategy 00:37:13.272 --> 00:37:17.526 that we try to follow here at RCP when we're, 00:37:18.486 --> 00:37:21.489 doing prognostic testing on diffuse large B-cell lymphoma. 00:37:21.489 --> 00:37:24.659 So the first step is it really needs to be 00:37:24.992 --> 00:37:27.662 a de novo diagnosis of Dlbcl. 00:37:27.662 --> 00:37:30.665 If it's relapsed disease or post-transplant, then for prolific 00:37:30.706 --> 00:37:34.627 disorder or transformation of liquid or lymphoma or something. 00:37:35.461 --> 00:37:38.839 These are all settings in which you can encounter Dlbcl. 00:37:39.382 --> 00:37:41.884 But the prognostic significance of all 00:37:41.884 --> 00:37:44.887 this testing is really uncertain in those settings. 00:37:45.096 --> 00:37:49.141 And then if you have different subtypes of diffuse large B-cell lymphoma, like a T 00:37:49.141 --> 00:37:53.271 cell history assay rich or EBV positive or primary mediastinal 00:37:53.271 --> 00:37:56.274 or primary central nervous system, 00:37:56.357 --> 00:37:59.277 the testing is not indicated in any of those. 00:37:59.277 --> 00:38:02.822 So once we've excluded all of these things, we're in the category 00:38:02.822 --> 00:38:04.699 of diffuse large B-cell Inos. 00:38:05.741 --> 00:38:07.451 So we'll start with the fish. 00:38:07.451 --> 00:38:08.911 We perform mic fish. 00:38:08.911 --> 00:38:11.539 And if it's positive for a MiG translocation 00:38:11.539 --> 00:38:14.542 we will add the BCL two and BCL six probes. 00:38:15.001 --> 00:38:19.422 If one of them is positive as well, or both will be classifying 00:38:19.422 --> 00:38:24.093 this as a high grade B-cell lymphoma with MC and BCL two or BCL six. 00:38:24.427 --> 00:38:27.930 And again, that's the most aggressive type of lymphoma 00:38:27.930 --> 00:38:30.933 in this area. 00:38:31.058 --> 00:38:34.603 If not, it goes back over here to not otherwise specified. 00:38:34.603 --> 00:38:38.858 So then we're at the same time doing immunohistochemistry for MC and BCL two. 00:38:39.734 --> 00:38:44.071 And we also do, the marker CD ten, BCL six, and mom one. 00:38:44.822 --> 00:38:49.201 This is referred to as the Hahn's classification of cell of origin. 00:38:49.201 --> 00:38:51.495 But it's really getting back to the 00:38:51.495 --> 00:38:55.541 activated B-cell or germinal center subtypes. So, 00:38:56.542 --> 00:39:00.046 if MC and BCL two are negative, it stays as this and OS. 00:39:00.046 --> 00:39:03.049 But if MC and B B-cell two protein they're positive 00:39:03.174 --> 00:39:06.344 we will classify it as diffuse large B-cell lymphoma. 00:39:06.344 --> 00:39:08.346 Double suppressor. 00:39:08.346 --> 00:39:11.140 So there's a lot to go through there. 00:39:11.140 --> 00:39:12.683 It's a complicated algorithm. 00:39:12.683 --> 00:39:16.604 But at the end of the day, this is a strategy that provides the best 00:39:16.604 --> 00:39:19.815 prognostic information we can for these patients. 00:39:22.985 --> 00:39:25.363 So when should we 00:39:25.363 --> 00:39:28.366 perform fish on the few B B-cell lymphoma? 00:39:29.075 --> 00:39:33.329 We just we talk about this a lot in our group with our clinicians. 00:39:33.329 --> 00:39:35.414 And I get this question a lot. 00:39:35.414 --> 00:39:37.917 So really our practice here 00:39:37.917 --> 00:39:40.920 is to perform fish for MC on 00:39:41.128 --> 00:39:45.007 on almost all of our new diagnosis of diffuse large B-cell lymphoma. 