J Pathol Transl Med.  2017 May;51(3):224-241. 10.4132/jptm.2017.04.09.

Molecular Testing of Lymphoproliferative Disorders: Current Status and Perspectives

• Jeon YK
• Yoon SO
• Paik JH
• Kim YA
• Shin BK
• Kim HJ
• Cha HJ
• Kim JE
• Huh J
• Ko YH
• The Hematopathology Study Group of the Korean Society of Pathologists
• The Molecular Pathology Study Group of Korean Society of Pathologists

• KMID: 2392591
• DOI: http://doi.org/10.4132/jptm.2017.04.09

Abstract

Molecular pathologic testing plays an important role for the diagnosis, prognostication and decision of treatment strategy in lymphoproliferative disease. Here, we briefly review the molecular tests currently used for lymphoproliferative disease and those which will be implicated in clinical practice in the near future. Specifically, this guideline addresses the clonality test for B- and T-cell proliferative lesions, molecular cytogenetic tests for malignant lymphoma, determination of cell-of-origin in diffuse large B-cell lymphoma, and molecular genetic alterations incorporated in the 2016 revision of the World Health Organization classification of lymphoid neoplasms. Finally, a new perspective on the next-generation sequencing for diagnostic, prognostic, and therapeutic purpose in malignant lymphoma will be summarized.

Keyword

Lymphoproliferative disorders; Malignant lymphoma; Pathology, molecular; Molecular diagnostics; Clonality test; Gene translocation; In situ hybridization, fluorescence; Next-generation sequencing

MeSH Terms

Classification
Cytogenetics
Diagnosis
In Situ Hybridization, Fluorescence
Lymphoma
Lymphoma, B-Cell
Lymphoproliferative Disorders*
Molecular Biology
Pathology, Molecular
T-Lymphocytes
World Health Organization

Figure

• 

  Fig. 1. Structure of IGH genes and the BIOMED-2 multiplex IGH gene polymerase chain reaction assay.

• 

  Fig. 2. Structure of TCRG genes and the BIOMED-2 multiplex TCRG gene polymerase chain reaction assay.

• 

  Fig. 3. Representative results and interpretation of BIOMED-2 multiplex IGH PCR analyzed by gene scanning. Clonal IGH gene rearrangement was detected in case (A), but not in case (B).

• 

  Fig. 4. Fluorescence in situ hybridization analysis to detect gene translocations using dual color, dual fusion probe (arrow, fused IGH and BCL2 genes) (A) and dual color, break apart probe (arrows, splitted MALT1 genes) (B). MALT, mucosa-associated lymphoid tissue.

• 

  Fig. 5. Representative immunohistochemical algorithms for the subgrouping of diffuse large B-cell lymphoma [42, 43]. GCB, germinal center B-cell.

• 

  Fig. 6. Genes involved in B-cell receptor signaling which converges to mitogen-activated protein (MAP) kinase pathway, nuclear factor κB (NF-κB) pathway, and phosphoinositide 3-kinase (PI3K) pathway are the therapeutic targets of B-cell lymphomas. MAPK, mitogen-activated protein kinase; MAPKK, mitogen-activated protein kinase kinase; MAPKKK, mitogen-activated protein kinase kinase kinase; mTOR, mammalian target of rapamycin. Modified from Young et al. Semin Hematol 2015;52:77-85, with permission of Elsevier [73].

• 

  Fig. 7. T-cell receptor signaling-related genes in nodal lymphomas of follicular helper T-cell phenotype are therapeutic targets. PI3K, phosphoinositide 3-kinase; NF-κB, nuclear factor κB; MAPK, mitogen-activated protein kinase. Modified from Vallois et al. Blood 2016;128:1490- 502, with permission of American Society of Hematology [84].

Reference

1. Tonegawa S. Somatic generation of antibody diversity. Nature. 1983; 302:575–81.

2. Davis MM, Bjorkman PJ. T-cell antigen receptor genes and T-cell recognition. Nature. 1988; 334:395–402.

3. van Dongen JJ, Langerak AW, BrÜggemann M, et al. Design and standardization of PCR primers and protocols for detection of clonal immunoglobulin and T-cell receptor gene recombinations in suspect lymphoproliferations: report of the BIOMED-2 Concerted Action BMH4-CT98-3936. Leukemia. 2003; 17:2257–317.

