Just Accepted by Leukemia & Lymphoma
Molecular Pathology of Myelodysplastic Syndromes: New Developments and Implication in Diagnosis and Treatment Xiaohui Zhang, Jeffrey E. Lancet, Ling Zhang DOI: 10.3109/10428194.2015.1037756
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
Abstract Myelodysplastic syndromes (MDS) diagnosis has been based on clinical presentations, laboratory and morphological findings, and molecular and cytogenetic profiles. With the advent of single nucleotide polymorphism (SNP) microarrays and high throughput sequencing technologies, a tremendous amount of progress has been made toward better understanding of MDS genetic and molecular changes. Recurring genetic abnormalities have been revealed in up to 80-90% of MDS patients. We herein review clinical and pathological basics of MDS, the most up-to-date advances in molecular diagnostics of MDS, including current understanding on cytogenetic and molecular markers of MDS, and their implications in MDS diagnosis and therapy selection
© 2015 Informa UK, Ltd. This provisional PDF corresponds to the article as it appeared upon acceptance. Fully formatted PDF and full text (HTML) versions will be made available soon. DISCLAIMER: The ideas and opinions expressed in the journal’s Just Accepted articles do not necessarily reflect those of Informa Healthcare (the Publisher), the Editors or the journal. The Publisher does not assume any responsibility for any injury and/or damage to persons or property arising from or related to any use of the material contained in these articles. The reader is advised to check the appropriate medical literature and the product information currently provided by the manufacturer of each drug to be administered to verify the dosages, the method and duration of administration, and contraindications. It is the responsibility of the treating physician or other health care professional, relying on his or her independent experience and knowledge of the patient, to determine drug dosages and the best treatment for the patient. Just Accepted articles have undergone full scientific review but none of the additional editorial preparation, such as copyediting, typesetting, and proofreading, as have articles published in the traditional manner. There may, therefore, be errors in Just Accepted articles that will be corrected in the final print and final online version of the article. Any use of the Just Accepted articles is subject to the express understanding that the papers have not yet gone through the full quality control process prior to publication.
Molecular Pathology of Myelodysplastic Syndromes: New Developments and Implication in Diagnosis and Treatment
Xiaohui Zhang1, Jeffrey E. Lancet2, Ling Zhang1 1
Department of Hematopathology and Laboratory Medicine, 2Department of Malignant
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
Hematology, H. Lee Moffitt Cancer Center and Research Institute, Tampa, FL 33612
Corresponding author: Ling Zhang, Department of Hematopathology and Laboratory Medicine, H. Lee Moffitt Cancer Center, MCC-LAB, 12902 USF Magnolia Dr., Tampa, Florida 33612. Tel: (813)745-8198. FAX: (813)745-3570. Email: [email protected]
Short title: Advances in Myelodysplastic Syndromes Abstract Myelodysplastic syndromes (MDS) diagnosis has been based on clinical presentations, laboratory and morphological findings, and molecular and cytogenetic profiles. With the advent of single nucleotide polymorphism (SNP) microarrays and high throughput sequencing technologies, a tremendous amount of progress has been made toward better understanding of MDS genetic and molecular changes. Recurring genetic abnormalities have been revealed in up to 80-90% of MDS patients. We herein review clinical and pathological basics of MDS, the most up-to-date advances in molecular diagnostics of MDS, including current understanding on cytogenetic and molecular markers of MDS, and their implications in MDS diagnosis and therapy selection
Keywords: Myelodysplastic syndromes, molecular diagnostics, cytogenetics, therapy
1
Introduction Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic stem cell diseases characterized by peripheral cytopenias, ineffective hematopoiesis, and increased risk of transformation to acute myeloid leukemia (AML). They occur principally in older adults (median age of 70 years) and at an annual incidence of 3-5/100,000 persons in general population, although with a much higher incidence (up to 75 per 100,000 persons) in those who are 65 years or older.1 The majority of patients present with symptoms related to cytopenia(s), and most patients present with anemia and transfusion dependence. Less frequent initial manifestations
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
include neutropenia and/or thrombocytopenia. Classification by WHO in 2008 defined 6 adult and 1 childhood MDS categories based on morphology, cytopenias, blast count, and cytogenetics. The diseases include refractory cytopenias with unilineage dysplasia (RCUD) including refractory anemia (RA), refractory neutropenia (RN) and refractory thrombocytopenia (RT), refractory anemia with ring sideroblasts (RARS), refractory cytopenia with multilineage dysplasia (RCMD), refractory anemia with excess blasts (RAEB, type I and II), myelodysplastic syndrome - unclassifiable (MDS-U), and myelodysplastic syndrome with isolated del(5q) (Table 1). Clinically, MDS can also be divided into de novo and secondary or therapy related. Persistent cytopenia without dysplasia and without one of the specific cytogenetic abnormalities should be viewed as “idiopathic cytopenia of undetermined significance” (ICUS). In addition to morphological assessment of the bone marrow, conventional cytogenetics plays a major role in the evaluation of patients with MDS in regard to determination of clonality. Specific recurring chromosomal changes have been recognized by WHO 2008 classification and can be used in monitoring of the disease during follow up (Table 2). However, the current WHO classification does not recognize the presence of mutations in facilitating a diagnosis of MDS. The only cytogenetic finding associated with a specific WHO MDS classification is isolated del(5q). Several other assays are used to help in the diagnosis of MDS. These include the use of fluorescent in situ hybridization (FISH) and flow cytometric analysis of the marrow cells. Because of the cytogenetic heterogeneity in MDS, many clinicians believe that FISH probes do
2
not add much value to the routine metaphase cytogenetic analysis.2,3 Although it is not a standard of care for diagnosis in the USA, studies of the International/European LeukemiaNet Working Group proposed that flow cytometry can help to identify abnormal phenotypic patterns that may support an MDS diagnosis in the setting of minimal dysplasia.4 Meanwhile, our knowledge about the molecular pathogenesis of MDS has expanded during the past years. The advent of high-throughput sequencing technologies has begun to transform our understanding of the molecular paradigms of this disease. It is expected that new molecular insights will ultimately optimize clinical care.