00:39:45.091 --> 00:39:48.135 And then if it's positive we follow up with BCL two and BCL six. 00:39:48.719 --> 00:39:50.805 But you have to consider the clinical context. 00:39:50.805 --> 00:39:54.809 Will it really change the clinical approach or therapy. 00:39:55.142 --> 00:39:57.937 And the particular examples 00:39:57.937 --> 00:40:02.066 you know, that come to mind if it's a, you know, a very elderly patient who's, 00:40:02.817 --> 00:40:06.278 otherwise has comorbid comorbidities and is unwell, 00:40:06.737 --> 00:40:10.366 they're not going to be able to go through the chemotherapy 00:40:10.366 --> 00:40:13.119 that's required for curative intent. 00:40:13.119 --> 00:40:17.665 So the prognostic, information of the fish assays etc. 00:40:17.957 --> 00:40:20.000 is really based on curative intent therapy. 00:40:20.000 --> 00:40:24.088 So in this case it wouldn't really provide any useful information. 00:40:24.713 --> 00:40:27.716 And all of this ancillary testing is expensive. 00:40:27.716 --> 00:40:31.512 So I think we need to think about that before we order at all. 00:40:33.472 --> 00:40:35.433 Okay. 00:40:35.433 --> 00:40:38.727 So for the lymphoma, Dlbcl conclusions. 00:40:38.727 --> 00:40:42.523 The diagnosis again requires only morphology and immuno phenotype. 00:40:43.441 --> 00:40:47.903 The, recognizing this W.H.O. 00:40:48.028 --> 00:40:53.200 to 2017 category of high grade B-cell lymphoma requires fish. 00:40:53.200 --> 00:40:57.580 And this qualifies it really as a genetically defined lymphoma. 00:40:59.206 --> 00:41:01.167 But this testing should really only be performed 00:41:01.167 --> 00:41:04.170 if it will affect patient care. 00:41:04.545 --> 00:41:05.504 Okay. 00:41:05.504 --> 00:41:09.425 So moving to, another case here. 00:41:09.884 --> 00:41:12.428 Really interesting case from, 00:41:12.428 --> 00:41:15.306 your year and a half ago. 00:41:15.306 --> 00:41:17.016 Here is the tissue from low power. 00:41:17.016 --> 00:41:21.103 And we can really see no residual architecture that looks like a lymph node. 00:41:21.103 --> 00:41:23.230 So we know that it's diffuse. 00:41:23.230 --> 00:41:27.026 When we look at it closely, we can see a lot of very large cells. 00:41:28.486 --> 00:41:30.196 When we do our B-cell marker. 00:41:30.196 --> 00:41:33.449 All of these large cells are negative for CD 20. 00:41:33.449 --> 00:41:36.452 So this is not a B-cell lymphoma. 00:41:36.452 --> 00:41:38.454 So it gets more interesting. 00:41:39.455 --> 00:41:41.624 This is CD3 which is a T cell marker. 00:41:41.624 --> 00:41:44.502 Now only some of the large cells are positive. 00:41:44.502 --> 00:41:45.878 Many of them are still negative. 00:41:45.878 --> 00:41:49.048 But this is a clue that we might be dealing with a T cell lymphoma. 00:41:50.466 --> 00:41:53.385 So we did additional markers along those lines. 00:41:53.385 --> 00:41:54.803 This is CD 30. 00:41:54.803 --> 00:41:57.932 And it is strongly and diffusely positive in this lymphoma. 00:41:58.390 --> 00:42:00.976 And CD two which is another T cell 00:42:00.976 --> 00:42:03.979 marker is also strongly and diffusely positive. 00:42:04.647 --> 00:42:06.023 And this is T1. 00:42:06.023 --> 00:42:08.776 This is a cytotoxic T cell marker. 00:42:08.776 --> 00:42:12.279 So now we've got additional T cell markers with the CD 30 00:42:13.030 --> 00:42:16.534 showing us that this is a CD 30 positive T cell lymphoma. 