4. Gilfillan S, Dierich A, Lemeur M, Benoist C, Mathis D. Mice lacking TdT: mature animals with an immature lymphocyte repertoire. Science. 1993; 261:1175–8.

5. van Dongen JJ, Szczepanski T, Adriaansen HJ. Immunobiology of leukemia. In : Henderson ES, Lister TA, Greaves MF, editors. 7th ed. Leukemia. Philadelphia: WB Saunders Company;2002. p. 85–130.

6. Langerak AW, Groenen PJ, Bruggemann M, et al. EuroClonality/BIOMED-2 guidelines for interpretation and reporting of Ig/TCR clonality testing in suspected lymphoproliferations. Leukemia. 2012; 26:2159–71.

7. Sherwood AM, Desmarais C, Livingston RJ, et al. Deep sequencing of the human TCRγ and TCRβ repertoires suggests that TCRβ rearranges after αβ and γδ T cell commitment. Sci Transl Med. 2011; 3:90ra61.

8. Bailey NG, Elenitoba-Johnson KS. Molecular diagnostics of T-cell lymphoproliferative disorders. Cancer J. 2014; 20:48–60.

9. Schafernak KT, Variakojis D, Goolsby CL, et al. Clonality assessment of cutaneous B-cell lymphoid proliferations: a comparison of flow cytometry immunophenotyping, molecular studies, and immunohistochemistry/in situ hybridization and review of the literature. Am J Dermatopathol. 2014; 36:781–95.

10. FÖdinger M, Winkler K, Mannhalter C, Chott A. Combined polymerase chain reaction approach for clonality detection in lymphoid neoplasms. Diagn Mol Pathol. 1999; 8:80–91.

11. Thériault C, Galoin S, Valmary S, et al. PCR analysis of immunoglobulin heavy chain (IgH) and TcR-γ chain gene rearrangements in the diagnosis of lymphoproliferative disorders: results of a study of 525 cases. Mod Pathol. 2000; 13:1269–79.

12. Benhattar J, Delacretaz F, Martin P, Chaubert P, Costa J. Improved polymerase chain reaction detection of clonal T-cell lymphoid neoplasms. Diagn Mol Pathol. 1995; 4:108–12.

13. Signoretti S, Murphy M, Cangi MG, Puddu P, Kadin ME, Loda M. Detection of clonal T-cell receptor γ gene rearrangements in paraffin-embedded tissue by polymerase chain reaction and nonradioactive single-strand conformational polymorphism analysis. Am J Pathol. 1999; 154:67–75.

14. Patel KP, Pan Q, Wang Y, et al. Comparison of BIOMED-2 versus laboratory-developed polymerase chain reaction assays for detecting T-cell receptor-γ gene rearrangements. J Mol Diagn. 2010; 12:226–37.

15. van Krieken JH, Langerak AW, Macintyre EA, et al. Improved reliability of lymphoma diagnostics via PCR-based clonality testing: report of the BIOMED-2 Concerted Action BHM4-CT98-3936. Leukemia. 2007; 21:201–6.

16. Langerak AW, Molina TJ, Lavender FL, et al. Polymerase chain reaction-based clonality testing in tissue samples with reactive lymphoproliferations: usefulness and pitfalls. A report of the BIOMED-2 Concerted Action BMH4-CT98-3936. Leukemia. 2007; 21:222–9.

17. Brüggemann M, White H, Gaulard P, et al. Powerful strategy for polymerase chain reaction-based clonality assessment in T-cell malignancies Report of the BIOMED-2 Concerted Action BHM4 CT98-3936. Leukemia. 2007; 21:215–21.

18. Kuppers R, Klein U, Hansmann ML, Rajewsky K. Cellular origin of human B-cell lymphomas. N Engl J Med. 1999; 341:1520–9.

19. Davis TH, Yockey CE, Balk SP. Detection of clonal immunoglobulin gene rearrangements by polymerase chain reaction amplification and single-strand conformational polymorphism analysis. Am J Pathol. 1993; 142:1841–7.

