Genetic susceptibility to leukemia
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WHO 2016 classification of Leukemia includes a new category called AML with germline mutations. This presentation is an overview to this entity.
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Genetic susceptibility to leukemia
- 1. Genetic Susceptibility to leukemia Presenter: Dr Ankit Jitani 1
- 2. Learning objectives • Familial cancer history • Cell cycle, check points and Tumor suppressor gene “double hit” concept • Familial cancer genetics • WHO 2016 genetic predisposition classification • Components genes in the classification • Approach to diagnosis and management differences 2
- 4. 4
- 6. Knudson’s Hypothesis Robbins & Cotrans Pathological Basis of disease, 8th Edition Dr Alfred Knudson 6
- 7. Cell cycle & check points Robbins & Cotrans Pathological Basis of disease, 8th Edition 7
- 8. RB gene Robbins & Cotrans Pathological Basis of disease, 8th Edition 8
- 9. TP53 Robbins & Cotrans Pathological Basis of disease, 8th Edition 9
- 10. Leukemogenesis 10 Molecular Pathogenesis and Therapy of Acute Myeloid Leukemia Ross L. Levine M.D. Human Oncology and Pathogenesis Program Leukemia Service, Department of Medicine Memorial Sloan Kettering Cancer Center
- 12. Loss of function mutation • Haploinsufficiency ▫ Loss of gene function caused by damage to a single allele • Dominant negative mutation ▫ Inactive protein from the mutant allele reduces the function of the normal allele 12
- 13. WHO Classification of MN with Germline Predisposition • Myeloid neoplasms with germline predisposition without a pre-existing disorder or organ dysfunction ▫ AML with germline CEBPA mutation ▫ MN with germline DDX41 mutation* • Myeloid neoplasms with germline predisposition and pre- existing platelet disorders ▫ MN with germline RUNX1 mutation* ▫ MN with germline ANKRD26 mutation* ▫ MN with germline ETV6 mutation* • Myeloid neoplasms with germline predisposition and other organ dysfunction ▫ MN with germline GATA2 mutation ▫ MN associated with BM bone marrow failure syndromes ▫ MN associated with telomere biology disorders ▫ JMML associated with neurofibromatosis, Noonan syndrome or Noonan syndrome-like disorders ▫ MN associated with Down syndrome* Arber DA et al, Blood 2016; doi:10.1182/blood-2016-03-643544 13
- 14. Blood 2016; 127 (8):960 14
- 15. 15
- 16. CCAAT/enhancer binding protein α • Transcription factor involved in lineage-specific myeloid differentiation • Encoded on chromosome 19q13.1 • Somatic mutations- 10-15% of CN-AML • 7-11% cases of CEBPA mutation may be familial 16
- 17. CCAAT/enhancer binding protein α 17
- 18. • 10 CEBPA-mutated families, representing 24 members • All cases presented de novo • Without preceding dysplastic or cytopenic phase • All cases CEBPAdm were detected • M1 or M2 morphology • Aberrant expression of CD7 18
- 19. OS in all cases OS in< 45 yrs age Survival after relapse for all patients with relapse 19
- 20. • CR1 rate of familial AML was 91% • Cumulative incidence of relapse at 10 years : ▫ Familial CEBPA: 56% ▫ Sporadic CEBPAsm: 62% ▫ Sporadic CEBPAdm: 44% • Median time to relapse: ▫ Familial CEBPA: 27 months ▫ Sporadic CEBPAsm: 11.8 months ▫ Sporadic CEBPAdm: 11.2 months 20
- 21. • Median survival after relapse: ▫ 8 years (familial) vs 16 months (CEBPAdm) vs 3.5 months (CEBPAsm) • Recurrence of familial AML retain sensitivity to chemotherapy • Allogeneic SCT to prevent future episodes • Challenge: identifying & excluding asymptomatic carrier as donor Tawana k et al Blood 2015 :blood-2015-05-647172 21
- 22. DDX41 (DEAD-box RNA helicase) • Class of tumor suppressor genes • Translation initiation & mRNA splicing • Role in innate immunity: cytoplasmic DNA sensor in dendritic cells • Germline frameshift mutation, second somatic hit • Deletion of 5q35.3– haploinsufficiency • Late-onset MDS/AML 22
- 23. • 7 families, 111 members • 27 cases of AML/MDS • Germline mutation: 19 cases • Monocytosis, CD14 + • Normal karyotype (70% versus 47% p = 0.0045) • Associated with TP53, RUNX1 mutation Polprasert C et al Cancer Cell 2015; 27, 658–670 23