00:42:17.034 --> 00:42:19.537 And in young patients we often think about 00:42:19.537 --> 00:42:22.581 ALK positive anaplastic large cell lymphoma. 00:42:22.581 --> 00:42:24.583 But ALK one was negative in this case. 00:42:26.168 --> 00:42:27.127 So again 00:42:27.127 --> 00:42:30.548 it's positive for CD2, CD 30 and T one. 00:42:30.548 --> 00:42:32.258 So we know it's a T cell. 00:42:32.258 --> 00:42:35.261 In addition it had CD 45 which is 00:42:36.220 --> 00:42:38.264 leukocyte common antigen expressed 00:42:38.264 --> 00:42:41.267 on all types of leukocytes. 00:42:41.350 --> 00:42:44.270 It had CD4 and CD seven negative for all the B 00:42:44.270 --> 00:42:48.107 cell markers and some T cell markers, CD eight and CD five. 00:42:48.107 --> 00:42:51.735 So the diagnosis based on morphology and immuno phenotype 00:42:51.735 --> 00:42:55.114 is anaplastic large cell lymphoma ALK one negative. 00:42:56.115 --> 00:42:59.118 We considered Hodgkin lymphoma 00:42:59.868 --> 00:43:02.496 which would have been more common in this age group. 00:43:02.496 --> 00:43:03.706 But it doesn't look like it. 00:43:03.706 --> 00:43:05.958 By morphology. It's negative for Pax five. 00:43:05.958 --> 00:43:07.376 It's negative for CD 15. 00:43:07.376 --> 00:43:10.337 And it did have CD three expression. 00:43:10.337 --> 00:43:12.923 The other consideration in the T cell lymphoma 00:43:12.923 --> 00:43:16.051 world would be peripheral T cell lymphoma not otherwise specified. 00:43:16.594 --> 00:43:22.182 But with this, degree of CD 30 expression, we really can't call it that. 00:43:22.182 --> 00:43:26.979 It needs to be in the ALK, alk negative alk cll category. 00:43:29.440 --> 00:43:33.360 So additional studies now several years ago we would have been done. 00:43:33.360 --> 00:43:36.363 This would have gone out as an ALK negative LCL. 00:43:36.488 --> 00:43:39.491 But now we know there's more we can do. So 00:43:41.869 --> 00:43:44.413 we performed fish for, 00:43:44.413 --> 00:43:48.167 this dust 22 IRA for rearrangement, and I've blown it up here. 00:43:48.167 --> 00:43:50.210 It's on chromosome six, 00:43:50.210 --> 00:43:53.589 and there are probes on either side of both of these genes. 00:43:53.589 --> 00:43:56.383 So the rearrangement can involve either one of these genes. 00:43:56.383 --> 00:43:58.427 And it will be detected with this probe. 00:43:58.427 --> 00:44:01.555 And then we did an immunohistochemical stain for P60 three. 00:44:01.555 --> 00:44:04.558 So why did we do those things. 00:44:04.725 --> 00:44:06.977 Well this is what we used to know 00:44:06.977 --> 00:44:10.856 is that the lymphomas that were ALK positive, this is again 00:44:10.856 --> 00:44:14.443 just ALK anaplastic large cell lymphomas that are out positive 00:44:14.610 --> 00:44:19.156 do a lot better than anaplastic large cell lymphomas that are ALK negative. 00:44:19.948 --> 00:44:22.576 So this we've known this for for some time. 00:44:22.576 --> 00:44:24.370 And this was always the important watershed. 00:44:24.370 --> 00:44:26.955 Is it ALK positive or ALK negative. 00:44:26.955 --> 00:44:29.541 But now things are getting a little bit more complicated. 00:44:29.541 --> 00:44:34.672 So again the ALK positives are this black line that's dotted here. 00:44:34.672 --> 00:44:36.006 So they still do really well. 00:44:36.006 --> 00:44:40.386 But it turns out if you're ALK negative but have a dust 22 rearrangement 00:44:40.844 --> 00:44:44.473 you do just as well as and ALK negative LCL, I'm sorry. 