20. Bourguin A, Tung R, Galili N, Sklar J. Rapid, nonradioactive detection of clonal T-cell receptor gene rearrangements in lymphoid neoplasms. Proc Natl Acad Sci U S A. 1990; 87:8536–40.

21. Medeiros LJ, Carr J. Overview of the role of molecular methods in the diagnosis of malignant lymphomas. Arch Pathol Lab Med. 1999; 123:1189–207.

22. Boeckx N, Willemse MJ, Szczepanski T, et al. Fusion gene transcripts and Ig/TCR gene rearrangements are complementary but infrequent targets for PCR-based detection of minimal residual disease in acute myeloid leukemia. Leukemia. 2002; 16:368–75.

23. Szczepanński T, Beishuizen A, Pongers-Willemse MJ, et al. Cross-lineage T cell receptor gene rearrangements occur in more than ninety percent of childhood precursor-B acute lymphoblastic leukemias: alternative PCR targets for detection of minimal residual disease. Leukemia. 1999; 13:196–205.

24. Szczepanński T, Langerak AW, van Dongen JJ, van Krieken JH. Lymphoma with multi-gene rearrangement on the level of immunoglobulin heavy chain, light chain, and T-cell receptor β chain. Am J Hematol. 1998; 59:99–100.

25. Moreau EJ, Langerak AW, van Gastel-Mol EJ, et al. Easy detection of all T cell receptor gamma (TCRG) gene rearrangements by Southern blot analysis: recommendations for optimal results. Leukemia. 1999; 13:1620–6.

26. Langerak AW, Wolvers-Tettero IL, van Dongen JJ. Detection of T cell receptor beta (TCRB) gene rearrangement patterns in T cell malignancies by Southern blot analysis. Leukemia. 1999; 13:965–74.

27. Kluin P, Schuuring E. Molecular cytogenetics of lymphoma: where do we stand in 2010? Histopathology. 2011; 58:128–44.

28. Niitsu N, Okamoto M, Miura I, Hirano M. Clinical features and prognosis of de novo diffuse large B-cell lymphoma with t(14;18) and 8q24/c-MYC translocations. Leukemia. 2009; 23:777–83.

29. Johnson NA, Slack GW, Savage KJ, et al. Concurrent expression of MYC and BCL2 in diffuse large B-cell lymphoma treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone. J Clin Oncol. 2012; 30:3452–9.

30. Horn H, Ziepert M, Becher C, et al. MYC status in concert with BCL2 and BCL6 expression predicts outcome in diffuse large B-cell lymphoma. Blood. 2013; 121:2253–63.

31. Hu S, Xu-Monette ZY, Tzankov A, et al. MYC/BCL2 protein coexpression contributes to the inferior survival of activated B-cell subtype of diffuse large B-cell lymphoma and demonstrates high-risk gene expression signatures: a report from The International DLBCL Rituximab-CHOP Consortium Program. Blood. 2013; 121:4021–31.

32. Swerdlow SH, Campo E, Pileri SA, et al. The 2016 revision of the World Health Organization classification of lymphoid neoplasms. Blood. 2016; 127:2375–90.

33. Swerdlow SH, Campo E, Harris NL, et al. WHO classification of tumours of haematopoietic and lymphoid tissues. 4th ed. Lyon: IARC Press;2008.

34. Green TM, Young KH, Visco C, et al. Immunohistochemical double-hit score is a strong predictor of outcome in patients with diffuse large B-cell lymphoma treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone. J Clin Oncol. 2012; 30:3460–7.

35. Campbell LJ. Cancer cytogenetics: methods and protocols. 2nd ed. New York: Humana Press;2011.

36. Wolff DJ, Bagg A, Cooley LD, et al. Guidance for fluorescence in situ hybridization testing in hematologic disorders. J Mol Diagn. 2007; 9:134–43.

37. McGowan-Jordan J, Simons A, Schmid M. ISCN 2016: an international system for human cytogenomic nomenclature (2016). Basel: Karger;2016.