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
A. Genetic Methodologies Common genetic alterations in MDS occur at different levels, including chromosome, DNA, epigenetics, and RNA levels. Currently, several genetic methodologies are being employed to detect cytogenetic and molecular aberrations. Conventional cytogenetics study is central to MDS diagnosis, subclassification, and prognostication, recommended in initial MDS workup and subsequent monitoring. FISH uses sequence-specific probes to determine chromosomal gains, losses, and translocations in the targeted regions. Interphase cells, and even formalin-fixed, paraffin-embedded tissue, can be used for analysis. Additionally, small and cryptic cytogenetic abnormalities may be detected. Studies demonstrated that FISH detected approximately 70% of the cytogenetic abnormalities detected by conventional karyotyping. As such, it is recommended that the primary use of FISH should be in the setting of unsuccessful conventional cytogenetics such as inadequate metaphases obtained.5 Single-nucleotide polymorphism array (SNP) is a genome-wide cytogenetic analysis, hybridizing tumor DNA to microarray probes specific for allelic variants of a single-nucleotide polymorphism. This has the ability to capture additional cryptic gains and losses and to detect copy-number neutral loss of heterozygosity (CN-LOH) or uniparental disomy (UPD) at high resolution, which are undetectable by conventional cytogenetics. The combination of SNP and metaphase conventional cytogenetics can reveal chromosomal alterations in 75% of patients, compared with 50% of patients by conventional cytogenetics alone,6 thereby emerging as a very useful complementary tool for karyotyping. Limitations of SNP include the inability to detect balanced chromosomal translocations. Next-generation sequencing (NGS) is a powerful
3
sequencing approach that is fundamentally different from traditional sequencing methods such as Sanger sequencing. For the purpose of MDS diagnostics, targeted amplicon deep-sequencing has been shown to be more suitable than genome-wide sequencing. NGS is sensitive enough to detect low-frequency variants, as low as 1-2%.7 With this technology, our knowledge about molecular pathogenesis has been significantly expanded. Well-defined molecular pathways, as well as newly identified pathways, have improved our understanding of MDS, hopefully translating into improved clinical care of MDS patients in the future. B. Importance of Cytogenetics in MDS
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
Common recurring chromosomal changes are listed in Table 2. The most common ones are deletion 5q (approximately 15%), del(7q)/monosomy 7 (approximately 10%), and trisomy 8 (approximately 10%). Complex karyotypes are found in 10% to 20% of de novo MDS. They typically include chromosomes 5 and/or 7 and are associated with an unfavorable clinical course.8 Overall, approximately 50% of de novo MDS and 80% of secondary MDS cases show detectable cytogenetic aberrations8. It is known that certain cytogenetic changes are associated with distinct clinical or morphologic features. Chromosome 17p loss is associated with pseudo Pelger-Huët anomaly, small vacuolated neutrophils, TP53 mutation, and an unfavorable clinical course.9 Inversion of 3q21q26 or t(3;3)(q21;q26.2) is often associated with atypical megakaryocytic proliferation, increased blasts, and rapid evolution to AML.10 Cases with inv(3)/t(3;3) or t(6;9) should be closely monitored for disease progression to AML. MDS with isolated del(5q) is the only cytogenetically defined MDS subtype currently recognized by the WHO classification. Morphologically, it is characterized by single or hypolobated megakaryocytes with
Molecular Pathology of Myelodysplastic Syndromes: New Developments and Implication in Diagnosis and Treatment Xiaohui Zhang, Jeffrey E. Lancet, Ling Zhang DOI: 10.3109/10428194.2015.1037756
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
Abstract Myelodysplastic syndromes (MDS) diagnosis has been based on clinical presentations, laboratory and morphological findings, and molecular and cytogenetic profiles. With the advent of single nucleotide polymorphism (SNP) microarrays and high throughput sequencing technologies, a tremendous amount of progress has been made toward better understanding of MDS genetic and molecular changes. Recurring genetic abnormalities have been revealed in up to 80-90% of MDS patients. We herein review clinical and pathological basics of MDS, the most up-to-date advances in molecular diagnostics of MDS, including current understanding on cytogenetic and molecular markers of MDS, and their implications in MDS diagnosis and therapy selection
