![• Therapy related myeloid neoplasm
– Alkylating agents/ionizing radiation:
• whole or partial loss of chromosome 5 and/or 7 ,
associated with one or more additional chromosomal
abnormalities(e.g. del(13q), del(20q), del(11q),
del(3p), -17, -18, -21, +8) in a Complex karyotype
– Topoisomerase II inhibitor
• Balanced chromososmal translocations that involve
rearrangement 11q23 [t(9;11)(p22,q23) & t(11;19)
(q23;p13)], 21q22 [t(8;21) (q22;q22), t(3;21)
(q26.2;q22.1)] , t(15;17)(q22;q12) and inv (16)
(p13q22)](https://image.slidesharecdn.com/who2008-250516155734-523ae3fc/85/WHO-2008-classification-of-hematolymphoid-neoplasms-ppt-28-320.jpg)
WHO 2008 classification of hematolymphoid neoplasms.ppt
- 1. WHO 2008
- 2. Investigations required to classify Hematolymphoid Neoplasms as per WHO classification 2008 Hematopathology Laboratory, Tata Memorial Hospital, Mumbai.
- 3. • Cytochemistry • Flow cytometry • Immunohistochemistry • Cytogenetics & molecular cytogenetics • Molecular diagnostics • Follow up investigations or treatment modalities
- 4. Cytochemistry • Myeloperoxidase: myeloid lineage • NSE : monocytic lineage • CAE : • Iron / Perl’s stain : for demonstration of ring sideroblasts • PAS: erythroid precursors • LAP/NAP(neutrophil alkaline phosphatase) • TRAP: Hairy cell leukemia
- 6. Acute lymphoblastic leukemia • B lymphoblastic leukemia • CD19, CD10, CD22, CD24, PAX5(specific), HLA-DR • cytoCD22, cytoCD79a • CD34, TdT • NG2 for MLL gene rearrangement • T lymphoblastic leukemia • CD1a,CD2, CD5, CD4, CD8, CD7, CD56, CD16, CD10 • CD3, cytoCD3(lineage specific) • CD34, TdT
- 7. Acute myeloid leukemia • Myeloid cell – CD13, CD33, CD117, CD15, CD65, MPO • Erythroid precursors: – CD235a/glycophorin , CD71, antibody to Hb A • Megakaryocytes: – CD41, CD42, CD61 • Monocytic markers – CD4, CD11b, CD11c, CD14, CD64, CD36, CD163, CD68, lysozymes • Basophils: – CD123, CD203c, (CD9)
- 8. Myelodysplastic syndrome • Myeloid maturation pattern • Refractory anemia/RARS • Aberrant immunophenotypic features of erythroid precursors • Refractory anemia with excess of blasts: blasts are positive for CD34, CD117, CD38, HLADR, CD13, CD33,CD15,CD11b, CD65, aberrant expression of CD7 OR CD56.
- 9. Lymphoma • Bcell markers: CD20, CD19, CD10, CD22, CD24, PAX5(specific), , CD5, surface Ig, kappa, lambda light chain, CD103, CD11c, CD25, • ALK, NPM, CD30, • T cell markers: CD1a,CD2, CD5, CD4, CD8, CD7, CD56, CD16, TCR αβ, TCR γδ • NK cell markers: cyto CD3ε, CD56 • Plasma cell markers: CD38, CD138, CD56, CD19, cyto kappa, cyto lambda
- 10. Lymphoma • CLL: expression of CD38 & ZAP-70: adverse prognosis • Mantel cell lymphoma : cyclin D1 • Hairy cell leukemia: CD123 • Annexin A1: most specific marker fro HCL as it is not expressed inany other B cell lymphoma: by immunohistochemistry: but not for monitoring minimal residual disease • T-bet, DBA.44, TRAP
- 12. Acute lymphoblastic leukemia • B lymphoblastic leukemia • CD20, CD10, CD22, CD24, PAX5(specific), HLA-DR • CD79a • CD34, TdT • T lymphoblastic leukemia • CD1a,CD2, CD5, CD4, CD8, CD7, CD56, CD16, CD10 • CD3, cytoCD3(lineage specific) • CD34, TdT