38. Alizadeh AA, Eisen MB, Davis RE, et al. Distinct types of diffuse large B-cell lymphoma identified by gene expression profiling. Nature. 2000; 403:503–11.

39. Wright G, Tan B, Rosenwald A, Hurt EH, Wiestner A, Staudt LM. A gene expression-based method to diagnose clinically distinct subgroups of diffuse large B cell lymphoma. Proc Natl Acad Sci U S A. 2003; 100:9991–6.

40. Testoni M, Zucca E, Young KH, Bertoni F. Genetic lesions in diffuse large B-cell lymphomas. Ann Oncol. 2015; 26:1069–80.

41. Carbone A, Gloghini A, Kwong YL, Younes A. Diffuse large B cell lymphoma: using pathologic and molecular biomarkers to define subgroups for novel therapy. Ann Hematol. 2014; 93:1263–77.

42. Hans CP, Weisenburger DD, Greiner TC, et al. Confirmation of the molecular classification of diffuse large B-cell lymphoma by immunohistochemistry using a tissue microarray. Blood. 2004; 103:275–82.

43. Choi WW, Weisenburger DD, Greiner TC, et al. A new immunostain algorithm classifies diffuse large B-cell lymphoma into molecular subtypes with high accuracy. Clin Cancer Res. 2009; 15:5494–502.

44. Meyer PN, Fu K, Greiner TC, et al. Immunohistochemical methods for predicting cell of origin and survival in patients with diffuse large B-cell lymphoma treated with rituximab. J Clin Oncol. 2011; 29:200–7.

45. Gutiérrez-García G, Cardesa-Salzmann T, Climent F, et al. Gene-expression profiling and not immunophenotypic algorithms predicts prognosis in patients with diffuse large B-cell lymphoma treated with immunochemotherapy. Blood. 2011; 117:4836–43.

46. Intlekofer AM, Younes A. Precision therapy for lymphoma: current state and future directions. Nat Rev Clin Oncol. 2014; 11:585–96.

47. Scott DW, Wright GW, Williams PM, et al. Determining cell-of-origin subtypes of diffuse large B-cell lymphoma using gene expression in formalin-fixed paraffin-embedded tissue. Blood. 2014; 123:1214–7.

48. Scott DW, Mottok A, Ennishi D, et al. Prognostic significance of diffuse large B-cell lymphoma cell of origin determined by digital gene expression in formalin-fixed paraffin-embedded tissue biopsies. J Clin Oncol. 2015; 33:2848–56.

49. Dietrich S, Pircher A, Endris V, et al. BRAF inhibition in hairy cell leukemia with low-dose vemurafenib. Blood. 2016; 127:2847–55.

50. Raess PW, Mintzer D, Husson M, et al. BRAF V600E is also seen in unclassifiable splenic B-cell lymphoma/leukemia, a potential mimic of hairy cell leukemia. Blood. 2013; 122:3084–5.

51. Blombery PA, Wong SQ, Hewitt CA, et al. Detection of BRAF mutations in patients with hairy cell leukemia and related lymphoproliferative disorders. Haematologica. 2012; 97:780–3.

52. Arcaini L, Zibellini S, Boveri E, et al. The BRAF V600E mutation in hairy cell leukemia and other mature B-cell neoplasms. Blood. 2012; 119:188–91.

53. Mason EF, Brown RD, Szeto DP, et al. Detection of activating MAP2K1 mutations in atypical hairy cell leukemia and hairy cell leukemia variant. Leuk Lymphoma. 2017; 58:233–6.

54. Waterfall JJ, Arons E, Walker RL, et al. High prevalence of MAP2K1 mutations in variant and IGHV4-34-expressing hairy-cell leukemias. Nat Genet. 2014; 46:8–10.

55. Kapoor P, Ansell SM, Fonseca R, et al. Diagnosis and management of waldenstrom macroglobulinemia: Mayo stratification of macroglobulinemia and risk-adapted therapy (mSMART) guidelines 2016. JAMA Oncol 2017 Jan 5 [Epub]. https://doi.org/10.1001/jamaoncol.2016.5763.

