editorial comment
Publishing data from failed cytogenetic assays– what can we learn? Peter Hokland1 and Finbarr Cotter2 1
Department of Haematology, Aarhus University Hospital, Aarhus, Denmark and 2Centre for Haematology-Oncology, Barts Cancer Institute, Barts and London School of Medicine and Dentistry, London, UK
In this issue we publish data on the survival of patients with acute myeloid leukaemia (AML), in whom the initial cytogenetic analysis had failed (Medeiros et al, 2013). Very puzzlingly, these patients had an adverse outcome, even after adjustment of demographic variables, which are well-known players in this disease. Needless to say, our expert reviewers had several comments to this manuscript. First of all, was this just a result of mass significance? Hardly. The statistics appear solid and, as mentioned, accepted confounding factors have been considered. So, that reason can be disposed of. One reviewer quite rightly pointed out that, in the ideal world, failed cytogenetics should be a thing of the past. This might be the case in some countries with highly optimized shipment of samples to expert laboratories, and we should probably all aim to aspire to such a scenario. A step in this direction could be to simply wait for a preliminary quality control of the first sample and then securing a second one if the first was deemed insufficient. However, that may not be possible for all patients, some of whom will require prompt cytoreduction. In any case, and quite sensibly, patients with “pre-analytical” errors related to shipment and processing (wrong anti-coagulant?) were excluded. Consequently, the reported impact on survival relates only to those cases in which the cytogenetic analysis was unsuccessful despite optimal initial handling. Likewise, an analysis was not considered as failed if a clonal aberration was observed, even if the magical number of 20 metaphases was not reached. Let us then have a closer look at the remainder of the material, where we must assume that a biological factor(s) results in the adverse course of disease: in fact, the data set, with its mindboggling conclusion, is a compilation of data from a number of protocols carried out in the “Southwestern Oncology Group (SWOG) area,” which – despite its
Correspondence: Professor P. Hokland, Department of Haematology, Aarhus University Hospital, Tage-Hansens Gade 2, 8000 Aarhus C, DENMARK. Email: [email protected]
ª 2013 John Wiley & Sons Ltd British Journal of Haematology, 2014, 164, 163–164
name-encompasses all of the US. Moreover, it covers protocols dating back to the mid-1980’s, i.e., more than 25 years ago. In light of this, it is quite understandable that archival material was not available from a sufficient number of patients to redo the analysis. On the other hand, data on the frequency of failed assays over time would have been informative to have. Thus, was failure to secure a karyotype mainly a thing of the past, or is it a phenomenon we have to accept, even today? Moreover, was the failure rate of the many different laboratories contributing data comparable, or did a few contribute a major part of the failures? In line with this (and technical issues are especially important in cytogenetics, where it is known that certain lot numbers of glass slides are less conducive for optimal banding than others), would data on cell concentrations upon arrival and a review of specific culture conditions as well as assessment of banding at the different centres have contributed information? Laboratory issues aside, the SWOG data also provide an opportunity to compare the approach to cytogenetics between large and highly respected groups and maybe even across the Atlantic. Thus, data from 2010 from the UK-AML trials would seem ideal, given that they are perhaps the largest and most rapidly accruing trials around. First of all, it is noticeable that similar rates of failure of cytogenetic analysis, as defined by inadequate banding and/or insufficient number of metaphases (around 5%), to the SWOG data were seen in adults (Grimwade et al, 2010) and in paediatric cases (Harrison et al, 2010). In other words, failed cytogenetics is certainly not just an American phenomenon. Once more, despite the attempts to exclude pre-analytical errors, the reasons for failure cannot be judged in detail, especially the contribution of delayed transit times (or indeed temperature changes during transit in the icy cold winters of Central US?) and the use of wrong anticoagulants. A basic difference between the SWOG and the UK-AML studies pertains to the presence or absence of a centralized review of the karyotype samples. This was done in the SWOG trials, but not in the UK ones. On the other hand, proficiency-testing programmes were in place for the UK trials, but not in the SWOG ones. It is impossible to evaluate the First published online 24 October 2013 doi:10.1111/bjh.12623