- 13. Acute myeloid leukemia • MPO • CD4, CD68 • CD235a/glycophorin , CD71, antibody to Hb A • Acute megakaryoblastic leukemia: – Platelet glycoprotein(CD42b, CD61): lineage specific – von Willebrand’s factor, LAT( linker of activation of T cells)
- 14. Myelodysplastic syndrome • CD34: to confirm the presence of an increased number of blasts • Cd61, cd42B: to identify micromegakaryodytes or other small dysplastic forms
- 15. Myeloproliferative Neoplasms • CD 34: CML- AP/BP
- 16. Lymphoma • Bcell markers: CD20, CD19, CD10, CD22, CD24, PAX5(specific), , CD5, surface Ig, kappa, lambda light chain, CD103, CD11c, CD25, • EMA, ALK, NPM, CD30, • BCL2, BCL6, Annexin A1, • T cell markers: CD1a,CD2, CD5, CD4, CD8, CD7, CD56, CD16, TCR αβ, TCR γδ, CXCL13,PD1, FoxP3 • NK cell markers: cyto CD3ε, CD94, CD161, CD16, CD56
- 17. Plasma cell neoplasms • Plasma cell markers: • CD38, CD138, CD79a, • Aberrant expression of CD56 • Aberrant expression CD117, CD20, CD52, CD10 • CD19: negative • cytoplasmic Ig, cyto kappa, cyto lambda
- 18. Classical Hodgkin’s Lymphoma • HRS cells : • CD45, J chain, CD20, CD79a : negative, • CD30, CD15, PAX5/BSAP: positive • IRF4/MUM1: positive • Express LMP1 & EBNA-1 without EBNA-2 • EMA, ALK: negative • OCT-2 & BOB.1 and PU.1: negative
- 19. NLPHL • LP cells: • Positive for: • CD20, CD79a, CD75, BCL-6, CD45 & J chain • EMA > than 50% cases • OCT-2,BOB.1 AID: coexpressed • Negative for : CD15, CD30 • Background cells: CD4+/CD57+ T cells & express markers like c-MAF. PD-1, BCL 6, IFR4/MUM1, CD134& NOT IL2, IL4, CD8, CD40, TIA1
- 20. Histiocytic & dendritic cell neoplasms • Langerhan’s cell histiocytosis/ sarcoma • CD1a, langerin, S100 • Other markers: CD68, vimentin, HLA-DR
- 22. Acute lymphoblastic leukemia • B lymphoblastic leukemia with recurrent genetic abnormalities • t(9;22) (q34;q11.2)BCR-ABL1 • t(v;11q23) MLL rearranged • t(12;21)(p13;q22)(ETV6-RUNX1) / TEL-AML1 • Hyperdiplody : flow cytometry/ conventional cytogenetics/ FISH • Hypodiploidy • ALL (with eosinophilia) t(5;14) IL-3-IGH • t(1;19) E2A-PBX1/TCF3-PBX1: poor prognosis
- 23. Acute lymphoblastic leukemia • T lymphoblastic leukemia : most common recurrent cytogenetic abnormalities • Abnormal α & δ TCR loci at : 14q11.2 • β TCR locus at : 7q35 • γ TCR locus at : 7p14-15 • Other abnormalities: del(9p): loss of CDKN2A
- 24. Acute myeloid leukemia • Acute myeloid leukemia with recurrent genetic abnormalities • t(8;21) (q22;q22) RUNX1-RUNX1T1 / AML-ETO • inv(16) or t(16;16) (p13.1;q22)CBFB-MYH11 • t(15;17) (q22;q12) PML-RARA • t(11;17) (q23;q12) ZBTB16-RARA • t(5;17) (q35;q12) NPM1-STAT5B
- 25. • Acute myeloid leukemia with recurrent genetic abnormalities • t(9;11) (p22;q23)MLLT3-MLL: – 1/3rd MLL : not detected by conventional karyotyping, so FISH or molecular studies are not important. • t(6;9) (p23;q34)DEK-NUP214 • Inv(3) or t(3;3) (q21;q26.2)RUN1-EVI1 • t(1;22) (p13;q13) RBM15-MKL1