- 24. • 289 families screened • 9 families with germline DDX41 mutation • Mean age of onset of MDS or AML: 62 yrs • Normal peripheral blood counts until late • Leukopenia & macrocytosis • Erythroid dysplasia Blood. 2016;127(8):1017-1023 24
- 25. DDX41 Out of frame Polprasert C et al Cancer Cell 2015; 27, 658–670 25 Same or different allele
- 26. DDX41 expression DDX41: response to lenalidomide Polprasert C et al Cancer Cell 2015; 27, 658–670 DDX41 mutants DDX41 mRNA expression 26
- 27. DDX41 mutation: Poor prognosis Polprasert C et al Cancer Cell 2015; 27, 658–670 27
- 28. GATA2 • GATA family of transcription factors • Essential for hematopoietic stem and progenitor cells, vascular & neural development • Erythroid cells, mast cells, and megakaryocytes • Non-hematopoietic embryonic stem cells • Forced expression of GATA2 promotes proliferation at the expense of differentiation 28
- 29. GATA2 • 18 patients susceptibility to: ▫ disseminated non-tuberculous mycobacterial infections, ▫ viral infections- HPV ▫ fungal infections ▫ PAP • Monocytopenia and lymphocytopenia • 9 developed AML/MDS 29
- 30. GATA2 • DNA samples from 13 of the 16 kindreds • 10 showed GATA2 mutation • Infections and pulmonary features predate leukemia • MonoMAC: ▫ Younger age of onset of MDS (33 vs 60-70 years) ▫ BM features including hypocellularity, significant fibrosis, and multilineage dysplasia 30
- 31. • Emberger syndrome: ▫ Lymphedema of lower extremities and genitals ▫ Myelodysplasia progressing to AML ▫ Cutaneous warts ▫ Sensorineural deafness • DCML deficiency ▫ dendritic cell, monocyte, B & NK lymphoid cell deficiency ▫ Infections & MDS/AML GATA2 Wang X el at Haematologica. 2015 ; 100(10): e398–e401 31
- 32. GATA2 Spinner MA et al Blood 2014 123:809-821 32
- 33. GATA2 outcome: conflicting data Poorer response to treatment Lower CR rate Resistant disease 33 Vigilant bone marrow monitoring
- 34. 34
- 35. 51 cause FDP, 3 predisposes to leukemia Blood 2016 127:2814-2823 35
- 36. Germline predisposition in FPD Predisposition Inheritence Genes Altered pathway Hemat malignancy Familial platelet disorder asso with myeloid malignancy AD RUNX1 TF network MDS, AML, T-ALL Thrombocytopenia 2 AD ANKRD26 MAPK signaling MDS, AML, CML Thrombocytopenia 5 AD ETV6 TF network MDS, ALL ASH education book 2016, 302-308 36
- 37. 2 types of RUNX1 mutation Dominant negative complete loss of RUNX1 activity Additional somatic mutation also affecting 2nd RUNX1 allele Monoallelic RUNX1 deletion produce haploinsufficiency Leads to thrombocytopenia alone Contribute to leukemia development Iléana Antony-Debré et al Blood 2015 125:930-940 37
- 38. • Defective downregulation of ANKRD26 by RUNX1/FLI1 • ANKRD26 overexpression • Mutated gene activate/phosphorylate MAPK/ERK pathway • ERK activation in megs: THC2 • ERK activation in myeloid cells: AML Proto-oncogene Marconi et al. Journal of Hematology & Oncology (2017) 10:18 5’UTR 38
- 39. ETV6: transcriptional repressor Proto-oncogene & TSG • Thrombocytopenia • Elevated MCV • Lymphoid: Myeloid neoplasm- 2:1 • Chemotherapy hypersensitivityNoetlzi L et al. Nature Genetics; 2015 nuclear localization and transcriptional repression DNA binding activities 39
- 40. • 10 families investigated • 5 families had a history of at least one person with platelet abnormalities • All demonstrated RUNX1 mutations • Incidence of MDS/AML among carriers of RUNX1 mutation was 35% • Transplantation: high incidence of early relapse Blood. 2008;112:4639-4645 40
- 41. 41
- 42. Syndromic predisposition Predisposition Inheritence Genes Altered pathway Hemat malignancy Fanconi Anemia AR, XLR FANC FANCS/BRCA1 & FANCD1/BRCA2 DNA damage/repair MDS, AML Dyskeratosis congenita AD, AR, XLR DKC1, TERC, TERT, TINF2.. Telomere maintenance MDS, AML Down Syndrome Sporadic Unknown Multifactorial AMKL, ALL RASopathies AD NF1, PTPN11, KRAS, NRAS, CBL RAS Signaling JMML Li Fraumani AD TP53 DNA damage/repair AML, ALL, sAML ASH education book 2016, 302-308 42