Editorial comment consequences of these different approaches across the Atlantic in the integration of cytogenetics in such high quality multicentre studies, but there is no doubt that they can be important. Thus, the UK studies failed to observe an adverse impact on treatment outcome of failed cytogenetics. Rather, in this setting, “no sample” patients had turned out to be the ones with adverse outcome at the 10-year mark (34% vs. 46% with failed v. 40% with successful), maybe because these patients were deemed to have such a poor performance status that a bone marrow aspiration was not performed. We are then left to contemplate what biologically distinct group of patients is identified in the SWOG study. Were the patients, e.g., the ones with lower numbers of harvested cells, reflecting a hypocellular marrow [a parallel to the adverse outcome of hypocellular MDS patients (Tong et al, 2012)]? Or were they the ones with too few metaphases not quite reaching the magical number of 20? On firmer ground, it can also be argued that the present observation could be considered to be a preferential failure in older patients. In fact, a trend (P = 008) was observed towards an adverse effect on age. If so, they would not be the patients with
References Becker, H., Marcucci, G., Maharry, K., Radmacher, M.D., Mrozek, K., Margeson, D., Whitman, S.P., Wu, Y.Z., Schwind, S., Paschka, P., Powell, B.L., Carter, T.H., Kolitz, J.E., Wetzler, M., Carroll, A.J., Baer, M.R., Caligiuri, M.A., Larson, R.A. & Bloomfield, C.D. (2010) Favorable prognostic impact of NPM1 mutations in older patients with cytogenetically normal de novo acute myeloid leukemia and associated geneand microRNA-expression signatures: a cancer and leukemia group B study. Journal of Clinical Oncology, 28, 596–604. Cancer Genome Atlas Research Network. (2013) Genomic and epigenomic landscapes of adult de
164
normal karyotypes, because they would include patients with NPM1 mutations, who in fact have a favourable outcome according to recent Cancer and Leukemia Group B data (Becker et al, 2010). While these deliberations on a surprising historic data set cannot then be boiled down to a simple explanation, it is clear what we should do in the future. Firstly, we still need the karyotypic assay, even in the age of genomic analyses. Secondly, we need to work to get the percentage of failed assays down. Thirdly, as also pointed out in the SWOG and UK-AML publications cited, molecular analysis is to be integrated in the diagnostic workup of these patients (Cancer Genome Atlas Research Network, 2013). Fourthly, biobanking at diagnosis for future analyses should be made an integral part of the multidisciplinary diagnostic process in these patients. Finally, should we regard a failure to achieve a successful karyotype as an adverse prognostic omen in the future? We feel that the jury is still out on that, but such a scenario should serve as a warning that the patient needs closer scrutiny during the diagnostic phase.
novo acute myeloid leukemia. New England Journal of Medicine, 368, 2059–2074. Grimwade, D., Hills, R.K., Moorman, A.V., Walker, H., Chatters, S., Goldstone, A.H., Wheatley, K., Harrison, C.J. & Burnett, A.K. (2010) Refinement of cytogenetic classification in acute myeloid leukemia: determination of prognostic significance of rare recurring chromosomal abnormalities among 5876 younger adult patients treated in the United Kingdom medical research council trials. Blood, 116, 354–365. Harrison, C.J., Hills, R.K., Moorman, A.V., Grimwade, D.J., Hann, I., Webb, D.K., Wheatley, K., de Graaf, S.S., van den Berg, E., Burnett, A.K. & Gibson, B.E. (2010) Cytogenetics of childhood acute myeloid leukemia: United
Kingdom medical research council treatment trials AML 10 and 12. Journal of Clinical Oncology, 28, 2674–2681. Medeiros, B., Othus, O., Estey, E.H., Fang, M. & Appelbaum, F.R. (2013) Unsuccesful diagnostic cytogenetic analysis is a poor prognostic feature in acute myeloid leukaemia. British Journal of Haematology, 164, 245–250. Tong, W.G., Quintas-Cardama, A., Kadia, T., Borthakur, G., Jabbour, E., Ravandi, F., Faderl, S., Wierda, W., Pierce, S., Shan, J., BuesoRamos, C., Kantarjian, H. & Garcia-Manero, G. (2012) Predicting survival of patients with hypocellular myelodysplastic syndrome: development of a disease-specific prognostic score system. Cancer, 118, 4462–4470.