56. Schmidt J, Federmann B, Schindler N, et al. MYD88 L265P and CXCR4 mutations in lymphoplasmacytic lymphoma identify cases with high disease activity. Br J Haematol. 2015; 169:795–803.

57. Martinez-Lopez A, Curiel-Olmo S, Mollejo M, et al. MYD88 (L265P) somatic mutation in marginal zone B-cell lymphoma. Am J Surg Pathol. 2015; 39:644–51.

58. Hamadeh F, MacNamara SP, Aguilera NS, Swerdlow SH, Cook JR. MYD88 L265P mutation analysis helps define nodal lymphoplasmacytic lymphoma. Mod Pathol. 2015; 28:564–74.

59. Salaverria I, Philipp C, Oschlies I, et al. Translocations activating IRF4 identify a subtype of germinal center-derived B-cell lymphoma affecting predominantly children and young adults. Blood. 2011; 118:139–47.

60. Salaverria I, Martin-Guerrero I, Wagener R, et al. A recurrent 11q aberration pattern characterizes a subset of MYC-negative high-grade B-cell lymphomas resembling Burkitt lymphoma. Blood. 2014; 123:1187–98.

61. Ferreiro JF, Morscio J, Dierickx D, et al. Post-transplant molecularly defined Burkitt lymphomas are frequently MYC-negative and characterized by the 11q-gain/loss pattern. Haematologica. 2015; 100:e275–9.

62. Stilgenbauer S, Schnaiter A, Paschka P, et al. Gene mutations and treatment outcome in chronic lymphocytic leukemia: results from the CLL8 trial. Blood. 2014; 123:3247–54.

63. Nadeu F, Delgado J, Royo C, et al. Clinical impact of clonal and subclonal TP53, SF3B1, BIRC3, NOTCH1, and ATM mutations in chronic lymphocytic leukemia. Blood. 2016; 127:2122–30.

64. te Raa GD, Malcikova J, Pospisilova S, et al. Overview of available p53 function tests in relation to TP53 and ATM gene alterations and chemoresistance in chronic lymphocytic leukemia. Leuk Lymphoma. 2013; 54:1849–53.

65. Rossi D, Fangazio M, Rasi S, et al. Disruption of BIRC3 associates with fludarabine chemorefractoriness in TP53 wild-type chronic lymphocytic leukemia. Blood. 2012; 119:2854–62.

66. Beà S, Valdés-Mas R, Navarro A, et al. Landscape of somatic mutations and clonal evolution in mantle cell lymphoma. Proc Natl Acad Sci U S A. 2013; 110:18250–5.

67. Xu-Monette ZY, Wu L, Visco C, et al. Mutational profile and prognostic significance of TP53 in diffuse large B-cell lymphoma patients treated with R-CHOP: report from an International DLBCL Rituximab-CHOP Consortium Program Study. Blood. 2012; 120:3986–96.

68. Cao Y, Zhu T, Zhang P, et al. Mutations or copy number losses of CD58 and TP53 genes in diffuse large B cell lymphoma are independent unfavorable prognostic factors. Oncotarget. 2016; 7:83294–307.

69. Jardin F, Pujals A, Pelletier L, et al. Recurrent mutations of the exportin 1 gene (XPO1) and their impact on selective inhibitor of nuclear export compounds sensitivity in primary mediastinal B-cell lymphoma. Am J Hematol. 2016; 91:923–30.

70. Camus V, Stamatoullas A, Mareschal S, et al. Detection and prognostic value of recurrent exportin 1 mutations in tumor and cell-free circulating DNA of patients with classical Hodgkin lymphoma. Haematologica. 2016; 101:1094–101.

71. Nakajima H, Kunimoto H. TET2 as an epigenetic master regulator for normal and malignant hematopoiesis. Cancer Sci. 2014; 105:1093–9.

72. Lemonnier F, Couronné L, Parrens M, et al. Recurrent TET2 mutations in peripheral T-cell lymphomas correlate with TFH-like features and adverse clinical parameters. Blood. 2012; 120:1466–9.

73. Young RM, Shaffer AL 3rd, Phelan JD, Staudt LM. B-cell receptor signaling in diffuse large B-cell lymphoma. Semin Hematol. 2015; 52:77–85.