- 43. Down’s syndrome • Risk: ▫ AML: 150x ▫ ALL: 33x • AMKL: 500 times higher risk then gen population • High risk of toxicity from conventional chemo • High cure rate: EFS 80% • ALL: CRLF2 overexpression, targeted Tx with JAK2 inhibitors Saida S et al Int J Hematol 2016, 103:365–372 Roll et al Cancer Res. 2010, 70(19): 7347–7352. 43
- 44. Fanconi Anemia • Cumulative incidence of AML: 10% to 37% • De novo or progress from MDS • Mechanism of clonal evolution ▫ Defective DNA repair ▫ Hypersensetivity to cytokine: TNF α • Associated genetic anomalies ▫ +3q (41%) ▫ –7/7q (17%) ▫ – 11q (14%) ▫ Cryptic rearrangements of RUNX1 (21%) de Latour et al Blood 2016 127:2971-2979 44
- 45. Fanconi anemia de Latour et al Blood 2016 127:2971-2979 45
- 46. t-AML • BC survivors with TRL have family histories suggestive of inherited cancer susceptibility • Frequently carry germline mutations in BC susceptibility genes 46
- 48. Approach • History ▫ Occurrence in relative ▫ Age at diagnosis ▫ Associated solid tumor ▫ Congenital anomaly ▫ H/O cytopenia ▫ Unusual response to chemotherapy 48
- 49. Approach • Physical examination ▫ Café-au-lait ▫ Hypopigmentation ▫ Premature gray hair ▫ Nail dystrophy ▫ Short stature ▫ Radial defect ▫ Lacy reticular pigmentation of skin ▫ leukoplakia 49
- 50. Genetic counseling and testing • Informed decision and prepare for outcomes • Might be no positive outcome in-spite of significant family history • Genetic variant of uncertain significance • Periodic CBC: 2-3 months to 1 yr (no consensus) • Baseline BMB: CEBPA, GATA2, IBMFS 50
- 51. Preferred sample for testing • Overlapping phenotype- gene panel tested • DNA from skin fibroblast • Active disease: not possible to differentiate somatic or germline mutation • Cases with AlloSCT: donor DNA • Buccal/saliva: may contain high level of lymphocytes 51
- 53. Treatment considerations • Related donor for AlloSCT: identify mutation negative donors • Increased chemotherapy toxicity: down’s syndrome/FA/ETV6 • Increased toxicity to conventional myeloablative conditioning regimens: TBD/IBMFS • Lack of proven intervention to at-risk relatives 53
- 54. Take home message • Diagnosis may be challenging with resource limitations • Implication in patient management • Genetic counseling of family and identify those at risk • Genes identified are tip of iceberg • Much remains to be known 54
- 55. Thank You 55
Editor's Notes
- Cancer: accumulation of mutation that deregulates cell differentiation, proliferation and survival. Usually in post zygotic cell. Dr Paul Broca 1st described hereditary basis
- Pathogenesis of retinoblastoma. Two mutations of the RB locus on chromosome 13q14 lead to neoplastic proliferation of the retinal cells. In the sporadic form both mutations at the RB locus are acquired by the retinal cells after birth. In the familial form, all somatic cells inherit one mutant RB gene from a carrier parent. The second mutation affects the RB locus in one of the retinal cells after birth. 1st Altered copy of the growth regulatory gene in germline and 2nd inactivation of the susceptible gene is somatic and at varoius location. RB gene identified in 1986
- The role of RB in regulating the G1-S checkpoint of the cell cycle. Hypophosphorylated RB in complex with the E2F transcription factors binds to DNA, recruits chromatin-remodeling factors (histone deacetylases and histone methyltransferases), and inhibits transcription of genes whose products are required for the S phase of the cell cycle. When RB is phosphorylated by the cyclin DŔCDK4, cyclin DŔCDK6, and cyclin EŔCDK2 complexes, it releases E2F. The latter then activates transcription of S-phase genes. The phosphorylation of RB is inhibited by CDKIs, because they inactivate cyclin-CDK complexes. Virtually all cancer cells show dysregulation of the G1-S checkpoint as a result of mutation in one of four genes that regulate the phosphorylation of RB; these genes are RB1, CDK4, the genes encoding cyclin D proteins, and CDKN2A (p16). EGF, epidermal growth factor; PDGF, platelet-derived growth factor; TGF-β, transforming growth factor-beta