ª 2013 John Wiley & Sons Ltd British Journal of Haematology, 2014, 164, 163–164
Publishing data from failed cytogenetic assays– what can we learn? Peter Hokland1 and Finbarr Cotter2 1
Department of Haematology, Aarhus University Hospital, Aarhus, Denmark and 2Centre for Haematology-Oncology, Barts Cancer Institute, Barts and London School of Medicine and Dentistry, London, UK
In this issue we publish data on the survival of patients with acute myeloid leukaemia (AML), in whom the initial cytogenetic analysis had failed (Medeiros et al, 2013). Very puzzlingly, these patients had an adverse outcome, even after adjustment of demographic variables, which are well-known players in this disease. Needless to say, our expert reviewers had several comments to this manuscript. First of all, was this just a result of mass significance? Hardly. The statistics appear solid and, as mentioned, accepted confounding factors have been considered. So, that reason can be disposed of. One reviewer quite rightly pointed out that, in the ideal world, failed cytogenetics should be a thing of the past. This might be the case in some countries with highly optimized shipment of samples to expert laboratories, and we should probably all aim to aspire to such a scenario. A step in this direction could be to simply wait for a preliminary quality control of the first sample and then securing a second one if the first was deemed insufficient. However, that may not be possible for all patients, some of whom will require prompt cytoreduction. In any case, and quite sensibly, patients with “pre-analytical” errors related to shipment and processing (wrong anti-coagulant?) were excluded. Consequently, the reported impact on survival relates only to those cases in which the cytogenetic analysis was unsuccessful despite optimal initial handling. Likewise, an analysis was not considered as failed if a clonal aberration was observed, even if the magical number of 20 metaphases was not reached. Let us then have a closer look at the remainder of the material, where we must assume that a biological factor(s) results in the adverse course of disease: in fact, the data set, with its mindboggling conclusion, is a compilation of data from a number of protocols carried out in the “Southwestern Oncology Group (SWOG) area,” which – despite its
Correspondence: Professor P. Hokland, Department of Haematology, Aarhus University Hospital, Tage-Hansens Gade 2, 8000 Aarhus C, DENMARK. Email: [email protected]
ª 2013 John Wiley & Sons Ltd British Journal of Haematology, 2014, 164, 163–164
name-encompasses all of the US. Moreover, it covers protocols dating back to the mid-1980’s, i.e., more than 25 years ago. In light of this, it is quite understandable that archival material was not available from a sufficient number of patients to redo the analysis. On the other hand, data on the frequency of failed assays over time would have been informative to have. Thus, was failure to secure a karyotype mainly a thing of the past, or is it a phenomenon we have to accept, even today? Moreover, was the failure rate of the many different laboratories contributing data comparable, or did a few contribute a major part of the failures? In line with this (and technical issues are especially important in cytogenetics, where it is known that certain lot numbers of glass slides are less conducive for optimal banding than others), would data on cell concentrations upon arrival and a review of specific culture conditions as well as assessment of banding at the different centres have contributed information? Laboratory issues aside, the SWOG data also provide an opportunity to compare the approach to cytogenetics between large and highly respected groups and maybe even across the Atlantic. Thus, data from 2010 from the UK-AML trials would seem ideal, given that they are perhaps the largest and most rapidly accruing trials around. First of all, it is noticeable that similar rates of failure of cytogenetic analysis, as defined by inadequate banding and/or insufficient number of metaphases (around 5%), to the SWOG data were seen in adults (Grimwade et al, 2010) and in paediatric cases (Harrison et al, 2010). In other words, failed cytogenetics is certainly not just an American phenomenon. Once more, despite the attempts to exclude pre-analytical errors, the reasons for failure cannot be judged in detail, especially the contribution of delayed transit times (or indeed temperature changes during transit in the icy cold winters of Central US?) and the use of wrong anticoagulants. A basic difference between the SWOG and the UK-AML studies pertains to the presence or absence of a centralized review of the karyotype samples. This was done in the SWOG trials, but not in the UK ones. On the other hand, proficiency-testing programmes were in place for the UK trials, but not in the SWOG ones. It is impossible to evaluate the First published online 24 October 2013 doi:10.1111/bjh.12623