- 26. • Acute myeloid leukemia with myelodysplasia-related changes • Complex karyotype : > 3 unrelated abnormalities • Unbalanced abnormalities -7/del(7q), -5/del(5q), i(17q)/t(17q), -13/del(13q), del(11q), del(12p)/t(12p), del(9q), idic(X)(q13)
- 27. • Acute myeloid leukemia with myelodysplasia-related changes • Balanced abnormalities t(11;16)(q23;p13.3), t(3;21)(q26.2,q22.1), t(1;3)(p36.3;q21.1), t(2;11)(p21;q23) t(5;12)(q33;p12), t(5;7)(q33;q11.2), t(5;17)(q33;p13), t(5;10)(q33;q21) t(3;5)(q25;q34)
- 28. • Therapy related myeloid neoplasm – Alkylating agents/ionizing radiation: • whole or partial loss of chromosome 5 and/or 7 , associated with one or more additional chromosomal abnormalities(e.g. del(13q), del(20q), del(11q), del(3p), -17, -18, -21, +8) in a Complex karyotype – Topoisomerase II inhibitor • Balanced chromososmal translocations that involve rearrangement 11q23 [t(9;11)(p22,q23) & t(11;19) (q23;p13)], 21q22 [t(8;21) (q22;q22), t(3;21) (q26.2;q22.1)] , t(15;17)(q22;q12) and inv (16) (p13q22)
- 29. • Myeloid proliferation related to Down syndrome • Transient abnormal myelopoiesis : – trisomy 21, acquired GATA1 • Myeloid leukemia associated with Down syndrome : – trisomy 21, somatic mutations of GATA1 – trisomy 8
- 30. Blastic plasmacytoid dendritic cell neoplasm • 2/3rd cases: abnormal karyotype • specific chromosomal are lacking but complex karyotypes are common: 5q21 or 5q34, 12q13, 13q13-21, 15q & loss of chromosome 9
- 31. Acute leukemias of ambiguous lineage • Mixed phenotype acute leukemia with t(9;22) (q34;q11.2)BCR-ABL • Mixed phenotype acute leukemia with t(v;11q23) MLL rearranged • MPAL (B+Myeloid): del6p, del 5q, structural abnormalities 7, complex karyotype • MPAL (T+Myeloid): MLL ,t(9;22)
- 32. Myelodysplastic syndrome • IPSS (International Prognostic Scoring System) for MDS: • Prognostic variables 0 0.5 1 1.5 2 • % bone marrow blasts <5% 5-10% 11-19% 20% • Karyotype: Good Inter. Poor • Cytopenia 0-1 2-3 • Karyotype: • Good - normal, -Y, del(5q), del(20q) • Poor - complex(>3 abnormalities) or chromosome 7 anomalie • Intermediate – other abnormalities
- 33. Myelodysplastic syndrome • Balanced abnormalities t(11;16)(q23;p13.3), t(3;21)(q26.2,q22.1), t(1;3)(p36.3;q21.1), t(2;11)(p21;q23) inv(3)(q12;q26.2) t(6;9)(p23;q34)
- 34. Myeloproliferative Neoplasms • Philadelphia chromosome T(9;22)BCR-ABL-1 • JAK-2 mutation – JAK -2 V617F , JAK -2 exon 12 mutation • MPL W515K/L • Additional abnormalitites: – CML-AP/BP: extra Ph, +8, +19, I(17q) – Polycythemia vera: common abnormalities +8, +9, del20q, del13, del9p – Primary myelofibrosis: del(13) & der6,t(1;6): strongly s/o, but not diagnostic
- 35. Myeloid & lymphoid neoplasm with eosinophilia & abnormalitites of PDGFRA, PDGFRB & FGFR1 • aberrant tyrosine kinase activity • MPN related with rearrangement of PDGFRA or PDGFRB are responsive to imatinib & some related tyrosine kinase inhibitors. • But specific therapy is not developed fro FGFR1 related diseases.