- The role of p53 in maintaining the integrity of the genome. Activation of normal p53 by DNA-damaging agents or by hypoxia leads to cell cycle arrest in G1 and induction of DNA repair, by transcriptional up-regulation of the cyclin-dependent kinase inhibitor CDKN1A (p21) and the GADD45 genes. Successful repair of DNA allows cells to proceed with the cell cycle; if DNA repair fails, p53 triggers either apoptosis or senescence. In cells with loss or mutations of p53, DNA damage does not induce cell cycle arrest or DNA repair, and genetically damaged cells proliferate, giving rise eventually to malignant neoplasms. B, p53 mediates gene repression by activating transcription of miRNAs. p53 activates transcription of the mir34 family of miRNAs. mir34s repress translation of both proliferative genes, such as cyclins, and anti-apoptotic genes, such as BCL2. Repression of these genes can promote either quiescence or senescence as well as apoptosis.
- Molecular Pathogenesis and Therapy of Acute Myeloid LeukemiaRoss L. Levine M.D. Human Oncology and Pathogenesis Program Leukemia Service, Department of Medicine Memorial Sloan Kettering Cancer Center
- sometimes, loss of a single allele of a tumor suppressor gene reduces levels or activity of the protein enough that the brakes on cell proliferation and survival are released. Loss of gene function caused by damage to a single allele is called haploinsufficiency. Such a finding indicates that dosage of the gene is important, and that two copies are required for normal function. Loss of one allele induces cancer susceptibility; loss of two alleles induces cancer. Middle, classical haploinsufficiency. Loss of one allele is sufficient for induction of cancer.
- Arber DA et al, Blood 2016; doi:10.1182/blood-2016-03-643544
- The genetic landscape of inherited HM. The order of the 11 established germ line mutations is depicted based on their date of discovery. Mutations are broadly assigned to 3 groups according to phenotype: HMs alone (blue), associated bone marrow failure syndromes (red), or characteristic cytopenias and/or platelet dysfunction (yellow). The incidence of these mutations varies considerably with .10 pedigrees reported for mutations in RUNX1, TERC, CEBPA, TERT, ANKRD26, GATA2, and now DDX41 with, in certain families/genes, apparent clustering of myeloid and lymphoid malignancies.
- essential role in mediating granulocytic differentiation and cellular growth arrest. The cumulative incidence of relapse in familial AML was 56% at 10 years (n5 11), and 3 patients experienced ‡3 disease episodes over a period of 17 to 20 years. Durable responses to secondary therapies were observed, with prolonged median survival after relapse (8 years) and long-term overall survival (10-year overall survival, 67%). Our data reveal that familial CEBPA-mutated AML exhibits a unique model of disease progression, associated with favorable long-term outcomes.
- Pathogenic CEBPA mutations occur primarily within 2 discrete regions (see figure). N-terminal mutations are characteristically frameshift insertions or deletions, leading to forced translation of an alternate 30-kDa protein from within an internal start site, which exerts dominant-negative effects on the full length CEBPA protein. C-terminal in-frame insertions/deletions occur within the DNA binding or leucine zipper domains and disrupt DNA binding and dimerization. Distribution of germline and acquired CEBPA mutations in familial AML. Transactivation domain (TAD) 1 amino acids (AA) 70 to 97; p30 start codon (ATG), AA 120; TAD2, AA 126 to 200; DNA-binding domain (DBD), AA 278 to 306; leucine zipper domain (LZD), AA 307 to 358. All germline mutations were localized to the N-terminal, causing a frameshift preceding the internal CEBPA-p30 start codon. Somatic mutations (detailed in supplemental Table 2) clustered in the C-terminal of the gene, with a hotspot located at p.K313.