Editorial comment consequences of these different approaches across the Atlantic in the integration of cytogenetics in such high quality multicentre studies, but there is no doubt that they can be important. Thus, the UK studies failed to observe an adverse impact on treatment outcome of failed cytogenetics. Rather, in this setting, “no sample” patients had turned out to be the ones with adverse outcome at the 10-year mark (34% vs. 46% with failed v. 40% with successful), maybe because these patients were deemed to have such a poor performance status that a bone marrow aspiration was not performed. We are then left to contemplate what biologically distinct group of patients is identified in the SWOG study. Were the patients, e.g., the ones with lower numbers of harvested cells, reflecting a hypocellular marrow [a parallel to the adverse outcome of hypocellular MDS patients (Tong et al, 2012)]? Or were they the ones with too few metaphases not quite reaching the magical number of 20? On firmer ground, it can also be argued that the present observation could be considered to be a preferential failure in older patients. In fact, a trend (P = 008) was observed towards an adverse effect on age. If so, they would not be the patients with
References Becker, H., Marcucci, G., Maharry, K., Radmacher, M.D., Mrozek, K., Margeson, D., Whitman, S.P., Wu, Y.Z., Schwind, S., Paschka, P., Powell, B.L., Carter, T.H., Kolitz, J.E., Wetzler, M., Carroll, A.J., Baer, M.R., Caligiuri, M.A., Larson, R.A. & Bloomfield, C.D. (2010) Favorable prognostic impact of NPM1 mutations in older patients with cytogenetically normal de novo acute myeloid leukemia and associated geneand microRNA-expression signatures: a cancer and leukemia group B study. Journal of Clinical Oncology, 28, 596–604. Cancer Genome Atlas Research Network. (2013) Genomic and epigenomic landscapes of adult de
164
normal karyotypes, because they would include patients with NPM1 mutations, who in fact have a favourable outcome according to recent Cancer and Leukemia Group B data (Becker et al, 2010). While these deliberations on a surprising historic data set cannot then be boiled down to a simple explanation, it is clear what we should do in the future. Firstly, we still need the karyotypic assay, even in the age of genomic analyses. Secondly, we need to work to get the percentage of failed assays down. Thirdly, as also pointed out in the SWOG and UK-AML publications cited, molecular analysis is to be integrated in the diagnostic workup of these patients (Cancer Genome Atlas Research Network, 2013). Fourthly, biobanking at diagnosis for future analyses should be made an integral part of the multidisciplinary diagnostic process in these patients. Finally, should we regard a failure to achieve a successful karyotype as an adverse prognostic omen in the future? We feel that the jury is still out on that, but such a scenario should serve as a warning that the patient needs closer scrutiny during the diagnostic phase.
novo acute myeloid leukemia. New England Journal of Medicine, 368, 2059–2074. Grimwade, D., Hills, R.K., Moorman, A.V., Walker, H., Chatters, S., Goldstone, A.H., Wheatley, K., Harrison, C.J. & Burnett, A.K. (2010) Refinement of cytogenetic classification in acute myeloid leukemia: determination of prognostic significance of rare recurring chromosomal abnormalities among 5876 younger adult patients treated in the United Kingdom medical research council trials. Blood, 116, 354–365. Harrison, C.J., Hills, R.K., Moorman, A.V., Grimwade, D.J., Hann, I., Webb, D.K., Wheatley, K., de Graaf, S.S., van den Berg, E., Burnett, A.K. & Gibson, B.E. (2010) Cytogenetics of childhood acute myeloid leukemia: United
Kingdom medical research council treatment trials AML 10 and 12. Journal of Clinical Oncology, 28, 2674–2681. Medeiros, B., Othus, O., Estey, E.H., Fang, M. & Appelbaum, F.R. (2013) Unsuccesful diagnostic cytogenetic analysis is a poor prognostic feature in acute myeloid leukaemia. British Journal of Haematology, 164, 245–250. Tong, W.G., Quintas-Cardama, A., Kadia, T., Borthakur, G., Jabbour, E., Ravandi, F., Faderl, S., Wierda, W., Pierce, S., Shan, J., BuesoRamos, C., Kantarjian, H. & Garcia-Manero, G. (2012) Predicting survival of patients with hypocellular myelodysplastic syndrome: development of a disease-specific prognostic score system. Cancer, 118, 4462–4470.
ª 2013 John Wiley & Sons Ltd British Journal of Haematology, 2014, 164, 163–164