00:44:44.473 --> 00:44:47.142 As in ALK positive CL alk. 00:44:47.142 --> 00:44:50.771 And so this red dashed line is the anaplastic large 00:44:50.771 --> 00:44:54.733 cell on problems that are negative for ALK and negative for dust 22 00:44:55.234 --> 00:44:59.196 so these are what's left of the negative ALK CLS. 00:44:59.738 --> 00:45:02.950 But we've moved some of them up into the good prognosis category. 00:45:02.950 --> 00:45:04.868 So it's really significant. 00:45:04.868 --> 00:45:08.330 This last line is an uncommon subtype 00:45:08.539 --> 00:45:11.375 that has a rearrangement of TP 63. 00:45:11.375 --> 00:45:14.169 And those those patients do very poorly. 00:45:14.169 --> 00:45:17.589 But it's a lot less common than the other, categories. 00:45:19.341 --> 00:45:21.427 So this is from that same paper. 00:45:21.427 --> 00:45:26.223 It shows us basically the four categories of anaplastic large cell lymphoma 00:45:26.473 --> 00:45:27.516 that we deal with now. 00:45:27.516 --> 00:45:30.978 So they all have CD 30 strong and diffuse. 00:45:31.520 --> 00:45:33.105 This one down here has ALK. 00:45:33.105 --> 00:45:36.400 So this is an ALK positive LCL while all the others are negative. 00:45:37.568 --> 00:45:40.237 This one is and ALK negative. 00:45:40.237 --> 00:45:42.072 But it has a dust 22 arrangement. 00:45:42.072 --> 00:45:45.492 So it's prognosis is just as good as the ALK positive LCL. 00:45:45.492 --> 00:45:48.162 And that's kind of the new information here. 00:45:48.162 --> 00:45:51.540 This is the very uncommon subtype that has P 63 00:45:52.332 --> 00:45:55.586 translocation shown by fish and overexpression of the protein. 00:45:55.586 --> 00:45:58.172 And these two are the ones that do very poorly. 00:45:58.172 --> 00:46:01.175 And then those that are negative for all the above, 00:46:02.176 --> 00:46:05.471 art are the sort of intermediate poor prognosis. 00:46:07.806 --> 00:46:10.809 Which is this dashed line here. 00:46:11.560 --> 00:46:12.936 So this is what this is 00:46:12.936 --> 00:46:16.899 the current, state of the art for anaplastic large cell lymphoma, 00:46:16.899 --> 00:46:20.819 which is really evolved beyond just ALK positive and negative. 00:46:21.695 --> 00:46:25.491 So when we're worrying about ALK positive LCL, of course, we know that 00:46:25.783 --> 00:46:30.245 detecting ALK expression by imaging has the chemistry makes the diagnosis. 00:46:30.829 --> 00:46:33.540 There are different patterns, 00:46:33.540 --> 00:46:37.002 the common lymphocytes, the acidic small cell and Hodgkin like pattern. 00:46:37.586 --> 00:46:41.256 These are important to be aware of for human pathologists because 00:46:41.507 --> 00:46:43.592 they can look a little bit unusual. 00:46:43.592 --> 00:46:46.637 And if you're not thinking about it, they you could potentially 00:46:46.637 --> 00:46:49.640 miss this, especially like a small cell. 00:46:49.807 --> 00:46:51.850 It's ironic that that name is, 00:46:51.850 --> 00:46:55.062 anaplastic large cell lymphoma, but there's a small cell variance. 00:46:55.062 --> 00:46:56.897 So you can see how that could get confusing. 00:46:58.690 --> 00:46:59.650 ALK negative 00:46:59.650 --> 00:47:03.278 CLS really look like an ALK positive LCL with large, 00:47:04.488 --> 00:47:08.075 very atypical diffuse cells, but they're negative for ALK. 00:47:08.826 --> 00:47:11.245 And there aren't variants like small cell 00:47:11.245 --> 00:47:14.248 or Hodgkin like recognized for ALK negative. 