- Clinical data were collected from 10 CEBPA-mutated families, representing 24 members with acute myeloid leukemia (AML). Germline CEBPA mutations clustered within the N-terminal and were highly penetrant, with AML presenting at a median age of 24.5 years (range, 1.75-46 years). In all diagnostic tumors tested (n518), doubleCEBPAmutations (CEBPAdm)were detected, with acquired (somatic) mutations preferentially targeting the C-terminal. Somatic CEBPA mutations were unstable throughout the disease course, with different mutations identified at recurrence
- Somatic CEBPA mutations were unstable throughout the disease course, with different mutations identified at recurrence. Such observations of late chemotherapysensitive recurrence support the hypothesis of novel, independent clones initiating new leukemic episodes. More importantly for clinicians, they reveal important biological and prognostic insights into the clinical disease course of familial AML.
- Probable ATP-dependent RNA helicase DDX41 is an enzyme that in humans is encoded by the DDX41 gene.[3][4] DEAD box proteins, characterized by the conserved motif Asp-Glu-Ala-Asp (DEAD), are putative RNA helicases. They are implicated in a number of cellular processes involving alteration of RNA secondary structure, such as translation initiation, nuclear and mitochondrial splicing, and ribosome and spliceosome assembly. Based on their distribution patterns, some members of the DEAD box protein family are believed to be involved in embryogenesis, spermatogenesis, and cellular growth and division. This gene encodes a member of this family. The function of this member has not been determined. Based on studies in Drosophila, the abstrakt gene is widely required during post-transcriptional gene expression Granulomatous and immune disorders presenting prior to HM in germ line mutation carriers were observed
- Somatic mutations of DDX41 affected the ATP binding domain (Figure 2A). Because germline mutations were predominantly out-of-frame insertions and they coincided with somatic DDX41 mutations suggested that defects of this gene may result in a loss of function. Consequently, deletion and mutations of this gene may be functionally equivalent. Deletions of the long arm of chr.5 involving the DDX41 locus (5q35.3) were present in 6% of all cases and 26% of the del(5q) cases
- DDX41 is located at the distal end of chromosome 5q, 5q35.3, and encodes a protein that contains three known domains and ATP binding sites, as illustrated. The pink bars visualize deletions of chromosome 5q in our MDS cohort that include the DDX41 locus. The red triangles indicate DDX41 mutations in patients with hematological malignancies from our cohort and TCGA. Red circles indicate the identified germline mutations of DDX41 (p.Q52fs, p.D140fs, p.M155I, and p.I396T). The p.R525H mutation was detected in 13 out of 1,045 cases. Purple triangles show DDX41 mutations in non hematological malignancies. Sanger sequencing confirming recurrent germline mutation (p.D140fs; left) and somatic mutation (p.R525H; right) of DDX41 are shown.
- Response rate to lenalidomide in patients with DDX41 mutants (n = 8/8) compared with WT cases (n = 55/103). p = 0.01. Response rate to lenalidomide treatment of patients with DDX41 mutations and/or low DDX41 expression (n = 7/9) compared with others (n = 2/10). Cases with high and low DDX41 expression were dichotomized by the mean of relative mRNA transcription levels (mean = 3.85 relative mRNA expression).
- Overall survival analysis in patients with DDX41 mutations or deletions compared with WT cases(HR = 3.5; 95% CI = 2.0–5.9; and p < 0.0001). (D) Overall survival analysis in patients with low DDX41 mRNA expression compared with patients with higher expression (HR = 1.6; 95% CI = 1.0– 2.2; and p = 0.029). Cases with high and low DDX41 expression were dichotomized by the mean of relative mRNA transcription levels (mean = 3.85 relative mRNA expression).