© 2015 Informa UK, Ltd. This provisional PDF corresponds to the article as it appeared upon acceptance. Fully formatted PDF and full text (HTML) versions will be made available soon. DISCLAIMER: The ideas and opinions expressed in the journal’s Just Accepted articles do not necessarily reflect those of Informa Healthcare (the Publisher), the Editors or the journal. The Publisher does not assume any responsibility for any injury and/or damage to persons or property arising from or related to any use of the material contained in these articles. The reader is advised to check the appropriate medical literature and the product information currently provided by the manufacturer of each drug to be administered to verify the dosages, the method and duration of administration, and contraindications. It is the responsibility of the treating physician or other health care professional, relying on his or her independent experience and knowledge of the patient, to determine drug dosages and the best treatment for the patient. Just Accepted articles have undergone full scientific review but none of the additional editorial preparation, such as copyediting, typesetting, and proofreading, as have articles published in the traditional manner. There may, therefore, be errors in Just Accepted articles that will be corrected in the final print and final online version of the article. Any use of the Just Accepted articles is subject to the express understanding that the papers have not yet gone through the full quality control process prior to publication.
Molecular Pathology of Myelodysplastic Syndromes: New Developments and Implication in Diagnosis and Treatment
Xiaohui Zhang1, Jeffrey E. Lancet2, Ling Zhang1 1
Department of Hematopathology and Laboratory Medicine, 2Department of Malignant
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
Hematology, H. Lee Moffitt Cancer Center and Research Institute, Tampa, FL 33612
Corresponding author: Ling Zhang, Department of Hematopathology and Laboratory Medicine, H. Lee Moffitt Cancer Center, MCC-LAB, 12902 USF Magnolia Dr., Tampa, Florida 33612. Tel: (813)745-8198. FAX: (813)745-3570. Email: [email protected]
Short title: Advances in Myelodysplastic Syndromes Abstract Myelodysplastic syndromes (MDS) diagnosis has been based on clinical presentations, laboratory and morphological findings, and molecular and cytogenetic profiles. With the advent of single nucleotide polymorphism (SNP) microarrays and high throughput sequencing technologies, a tremendous amount of progress has been made toward better understanding of MDS genetic and molecular changes. Recurring genetic abnormalities have been revealed in up to 80-90% of MDS patients. We herein review clinical and pathological basics of MDS, the most up-to-date advances in molecular diagnostics of MDS, including current understanding on cytogenetic and molecular markers of MDS, and their implications in MDS diagnosis and therapy selection
Keywords: Myelodysplastic syndromes, molecular diagnostics, cytogenetics, therapy
1
Introduction Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic stem cell diseases characterized by peripheral cytopenias, ineffective hematopoiesis, and increased risk of transformation to acute myeloid leukemia (AML). They occur principally in older adults (median age of 70 years) and at an annual incidence of 3-5/100,000 persons in general population, although with a much higher incidence (up to 75 per 100,000 persons) in those who are 65 years or older.1 The majority of patients present with symptoms related to cytopenia(s), and most patients present with anemia and transfusion dependence. Less frequent initial manifestations
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
include neutropenia and/or thrombocytopenia. Classification by WHO in 2008 defined 6 adult and 1 childhood MDS categories based on morphology, cytopenias, blast count, and cytogenetics. The diseases include refractory cytopenias with unilineage dysplasia (RCUD) including refractory anemia (RA), refractory neutropenia (RN) and refractory thrombocytopenia (RT), refractory anemia with ring sideroblasts (RARS), refractory cytopenia with multilineage dysplasia (RCMD), refractory anemia with excess blasts (RAEB, type I and II), myelodysplastic syndrome - unclassifiable (MDS-U), and myelodysplastic syndrome with isolated del(5q) (Table 1). Clinically, MDS can also be divided into de novo and secondary or therapy related. Persistent cytopenia without dysplasia and without one of the specific cytogenetic abnormalities should be viewed as “idiopathic cytopenia of undetermined significance” (ICUS). In addition to morphological assessment of the bone marrow, conventional cytogenetics plays a major role in the evaluation of patients with MDS in regard to determination of clonality. Specific recurring chromosomal changes have been recognized by WHO 2008 classification and can be used in monitoring of the disease during follow up (Table 2). However, the current WHO classification does not recognize the presence of mutations in facilitating a diagnosis of MDS. The only cytogenetic finding associated with a specific WHO MDS classification is isolated del(5q). Several other assays are used to help in the diagnosis of MDS. These include the use of fluorescent in situ hybridization (FISH) and flow cytometric analysis of the marrow cells. Because of the cytogenetic heterogeneity in MDS, many clinicians believe that FISH probes do
2
not add much value to the routine metaphase cytogenetic analysis.2,3 Although it is not a standard of care for diagnosis in the USA, studies of the International/European LeukemiaNet Working Group proposed that flow cytometry can help to identify abnormal phenotypic patterns that may support an MDS diagnosis in the setting of minimal dysplasia.4 Meanwhile, our knowledge about the molecular pathogenesis of MDS has expanded during the past years. The advent of high-throughput sequencing technologies has begun to transform our understanding of the molecular paradigms of this disease. It is expected that new molecular insights will ultimately optimize clinical care.