- 36. Myeloid neoplasm with abnormalities of PDGFRA rearrangement • FIP1L1-PDGFRA fusion gene resulting from a cryptic del(4)(q12) • Chromosomal rearrangement with 4q12 breakpoint – t(1;4)(q44;q12) – t(4;10)(q12;p11) – t(2;4)(p24;q12) STRN-PDGFRA – t(4;12)(p2?3;q1?2) ETV6-PDGFRA – t(4;22)(q12;p11)BCR-PDGFRA – FISH analysis using probe for the CHIC2 gene – RT-PCR, nested PCR
- 37. Myeloid neoplasm with abnormalities of PDGFRB rearrangement • Breakpoint 5q31 or 5q33 PDGFRB (FISH analysis) • Rearrangement of PDGFRB at 5q31-33 t(5;12)(q31~33;p12) ETV6- PDGFRB t(1;3;5) (p36;p21;q33)WDR48- PDGFRB der(1)t(1;5)(p34;q33), der(5)t(1;5)(p34;q15), der(11)ins(11;5)(p12;q15;q33): GPIAP1- PDGFRB t(1;5)(q21;q33)TPM3- PDGFRB t(1;5)(q23;q33)PDE4DIPB- PDGFRB t(4;5;5)(q23;q31;q33) PRKG2- PDGFRB
- 38. • Break point 5q33 PDGFRB t(3;5)(p21-25;q31-q35) GOLGA4-PDGFRB t(5;7)(q33-q11.2)HIP1-PDGFRB t(5;10)(q33;q21) CCDC6-PDGFRB t(5;12)(q31-33;q24) GIT2-PDGFRB t(5;14)(q33;q24)NIN-PDGFRB t(5;14)(q33;q32)KIAA1509-PDGFRB t(5;15)(q33;q22) TP53BP1-PDGFRB t(5;16) (q33;q13) NDE1- PDGFRB t(5;17) (q33;q13) RABEP1-PDGFRB t(5;17)(q33;q11.2) SPECC1-PDGFRB
- 39. • FISH analysis (with PDGFRB probe) is indicated in all patients with presumptive diagnosis of MPN. • RT-PCR, using primers suitable for all known breakpoint is recommended to confirm ETV6- PDGFRB . • FISH analysis dose not always demonstrate rearrangement of PDGFRB even when it is detectable on Southern Blot analysis • But molecular studies are not indicated if there is no 5q31~33 breakpoint on classical cytogenetic analysis
- 40. • Not all translocations characterized as t(5;12) (q31~33;p12) leads to ETV6- PDGFRB fusion. • Cases without a fusion gene are not of this category of MNP and are not likely to respond to imatinib. • But if molecular analysis is not available, a trail of imatinib is justified in patients with an MPN associated with t(5;12).
- 41. Myeloid neoplasm with FGFR1 abnormalities • Variety of translocation with an 8q11 brakpoint • t(8;13)(p11;q12)ZNF198- FGFR1 • t(8;9)(p11;q33) CEP110-FGFR1 • t(6;8)(q27;p11-12) FGFR1OP1-FGFR1: with polycythemia vera • t(8;22)(p11;q11) BCR-FGFR1: with basophilia • t(7;8)(q34;p11) TRIM24-FGFR1 • t(8;17)(p11;q23) MYO18A-FGFR1 • t(8;19)(p12;q13.3) HERVK-FGFR1 • ins(12;80(p11;p11p22) FGFR1OP2-FGFR1 • Trisomy 21: secondary cytogenetic abnormality
- 42. Myeloid neoplasm with FGFR1 abnormalities • Poor prognosis • No established tyrosine kinase inhibitor therapy • Interferon has induced a cytogenetic response in several patients • PKC142 is effective in occasional case.
- 43. Myelodysplastic/myeloproliferative diseases • Absence of Philadelphia chromosome, PDGFRA, PDGFRB • JAK-2 mutation +/- • RAS mutation • CMML: – Clonal cytogenetic abnormalities: non specific – structural abnormalities of 12p, trisomy8, del7/-7, i(17q) • Atypical CML: – t(8;9)(PCM1-JAK2)
- 44. Myelodysplastic/myeloproliferative diseases • JMML: – Monosomy 7: 25% – Normal karyotype: 65% – Other abnormalities: 10% • MDS/MPD, Unclassified – No specific cytogenetic or molecular abnormalities
- 45. Lymphoma • CLL: by FISH – del 13q14.3: (isolated) : favourable clinical course – trisomy 12 – Deletions of 11q22-23, 17p13 and 6q21: worse outcome
- 46. • Splenic marginal zone lymphoma – Allelic loss of chromosome 7q31-32 – Dysregulation CDK6 gene at 7q21 – Trisomy 3q CCND1 rearrnagement, t(11;14), cyclinD1 expression • Nodal marginal zone lymphoma: – trisomies 3, 18, 7 – Tranlocations associated with extranodal MZL are not detcted.