- Domain structure of GATA2 showing the positions of the alterations. The positions of the p.Thr354Met, p.Thr355del, AML-M5 (ref. 7) (green) and CML blast crisis alterations are shown with respect to zinc finger (ZF) 1 and 2, the transactivation domain (TA) and the nuclear localization signal (NLS). GATA2 belongs to a family of zinc finger transcription factors that are critical regulators of gene expression in hematopoietic cells. Although GATA1 functions primarily in erythropoiesis and megakaryopoiesis and GATA3 in T-cell lymphopoiesis, GATA2 is particularly critical for the genesis and function of hematopoietic stem and progenitor cells and thus all subsequent blood cell lineages. GATA2 haploinsufficiency perturbs normal hematopoietic stem cell homeostasis in murine models
- several first-degree relatives have a history of opportunistic infections and myeloid disorders, suggesting an etiologic link to this disorder. GATA2 plays an essential role in maintaining the proliferation and survival of early hematopoietic cells, as well as preferential differentiation to erythroid or megakaryocytic lineages Presence of monocyte and plasma cell at infection site
- GATA2 regulates phagocytosis by pulmonary alveolar macrophages, which may explain the common occurrence of pulmonary alveolar proteinosis in MonoMAC.15 The significant human papillomavirus and other viral infections in the MonoMAC syndrome probably reflect the profound absence of NK cells in MonoMAC. The relatively narrow spectrum of infections is distinct from the neutropenia of MDS and draws a surprising connection between infection susceptibility and predeliction to myeloid malignancy. 29% of patients with a germline GATA2 mutation had an acquired ASXL1 mutation identified within the hematopoietic malignancy,42 suggesting cooperativity between these two genes. ASXL1 encodes a polycomb-associated protein that co-localizes with ETS (E-twenty six) transcription factors and influences histone modifications.
- The mechanism by which GATA2 p.Thr354Met and p.Thr355del alterations function is distinct to that generally described for RUNX1 and CEBPA, which commonly act as classical tumor suppressor genes with a wide range of mutations and requiring functional disruption of both alleles. Transcription factors are well characterized as targets of dominant negative or constitutively active mutations in cancer, with RUNX1 mutations leading to a spectrum of outcomes including AML and acute lymphoblastic leukemia, consistent with both tumor suppressor gene and dominant oncogene models (A) Hypocellular bone marrow with trilineage hypoplasia (36-year-old woman), bone marrow core biopsy hematoxylin and eosin stain. (B-D) Atypical megakaryocytes, small mononuclear (B), and dysplastic forms with separated nuclear lobes
- Cutaneous M abscessus infection and (F) skin biopsy AFB smear showing abundant mycobacteria. (G) Computed tomography scan of the chest showing the typical crazy paving pattern of PAP and (H) lung biopsy hematoxylin and eosin stain showing alveolar filling with lipoproteinaceous material. (I) Recalcitrant periungual warts, (J) perineal condyloma, and vulvar/anal intraepithelial neoplasia. (K) Panniculitis/erythema nodosum involving the anterior shins. (L) Computed tomography scan of the abdomen with multiple hypodense liver lesions (arrows) and (M) liver biopsy showing EBV related spindle cell tumo rwith in situ hybridization of EBV-encoded messenger RNA using EBV probe antifluorescein antibody and bond polymer refine detection. (N) Unilateral lymphedema of the right lower extremity. (O) Magnetic resonance imaging of the brain with embolic infarcts in the right occipital and left parietal lobes (arrows) in the setting of culture-negative endocarditis.
- In the absence of cytogenetic abnormalities or overt dysplasia, the hypoplastic bone marrow of GATA2 deficiency may also be initially diagnosed as aplastic anemia. Vigilant bone marrow monitoring is prudent in patients with GATA2 deficiency, as the development of excess blasts or abnormal cytogenetics may influence the timing of HSCT. Somatic mutations in ASXL1 have also been implicated in leukemic transformation and might accelerate the timing for HSCT
- cartoon depicts the process of megakaryopoiesis and platelet formation. Each of the 51 known IPD genes are indicated and categorized according to their effect on megakaryocyte and platelet biology. *IPDs typically associated with phenotypes outside of the blood system. HSC, hematopoietic stem cell.