00:47:14.873 --> 00:47:17.876 And then finally the the P 63. 00:47:19.169 --> 00:47:21.338 So because this is rare, 00:47:21.338 --> 00:47:24.716 we have not yet developed a fish assay in our laboratory for this. 00:47:24.716 --> 00:47:29.221 And we use the immunohistochemistry for P 63 protein as a surrogate marker for 00:47:29.263 --> 00:47:32.891 because it's always going to be expressed in cases that have the rearrangement. 00:47:33.392 --> 00:47:36.728 But sometimes it'll be positive in cases without the rearrangements. So 00:47:38.230 --> 00:47:41.024 if we see expression 00:47:41.024 --> 00:47:45.445 then we can proceed to do the fish assay as a send out test. 00:47:45.445 --> 00:47:46.530 But if there's no expression 00:47:46.530 --> 00:47:50.617 then the the rearrangement of TP 63 is excluded and we can move on. 00:47:50.617 --> 00:47:52.244 It has good negative predictive value. 00:47:55.414 --> 00:47:58.417 Okay. 00:47:58.458 --> 00:47:59.793 Next case is a 17 year 00:47:59.793 --> 00:48:02.796 old girl with enlarged tonsils. 00:48:02.880 --> 00:48:04.965 This is from the original pathology report. 00:48:04.965 --> 00:48:06.884 It came to us in consultation. 00:48:06.884 --> 00:48:09.845 One tonsil they thought was a high grade lymphoma. 00:48:10.137 --> 00:48:13.181 The other looked reactive follicular hyperplasia. 00:48:14.016 --> 00:48:17.019 So I don't like the terminology high grade lymphoma in this case 00:48:17.019 --> 00:48:20.022 because that implies to me a Burkitt lymphoma. 00:48:20.689 --> 00:48:24.401 And, you know, possibly a double hit lymphoma, at least in older adults. 00:48:24.776 --> 00:48:27.696 And these require these have a, you know, poor prognosis 00:48:27.696 --> 00:48:30.741 when we try to require intensive chemotherapy, 00:48:31.700 --> 00:48:33.702 at least the double hits have a poor prognosis. 00:48:33.702 --> 00:48:34.912 Burkitt lymphoma, 00:48:34.912 --> 00:48:37.664 if recognized and treated properly, can have a good prognosis. 00:48:37.664 --> 00:48:40.626 But it's a very different treatment regimen. 00:48:41.543 --> 00:48:43.003 So this is what it looks like. 00:48:43.003 --> 00:48:44.630 You can see normal tonsil here 00:48:44.630 --> 00:48:48.258 with hyperplastic follicles that show, reactive germinal centers. 00:48:49.092 --> 00:48:51.720 And then this area we have these large nodules 00:48:51.720 --> 00:48:54.556 that are pushed together and it looks paler. 00:48:54.556 --> 00:48:58.018 And when we go closer sorry, this is not closer yet, 00:48:58.018 --> 00:49:01.063 but this is another area that shows a large diffuse area. 00:49:01.063 --> 00:49:02.397 So we've got this 00:49:02.397 --> 00:49:06.944 this area that has some abnormal large follicular appearing structure. 00:49:06.944 --> 00:49:08.654 And then a large diffuse area. 00:49:09.905 --> 00:49:10.781 And this is a high 00:49:10.781 --> 00:49:14.368 power showing the cell size again with a small lymphocyte for comparison. 00:49:14.701 --> 00:49:16.662 These are large cells. 00:49:16.662 --> 00:49:21.667 So we turn to our immunohistochemical stains. 00:49:21.667 --> 00:49:23.627 And we proved that there are B cells 00:49:23.627 --> 00:49:26.922 and not T cells with CD 20 and CD3 respectively. 00:49:27.714 --> 00:49:31.301 They express BCL six and they have a high proliferation index, 00:49:31.301 --> 00:49:35.097 which makes us think about the possibility of a diffuse large B-cell lymphoma. 