- 47. • Hairy cell leukemia: – no specific cytogenetic abnormality : numerical abnormalities of chromosome 5 & 7 have been described • Burkitt’s lymphoma : – MYC translocation/rearrrangement at band 8q24 by FISH – t(8;14)(q24;q32) (IGH :MYC) – t(8;22)(q24;q11)(lambda: MYC) – t(2;8)(p12;q24)(kappa: MYC)
- 48. Immunophenotypic & genetic features useful in distinguishing BL from DLBCL CHARACTERISTIC BL Intemediate BL/DLBCL DLBCL Proliferation(Ki67/MIB1) > 90% & homogeneous Yes Common rare < 90% & heterogeneous No Sometimes common BCL2 expression Negative/weak Yes Sometimes Sometimes Strong No Sometimes Sometimes
- 49. Genetic features BL Intemediate BL/DLBCL DLBCL MYC rearrangement Yes Common Rare IG-MYC Yes Sometimes Rare Non IG-MYC No Sometimes Rare BCL2 but no MYC rearrangement No Rare Sometimes BCL6 but no MYC rearrangement No Rare Sometimes Double hit No Sometimes Rare MYC-simple karyotype Yes Rare Rare MYC-complex karyotype Rare Common Rare Immunophenotypic & genetic features useful in distinguishing BL from DLBCL
- 50. Anatomic site distribution & frequency(%) of chromosomal translocations & trisomies 3 & 18 in MALT lymphomas Site t(11;18) (q21;q21) t(14;18) (q32;q21) t(3;14) (p14.1;q32) t(1;14) (p22;q32) +3 +18 stomach 6-26 1-5 0 0 11 6 Intestine 12-56 0 0 0-13 75 25 occular adnexa/orbit 0-10 0-25 0-20 0 38 13 Salivary glands 0-5 0-16 0 0-2 55 19 lung 31-53 6-10 0 2-7 20 7 skin 0-8 0-14 0-10 0 20 4 thyroid 0-17 0 0-50 0 17 0
- 51. Genetic abnormalities in folicular lymphoma • Cytogenetic Abnormalities (FISH : most sensitive & specific method) • t(14;18)(q23;q21) 80% • +7 20% • +18 20% • 3q27-28 15% • 6q23-26 15% • 17p 15% • Oncogene abnormalities • BCL2 rearranged 80% • BCL6 rearranged 15% • BCL65’ mutation 40%
- 52. Mantle cell lymphoma naïve B early classical blastoid lymphocyte MCL MCL MCL Germline ATM CHK2 t(11;14) ATM p16/CDK4/Rb cyclinD1 CHK2 ARF/MDM2/p53 Rb p27 increased high genomic instability proliferation
- 53. Mantle cell lymphoma • t(11;14)(q13;q32)IGH@-CCND1 – Very high levels of cyclin D1 expression – high proliferation- more aggressive clinical behaviour • High number of secondary non-random secondary chromosomal aberrations: – gain of 3q26, 7p21, 8q24(MYC)(aggressive), loss of 1p13-p31, 6q23-q27, 9p21, 11q22-q23, 13q11-q13, 13q14-q34, 17p13-pter – Trisomy12 • Cyclin D1 negative MCL: – high expression of cyclin D2 or cyclin D3 – t(2;12)(p12;p13)cyclinD2-kappa light chain
- 54. Genetic, molecular & clinical characteristics of DLBCL subgroups &PMBL recognized by expression profiling Characheristic ABC (activated B cell like) GCB (germinal center B cell like) PMBL (primary mediastinal large B cell lymphoma) t(14;18) 0 35 0 3q gain/ amplification 26 0 5 9p gain/amplification 6 0 35 12q12 5 20 5 Ongoing IG mutations No Yes No BCL2 rearrangement 0 20-25% 0 Rel amplification 0 15% SOCS1 inactivation 45% NFkB activation Yes No yes 5 year survival 15-30 50-60 65 Disease hallmarks Late relapses Pred. women <35 yrs , mediastinal
- 55. Plasma cell neoplasms • Plasma cell myeloma: • Conventional cytogenetics: abnormalities are detected in 1/3rd of myelomas • FISH detects the abnormalities in 90% of cases. • Numerical, structural abnormalities : trisomy, whole/ partial chromosomal deletion & translocation & complex abnormalities.