- Thrombocytopenia 2 is an autosomal dominant thrombocytopenia with moderate thrombocytopenia, demonstrating normal mean platelet volumes; elevated thrombopoietin levels; variable aggregation defects in response to collagen, adenosine diphosphate, or ristocetin; and bone marrow examinations revealing dysmegakaryopoiesis with small megakaryocytes and hypolobulated nuclei
- evolution to leukemia depends on the type of mutations (ie, mutations maintaining CBFβ-binding properties) to generate dominant-negative (DN) proteins that favor leukemic evolution RUNX1 alterations that predispose to leukemia, mainly DN-like mutants,3,4 deregulate critical hematopoietic stem cell (HSC) and progenitor target genes such as NR4A3,4 leading to the amplification of a pool of cells susceptible to acquiring additional somatic mutations, sometimes affecting the second RUNX1 allele. similar recurrent secondary aberrations have not been observed in FPD/AML; in particular, acquired mutations in the wild-type RUNX1 allele are not observed. Familial RUNX1 mutations generally cluster to the N-terminal region of the gene within the RHD in exons 3 to 5, often disrupting DNA binding but allowing ongoing dimerization with CBF-. Familial mutations have been reported less frequently in the C-terminal region, and these typically maintain DNA binding and dimerization with loss of the trans-activation region. The nature of RUNX1 mutations in de novo MDS/AML appears similar to that seen in FPD/AML with a predominance of point mutations in the RHD
- ETV6 is a 57-kDa protein with 452 amino acids and 3 functional domains: the N-terminal pointed (PNT), central regulatory and C-terminal DNA-binding (ETS) domains (Fig. 1c). The nuclear localization and transcriptional repression activity of ETV6 require homodimerization via the pointed domain. ETV6 modulates the activity of other ETS transcription factors such as FLI1 for which hemizygous deletions of the corresponding gene cause Paris-Trousseau–related thrombocytopenia and several other known ETV6 interaction partners that are present in platelets and presumably in megakaryocytes. The function of the central domain is not well understood, but this domain has been shown to undergo post-translational modifications and to be essential for the repressive function of ETV6 in in vitro reporter assays. Top, schematic of ETV6, which is composed of eight exons encoding UTRs (yellow) and protein-coding sequences (blue). The two mutations identified, c.641C>T in exon 5 and c.1252A>G in exon 7, are depicted. Bottom, schematic of the ETV6 protein showing its different domains, pointed (PNT), central and ETS (the DNA-binding domain), and the location of the amino acid changes. grade 3 myelosuppression following exposure to anti-metabolite therapy; this feature of chemotherapy hypersensitivity was shared by another patient with T-/myeloid mixed phenotype leukemia and a germline ETV6 mutation ETV6 is a clinically significant proto-oncogene in that it can fuse with other genes to drive the development and/or progression of certain cancers. However, ETV6 is also an anti-oncogene or tumor suppressor gene in that mutations in it that encode for a truncated and therefore inactive protein are also associated with certain types of cancers. It binds to and thereby inhibits FLI1, another member of the ETS transcription factor family, which is active in promoting the maturation of blood platelet-forming megakaryocytes and blocking the Cellular differentiation of erythroblasts into red blood cells; this results in the excessive proliferation and abnormal morphology of erythroblasts
- 18 FANC genes
- Proposed model of ML-DS pathogenesis. Evolution of myeloid leukemia in children with Down syndrome. Int J Hematol 2016, 103:365–372, with permission from the Japanese Society of Hematology. Trisomy 21. vertical lines; GATA1 mutation, black and white triangles; cohesin, CTCF, and other epigenetic regulator mutation, black circle; kinase-signaling molecule mutation, black star. cytokine receptor-like factor 2, CRLF2-overexpressing primary lymphoid progenitor cells from mouse fetal liver showed evidence of increased JAK2/STAT5 signaling (4). The frequency of JAK2 mutations in ALL has been reported to be about 10% in pediatric high-risk ALL and about 20% in DS-ALL
- 25%to40%ofpatientslack physicalabnormalitiesassociatedwiththedisease.97 Althoughthemedianageofdiagnosisis6.5yearsforboys and8yearsforgirls,thediseaseisdiagnosedthroughout pediatricandadultagegroups.Themedianageofonsetof bonemarrowfailureissevenyears.98 Mediansurvivalfor patientswithFAis24years,withacumulativeincidenceof 90%ofbonemarrowfailurebyage40years.99 At least 20%ofpatientswithFAdevelopmalignancy,withAML beingthemostcommondiagnosis The European Group for Blood and Marrow Transplantation analyzed 795 FA patients who underwent initial HSCT between 1972 and 2010, with 58 patients transplanted for MDS or AML.15 OS rates at 1, 3, and 5 years were 44%, 41%, and 35%, respectively. The cumulative incidence of relapse at 2 years was 11%. There was no difference in OS by donor type (related vs unrelated).