00:49:36.014 --> 00:49:40.852 This is just a nice, picture of the, proliferation marker MeV one. 00:49:40.852 --> 00:49:45.232 You can see in those abnormal follicles that they have a high uniform, 00:49:46.274 --> 00:49:47.109 proliferation. 00:49:47.109 --> 00:49:49.444 So all the positive cells are in cycle. 00:49:49.444 --> 00:49:53.532 If you look at normal, germinal centers, you can see that they're polarized 00:49:53.532 --> 00:49:56.952 where they're higher on one end and lower on the other end, higher and lower. 00:49:56.952 --> 00:49:58.453 That's what reactive ones look like. 00:49:58.453 --> 00:50:00.872 So these are distinctly abnormal on the stain. 00:50:03.250 --> 00:50:05.210 And this is a 00:50:05.210 --> 00:50:07.963 immunohistochemical stain for IRF for 00:50:07.963 --> 00:50:10.966 or Mom one. 00:50:11.091 --> 00:50:13.635 Which is strongly positive in this case okay. 00:50:13.635 --> 00:50:15.929 So let's go back to our differential diagnosis. 00:50:15.929 --> 00:50:17.347 It's not Burkitt lymphoma. 00:50:17.347 --> 00:50:21.101 The morphology was wrong for that follicular lymphoma. 00:50:21.101 --> 00:50:21.727 Grade three. 00:50:21.727 --> 00:50:23.812 Well the area that had the large follicular 00:50:23.812 --> 00:50:27.566 type structures would be considered for follicular lymphoma. 00:50:27.566 --> 00:50:28.483 Grade three. 00:50:28.483 --> 00:50:31.028 And the area that would look diffuse looks like it. 00:50:31.028 --> 00:50:33.530 If you saw large B-cell lymphoma. 00:50:33.530 --> 00:50:35.782 But the other entity that is now entered 00:50:35.782 --> 00:50:39.578 the picture is a large B-cell lymphoma with an IRA for rearrangement. 00:50:39.578 --> 00:50:41.621 And this is new in 2017. 00:50:41.621 --> 00:50:44.750 So we have to turn to fish once again. 00:50:45.042 --> 00:50:45.834 Fish for MC, 00:50:45.834 --> 00:50:51.089 BCL two and BCL six are all negative, but fish for era four was positive. 00:50:52.340 --> 00:50:53.133 So that puts it 00:50:53.133 --> 00:50:57.012 into a category of a large B-cell lymphoma with IRF for rearrangement. 00:50:57.012 --> 00:50:59.389 These are usually localized in the head and neck. 00:50:59.389 --> 00:51:02.392 In younger patients, they can be at any age, but 00:51:02.726 --> 00:51:06.563 where we see them more in in teenagers or young adults 00:51:07.147 --> 00:51:10.358 or even young children, these can look like a diffuse large 00:51:10.358 --> 00:51:13.653 B-cell lymphoma, like a high grade follicular lymphoma, 00:51:13.987 --> 00:51:16.782 or like a pediatric type follicular lymphoma. 00:51:16.782 --> 00:51:19.826 They're positive for BCL six and IRF for. 00:51:20.410 --> 00:51:23.622 And the key here is that these have a really good outcome after chemotherapy. 00:51:23.622 --> 00:51:26.625 And they require less intensive therapy than, 00:51:27.042 --> 00:51:29.711 that you would use for Burkitt lymphoma. 00:51:29.711 --> 00:51:35.175 So it's really important in the right clinical context to perform the IRF form, 00:51:35.258 --> 00:51:39.262 one stain and or do the fish to, try 00:51:39.262 --> 00:51:42.390 and make this diagnosis because it's going to make a big difference to the patient. 00:51:44.893 --> 00:51:45.310 This is 00:51:45.310 --> 00:51:49.272 just a study that we published, a year ago showing that while these 00:51:49.272 --> 00:51:54.611 are quite uncommon, overall, less than 1% of all lymphomas, they're, 00:51:55.070 --> 00:51:58.073 not that uncommon in children. 