- 56. Plasma cell neoplasms • Most frequent translocation involve IGH@ on chromosome 14q32 • Five major recurrent oncogenes are involved in 14q32 translocation: (40%) • Cyclin D1 (11q13) • C-MAF(16q23) • FGFR3/MMSET(4p16.3) • Cyclin D3 (6p21) • MAFB(20q11) • Hyperdiploidy with gain in odd numbered chromosome 3, 5, 7, 9, 11, 15, ,19, 21 • Monosomy or partial deletion of chromosome 13q14
- 57. Translocation & Cyclin D groups in plasma cell myelomas Group Prim. Translocations Gene D -Cyclin Ploidy Ferquency % Prognosis 6q21 6q21 CCND3 D3 NH 3 Good 11q13 11q13 CCND1 D1 D, NH 16 Good D1 None None D1 H 34 Good D1+D2 None None D1+D2 H 6 ? poor D2 None None D2 H, NH 17 ? None None None None NH 2 ? Good 4p16 4p16 FGFR3/ MMSET D2 NH>H 15 poor maf 16q23 C- maf D2 NH 5 poor 20q11 mafB D2 NH 2 poor
- 58. Classical Hodgkin’s lymphoma • Conventional cytogenetics and FISH: – Overexpression of p53 – Aneuplody – Hypertetraploidy – Fail to demonstrate recurrent & specific chromosomal changes • Comparative genomic hybridization: – recurrent gain of chromosomal sub regions on chromosomal arms 2p, 9p, 12qand high level amplifications on chromosomal bands 4q16, 4q23-q24 & 9p23-p24
- 59. NLPHL • clonal reangement of IG gene • IGH@-BCL6 translocation • Somatic hypermutation in PAX5, also in PIM1,MYC & RhoH/TTF • EBV infection negative
- 60. Mature T- & NK cell neoplasms
- 61. • T-cell prolymphocytic leukemia: – Insersion of chromosome 14 with breakpoints in the long arm at q14 & q32(80%) – Reciprocal tandem translocation t(14;14)(q11;q32) – t(X;14)(q28;q11): less common – Abnormalities of chromosome 8, idic(8p11).t(8;8)(p11- p12;q12) & trisomy 8q(70-80%) – Deletion at12p13(FISH) – Deletion at 11q23, locus for the ATM gane (molecular &FISH studies) – Abnormalities of chromosome 6(33%) & 17(26%)
- 62. • Chronic lymphoproliferative disorders of NK cells: – Normal karyotype (most of cases) – No rearrangement of immunoglobulin & T cell receptor gene – In females: X- chromosome inactivation: indirect marker of clonality • Aggressive NK-cell leukemia: – TCR genes : germline configuration – In females: X- chromosome inactivation: indirect marker of clonality – Other clonal cytogenetic abnormalities: del(6)(q21q25) , del(11q) •
- 63. • Extranodal NK/T- cell lymphoma, nasal type • Various cytogenetic aberration but no specific chromosomal translocation – Del(6)(q21;q25) or i(6)(p10): commonest • Other aberration on comparative genomic hybridization: – Gain of 2q – Loss od 1p36.23-p36.33,6q16.1-q27, 4q12, 5q34- q35.3, 7q21.3-q22.1, 11q22.3-q27, 4q12, 5q34- q35.3, 7q21.3-q22.1, 11q22.3-q23.3 & 15q11.2-q14
- 64. • Hepatosplenic T- cell lymphoma – Isochromosome 7q (FISH ) – With disease progression 2 to 5 copies of i(7)(q10) – Ring chromosome leading to 7q amplification – Trisomy 8 – Loss of sex chromosome • Sezary syndrome – Recurrent chromosome abnormalities : not detected – Complex caryotypes with numerical & structural alteration are common – Unbalanced translocation & deletions of chromosome 1p, 6q, 10q, 17p, 19