74. Okosun J, Bödör C, Wang J, et al. Integrated genomic analysis identifies recurrent mutations and evolution patterns driving the initiation and progression of follicular lymphoma. Nat Genet. 2014; 46:176–81.

75. Bouska A, Zhang W, Gong Q, et al. Combined copy number and mutation analysis identifies oncogenic pathways associated with transformation of follicular lymphoma. Leukemia. 2017; 31:83–91.

76. Younes A. Promising novel agents for aggressive B-cell lymphoma. Hematol Oncol Clin North Am. 2016; 30:1229–37.

77. Kurmasheva RT, Sammons M, Favours E, et al. Initial testing (stage 1) of tazemetostat (EPZ-6438), a novel EZH2 inhibitor, by the pediatric preclinical testing program. Pediatr Blood Cancer. 2017; 64:e26218.

78. Parrilla Castellar ER, Jaffe ES, Said JW, et al. ALK-negative anaplastic large cell lymphoma is a genetically heterogeneous disease with widely disparate clinical outcomes. Blood. 2014; 124:1473–80.

79. King RL, Dao LN, McPhail ED, et al. Morphologic features of ALK-negative anaplastic large cell lymphomas with DUSP22 rearrangements. Am J Surg Pathol. 2016; 40:36–43.

80. Manso R, Sánchez-Beato M, Monsalvo S, et al. The RHOA G17V gene mutation occurs frequently in peripheral T-cell lymphoma and is associated with a characteristic molecular signature. Blood. 2014; 123:2893–4.

81. Yoo HY, Sung MK, Lee SH, et al. A recurrent inactivating mutation in RHOA GTPase in angioimmunoblastic T cell lymphoma. Nat Genet. 2014; 46:371–5.

82. Sakata-Yanagimoto M, Enami T, Yoshida K, et al. Somatic RHOA mutation in angioimmunoblastic T cell lymphoma. Nat Genet. 2014; 46:171–5.

83. Palomero T, Couronné L, Khiabanian H, et al. Recurrent mutations in epigenetic regulators, RHOA and FYN kinase in peripheral T cell lymphomas. Nat Genet. 2014; 46:166–70.

84. Vallois D, Dobay MP, Morin RD, et al. Activating mutations in genes related to TCR signaling in angioimmunoblastic and other follicular helper T-cell-derived lymphomas. Blood. 2016; 128:1490502.

85. Rohr J, Guo S, Huo J, et al. Recurrent activating mutations of CD28 in peripheral T-cell lymphomas. Leukemia. 2016; 30:1062–70.

86. Streubel B, Vinatzer U, Willheim M, Raderer M, Chott A. Novel t(5;9)(q33;q22) fuses ITK to SYK in unspecified peripheral T-cell lymphoma. Leukemia. 2006; 20:313–8.

87. Pechloff K, Holch J, Ferch U, et al. The fusion kinase ITK-SYK mimics a T cell receptor signal and drives oncogenesis in conditional mouse models of peripheral T cell lymphoma. J Exp Med. 2010; 207:1031–44.

88. Wang C, McKeithan TW, Gong Q, et al. IDH2R172 mutations define a unique subgroup of patients with angioimmunoblastic T-cell lymphoma. Blood. 2015; 126:1741–52.

89. Cairns RA, Iqbal J, Lemonnier F, et al. IDH2 mutations are frequent in angioimmunoblastic T-cell lymphoma. Blood. 2012; 119:1901–3.

90. Jiang L, Gu ZH, Yan ZX, et al. Exome sequencing identifies somatic mutations of DDX3X in natural killer/T-cell lymphoma. Nat Genet. 2015; 47:1061–6.

91. Lee S, Park HY, Kang SY, et al. Genetic alterations of JAK/STAT cascade and histone modification in extranodal NK/T-cell lymphoma nasal type. Oncotarget. 2015; 6:17764–76.

92. Dobashi A, Tsuyama N, Asaka R, et al. Frequent BCOR aberrations in extranodal NK/T-cell lymphoma, nasal type. Genes Chromosomes Cancer. 2016; 55:460–71.