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
A. Genetic Methodologies Common genetic alterations in MDS occur at different levels, including chromosome, DNA, epigenetics, and RNA levels. Currently, several genetic methodologies are being employed to detect cytogenetic and molecular aberrations. Conventional cytogenetics study is central to MDS diagnosis, subclassification, and prognostication, recommended in initial MDS workup and subsequent monitoring. FISH uses sequence-specific probes to determine chromosomal gains, losses, and translocations in the targeted regions. Interphase cells, and even formalin-fixed, paraffin-embedded tissue, can be used for analysis. Additionally, small and cryptic cytogenetic abnormalities may be detected. Studies demonstrated that FISH detected approximately 70% of the cytogenetic abnormalities detected by conventional karyotyping. As such, it is recommended that the primary use of FISH should be in the setting of unsuccessful conventional cytogenetics such as inadequate metaphases obtained.5 Single-nucleotide polymorphism array (SNP) is a genome-wide cytogenetic analysis, hybridizing tumor DNA to microarray probes specific for allelic variants of a single-nucleotide polymorphism. This has the ability to capture additional cryptic gains and losses and to detect copy-number neutral loss of heterozygosity (CN-LOH) or uniparental disomy (UPD) at high resolution, which are undetectable by conventional cytogenetics. The combination of SNP and metaphase conventional cytogenetics can reveal chromosomal alterations in 75% of patients, compared with 50% of patients by conventional cytogenetics alone,6 thereby emerging as a very useful complementary tool for karyotyping. Limitations of SNP include the inability to detect balanced chromosomal translocations. Next-generation sequencing (NGS) is a powerful
3
sequencing approach that is fundamentally different from traditional sequencing methods such as Sanger sequencing. For the purpose of MDS diagnostics, targeted amplicon deep-sequencing has been shown to be more suitable than genome-wide sequencing. NGS is sensitive enough to detect low-frequency variants, as low as 1-2%.7 With this technology, our knowledge about molecular pathogenesis has been significantly expanded. Well-defined molecular pathways, as well as newly identified pathways, have improved our understanding of MDS, hopefully translating into improved clinical care of MDS patients in the future. B. Importance of Cytogenetics in MDS
Leuk Lymphoma Downloaded from informahealthcare.com by Gazi Univ. on 05/05/15 For personal use only.
Common recurring chromosomal changes are listed in Table 2. The most common ones are deletion 5q (approximately 15%), del(7q)/monosomy 7 (approximately 10%), and trisomy 8 (approximately 10%). Complex karyotypes are found in 10% to 20% of de novo MDS. They typically include chromosomes 5 and/or 7 and are associated with an unfavorable clinical course.8 Overall, approximately 50% of de novo MDS and 80% of secondary MDS cases show detectable cytogenetic aberrations8. It is known that certain cytogenetic changes are associated with distinct clinical or morphologic features. Chromosome 17p loss is associated with pseudo Pelger-Huët anomaly, small vacuolated neutrophils, TP53 mutation, and an unfavorable clinical course.9 Inversion of 3q21q26 or t(3;3)(q21;q26.2) is often associated with atypical megakaryocytic proliferation, increased blasts, and rapid evolution to AML.10 Cases with inv(3)/t(3;3) or t(6;9) should be closely monitored for disease progression to AML. MDS with isolated del(5q) is the only cytogenetically defined MDS subtype currently recognized by the WHO classification. Morphologically, it is characterized by single or hypolobated megakaryocytes with