- 65. • Angioimmunoblastic T-cell lymphoma: – Most frequent cytogenetic abnormalites: trisomy 3, trisomy 5, additional X chromosome – On comparative genetic hybridization : – Gain of 22q, 19 & 11q13& loss of 13q • T cell large granular lymphocytic leukemia: – No unique karyotypic abnormality but, Clonal, TCR gene rearrangement
- 66. Translocations & fusion proteins involving ALK at 2p23 Chromosomal anomaly ALK partner M Wt og ALK hybrid protein ALK staining pattern % of cases t(2;5)(p23;q35) NPM 80 N, diffuse Cyto. 84% t(1;2)(q25;p23) TPM3 104 diffuse Cyto. 13% Inv(2)(p23;q35) ATIC 96 diffuse Cyto. < 1% t(2;3)(p23;q21) TFG X long TFG long TFG strong 113 97 85 diffuse Cyto. diffuse Cyto. diffuse Cyto. < 1% t(2;17)(p23;q23) CTCL 250 granular cyto. < 1%
- 67. Translocations & fusion proteins involving ALK at 2p23 Chromosomal anomaly ALK partner M Wt og ALK hybrid protein ALK staining pattern % of cases t(2;X)(p23;q11-12) MSN 125 membrane 84% t(2;19)(p23;P13.1) TPM4 95 diffuse Cyto. < 1% t(2;22) p23;q11.2) MYH9 220 diffuse Cyto. < 1% t(2;17)(p23;q25) ALO17 ND diffuse Cyto. < 1% OTHERS ? ? N/ Cyto. < 1%
- 68. In situ hybridization • Epstein-Barr Virus infection : NHL & HL – EBER: EBV-encoded small nuclear RNA – LMP1 – LAMA • HHV 8 : – kaposy sarcoma – Primary effusion lymphoma – Large B cell lymphoma arising in HHV 8- associated multicentric Castleman disease • HTLV-1: – Adult T-cell leukemia/lymphoma
- 70. Acute lymphoblastic leukemia • B lymphoblastic leukemia • Clonal DJ rearrangement of IGH@ gene :100% • T cell receptor gene rearrangement : upto 70% • T lymphoblastic leukemia • T cell receptor gene rearrangement: 100% • rearrangement of IGH@ gene: 20% • so not helpful for lineage assignment
- 71. Acute lymphoblastic leukemia • t(9;22) (q34;q11.2)BCR-ABL1 • t(v;11q23) MLL rearranged • t(12;21)(p13;q22)(ETV6-RUNX1) / TEL-AML1 • t(5;14) IL-3-IGH • t(1;19) E2A-PBX1 / TCF3-PBX1
- 72. Acute myeloid leukemia • KIT mutation : t(8;21)(q22;q22), inv(16)(p13;q22), t(16;16)(p13.1;q22): poor prognosis • FLT3 mutation : FLT3-ITD, FLT3-TKD • MLL mutation : MLL -PTD • CEBPA • NPM1 • WT1 • BAALC • ERG • ERG • MN1
- 73. Myeloproliferative Neoplasms • BCR-ABL 1: RT-PCR, RQ-PCR • JAK-2 mutation – JAK -2 V617F , JAK -2 exon 12 mutation • MPL W515K/L mutation
- 74. • Mastocytosis • Point mutation within KIT protoncogene – D816V mutation : nested PCR • Systemic Mastocytosis with AML: – RUNX1-RUNX1T1 fusion • Systemic Mastocytosis with myeloproliferative disease: – JAK2 V617F mutation
- 75. • RT-PCR, using primers suitable for all known breakpoint is recommended to confirm ETV6- PDGFRB . Myeloid & lymphoid neoplasm with eosinophilia & abnormalitites of PDGFRA, PDGFRB & FGFR1
- 76. Lymphoma • NHL &HL (HRS cells) – Immunoglobulin (IG) heavy or light chain rearrangement • Mature T- & NK- cell neoplasms – TCR gene rearrangement
- 77. • Sezary syndrome: • Amplification of JUNB • Inactivation TP53 & p16 ink4a
- 78. • Langerhan’s cell histiocytosis: • Clonal X linked androgrn receptor gene assay • No consistent molecular genetic defect identified
- 79. Other investigations • Viral marker study • HIV • HBV • HCV • Bacterial infection • H. pylori infection
- 80. • Serum β2 microblobulin • Serum protein • Serum or urine protein electrophoresis • Serum calcium • Serum creatinine