- We defined provisional cytogenetic/molecular categories by reference to MDS/AML literature in FA6,7,27,33,34 and non-FA49,50patients (also see text). RUNX1-abn can include RUNX1 gene mutation, deletion, and/or translocation.7 Genetic reversion (hematopoietic somatic mosaicism) is not indicated in the figure to keep it simple. bThe timing of BM monitoring is discussed, especially because repeated aspiration is poorly tolerated in children, adolescents, and young adults.40 The overall consensus is that a 1-year basis of BM aspirate is reasonable and should be adapted in response to blood cell count changes, myelodysplasia signs, increased blast proportion, and/or cytogenetic evidence of clonal evolution. Conversely, BM monitoring is likely to be slightly delayed in FA children under age 10 years (except in BRCA2/FANCD1 patients), given the rarity at this age and relatively slow pace of clonal progression. cThe classical reduced-intensity conditioning (RIC) regimen consists of 90 mg/m2 of fludarabine (30 mg/m2 on days −4, −3, and −2) and 40 mg/kg of cyclophosphamide (10 mg/kg on days −5, −4, −3, and −2) in the case of matched related donor. One might argue that, in the case of MDS/AML, low-dose cyclophosphamide/fludarabine alone would not suffice as conditioning therapy due to the substantial number of residual host cells early after transplant with this approach and the risk of relapse; an alternative is the use of TBI (2-3 Gy) in patients with MDS/AML and an HLA- matched sibling donor.16,48 In the case of matched unrelated donor, the conditioning regimen consists of fludarabine (120 mg/m2), cyclophosphamide (40 mg/kg), and TBI (2 Gy). GVHD prophylaxis consists of mycophenolate acid and cyclosporine. Anti-thymocyte globulin is used in total doses of 5 mg/kg in the case of matched unrelated donor only. Others favor ex vivo T-cell depletion with an add-back of T cells to achieve a fixed graft T-cell dose of 1 × 105 CD3 cells per kilogram recipient.62 In the case of CB HSCT, we do not use anti-thymocyte globulin in the conditioning regimen. dOthers do not recommend cytoreduction, except in patients with BRCA2 mutations16,48,62; the sequential strategy comprising pretransplant chemotherapy with fludarabine (30 mg/m2 per day for 5 days) and cytarabine (1 g/m2 twice per day for 5 days) with granulocyte colony-stimulating factor injections (FLAG), followed 3 weeks later by an RIC regimen (4 days of cyclophosphamide, 10 mg/kg; 4 days of fludarabine, 30 mg/m2; and TBI, 2 Gy) delivered during chemotherapy-induced aplasia. Again, anti-thymocyte globulin is used in total doses of 5 mg/kg in the case of matched unrelated donor only. In the case of CB HSCT, we do not use anti-thymocyte globulin in the conditioning regimen. eScreening for malignancies, including oropharyngeal, dental, and gynecological examinations, forms part of long-term patient care. Long-term multidisciplinary surveillance is also mandatory for all patients post-HSCT.40 The multiple problems in early age, subsequent requirements for HSCT, and continuing poor prognosis in survivors due to cancer susceptibility are a source of stress for FA patients and their families. Adequate psychosocial support and a coordinated, multidisciplinary team with dedicated physicians are the cornerstones to successful management.40 CGH, comparative genomic hybridization; FLAG, fludarabine/cytarabine/granulocyte colony-stimulating factor; GVHD, graft-versus-host disease; SIC, severe isolated cytopenia; SNP, single nucleotide polymorphism.
- 8 female BC survivors were identified, A family cancer history was available for 70 subjects (80%), among whom 40 (57%) reported at least one first- or second-degree relative with breast, ovarian, or pancreatic cancer. Most BC patients developed t-MN (n=81; 92%), but 7 cases (8%) of t-ALL were also observed (Table 1). The median latency from first cytotoxic exposure to TRL diagnosis among the 86 patients for whom latency was available was 58 months (interquartile range (IQR), 28–105 months). Clonal cytogenetic abnormalities were observed in 77 of the 84 subjects (92%) with an available karyotype. Among these, abnormalities of chromosomes 5 and/or 7 and recurring balanced translocations were both common, occurring in 51% (n=43) and 35% (n=29) of patients,
- Adaptar ligation is don to attach the target genome to the flow cell (plate). Both the adapters attach to the corresponding oligos on the flow cell and the PCR reaction creats a double stranded bridge, bridge is then linearised and the reverse strand is washed off. When all the oligos of the flow cell is saturated, the sequencing reaction occurs with fluorescence tagged NTPs