00:51:58.406 --> 00:52:01.785 From looking at, patients that were treated on children's oncology 00:52:01.785 --> 00:52:03.161 group protocols. 00:52:03.161 --> 00:52:07.165 We found two of 32 cases, which is 6% overall. 00:52:07.415 --> 00:52:10.961 And these, numbers are in line with what's been published out 00:52:10.961 --> 00:52:15.507 of, other cohorts, including a large, the largest cohort from Germany. So, 00:52:17.092 --> 00:52:19.970 again, when you're looking at, at, young patients 00:52:19.970 --> 00:52:22.973 and kids, this is not all that uncommon. 00:52:23.765 --> 00:52:24.850 Okay. 00:52:24.850 --> 00:52:29.229 So wrapping up here, the genetically defined lymphomas are ALK 00:52:29.229 --> 00:52:32.482 positive CLL mantle cell lymphoma, 00:52:32.482 --> 00:52:35.485 which requires demonstration of the 1114. 00:52:35.527 --> 00:52:37.070 Again, we've talked about the high grade 00:52:37.070 --> 00:52:40.115 B-cell lymphoma with MEK and BCL two or BCL six. 00:52:40.448 --> 00:52:44.286 We can now add large B-cell lymphoma with IRF for rearrangements. 00:52:46.037 --> 00:52:47.664 So these are ones that 00:52:47.664 --> 00:52:51.418 really require this information to make this specific diagnosis. 00:52:51.418 --> 00:52:54.337 So I would consider them genetically defined lymphomas. 00:52:54.337 --> 00:52:57.924 You're not going to make the diagnosis without the genetic information. 00:52:58.258 --> 00:53:01.720 Now there are other lymphomas that have highly characteristic genetic changes 00:53:01.720 --> 00:53:02.721 that don't define them. 00:53:02.721 --> 00:53:06.308 So most Burkitt lymphomas have an IgG translocation. 00:53:06.308 --> 00:53:09.311 But we know there are some examples that don't. 00:53:09.311 --> 00:53:13.315 Follicular lymphomas 90% have a 1418 translocation. 00:53:13.315 --> 00:53:15.025 But that still leaves 10% that don't. 00:53:15.025 --> 00:53:19.029 So it's not a defining translocation in lymphoid plasma lymphoma and hairy 00:53:19.029 --> 00:53:19.696 cell leukemia. 00:53:19.696 --> 00:53:23.992 There are characteristic mutations that are present in the bulk of cases, 00:53:23.992 --> 00:53:28.121 but as of yet, we don't currently use those mutations to define them. 00:53:30.457 --> 00:53:30.916 Okay. 00:53:30.916 --> 00:53:33.877 So I believe this is my last slide. 00:53:34.002 --> 00:53:37.005 And really the takeaway here is that we 00:53:37.005 --> 00:53:41.718 are we now have some entities defined in our classification that we follow. 00:53:42.469 --> 00:53:47.057 By genetic features, whether these be mutations or translocations. 00:53:47.432 --> 00:53:48.308 The translocation 00:53:48.308 --> 00:53:52.229 the definitions have been around longer, but the mutation ones are really emerging. 00:53:52.687 --> 00:53:54.606 This is more common in AML. 00:53:54.606 --> 00:53:57.150 And I didn't talk at all about ALS. But there are. 00:53:57.150 --> 00:53:59.861 But it's similar in that, disease. 00:53:59.861 --> 00:54:02.864 But they're really starting to emerge in lymphoma as well. 00:54:03.031 --> 00:54:05.575 So when it's going to make a difference 00:54:05.575 --> 00:54:08.745 and when the prognostic information is important, consider 00:54:09.037 --> 00:54:12.958 doing the appropriate additional testing to get this precise classification.