The Challenges of Precision Medicine and New Advances in Molecular Diagnostic Testing in Hematolymphoid Malignancies: Impact on the VHA

A group of T-cell lymphoproliferative disorders with expression of T follicular helper cell markers can be identified with IHC. These disorders include angioimmunoblastic TCL; follicular TCL, a new entity that is a PTCL-NOS subset; and primary cutaneous CD4-positive small/medium T-cell lymphoproliferative disorder. The neoplastic cells should express at least 2 or 3 T follicular helper cell–related antigens, including CD279/PD1, CD10, BCL6, CXCL13, ICOS, SAP, and CCR5; the most commonly used are PD1, BCL6, and CD10. Recurrent fusion of ITK-SYK translocation t(5;9) or CTLA4-CD28 is also common in follicular TCL. Although recurrent mutation is found in these entities, conventional karyotyping or IHC should be sufficient for diagnosis.

Cutaneous γ-Δ T-Cell Lymphoma

Among cutaneous TCLs, primary cutaneous γ-Δ TCL is clinically aggressive (median survival, 15 months). By definition, the cells are of the TCR-γ-Δ phenotype. When available, this phenotype is best shown with IHC staining for TCR-γ or TCR-Δ with appropriate detection methods. In routine practice, however, absence of β-F1 expression is sufficient to infer the TCR-γ-Δ phenotype.⁴⁸

Peripheral T-Cell Lymphoma

Gene expression profiling analysis of PTCLs has identified at least 3 subtypes characterized by overexpression of GATA3, TBX21, and cytotoxic genes and expression of the corresponding proteins with IHC.⁴⁷ These subtypes are associated with different clinical behavior and therapy responses. The GATA3 subtype has an inferior prognosis and shows a high level of T helper type 2 cytokines, which can be identified with IHC. As IHC-stained GATA3 has been available as a marker of urothelial carcinoma at most IHC laboratories, GATA3 IHC staining also may be considered in the evaluation of PTCLs.

Many monoclonal antibody therapies are being used as primary or secondary regimens in the treatment of TCL. Clinical trials are working to establish their efficacy. If treatment with a monoclonal antibody is being considered, it is appropriate to conduct IHC to demonstrate the presence of the target antigen and at follow-up, to demonstrate the efficacy of treatment. These therapies include alemtuzumab, which targets CD52, and brentuximab, which targets CD30.

T-Cell Large Granular Lymphocytic Leukemia

T-cell LGL leukemia is a complex diagnosis that requires persistent clonal expansion of LGLs and clinically peripheral blood cytopenia. In many cases, the diagnosis is difficult to establish, as benign large granular lymphocytosis with clonal T cells may occur in conjunction with viral infections or autoimmune disorders. Somatic mutations in the STAT3 (signal transducer and activator of transcription 3) gene are found in 40% of patients with T-cell LGL leukemia.⁴⁹ More recently, somatic mutations in the STAT5B gene were identified in 2% of T-cell LGL leukemia subsets. The clinical course of T-cell LGL leukemia in patients with the STAT5B mutation is aggressive and fatal, clearly different from the relatively favorable course of typical T-cell LGL leukemia.⁵⁰ The HMG subcommittee recommends considering a STAT3 and STAT5B mutation study for selected cases in which it is difficult to distinguish true T-cell LGL leukemia from its reactive expansions.

T-Cell Prolymphocytic Leukemia

T-cell prolymphocytic leukemia (T-PLL) is a rare, aggressive disease and is most commonly associated with a prolymphocytic morphology and expression of CD4. However, since a specific immunophenotypic profile of T-PLL has not been identified, flow cytometry is not adequate in isolation for definitive classification as T-PLL.⁵¹ A diagnosis of T-PLL often requires cytogenetics or a FISH study to confirm a suspected case. Most TPLL cases harbor characteristic chromosomal abnormalities involving 14q11.2 (TCR α/Δ), 14q32 (TCL1 gene), or Xq28 (MTCP1 gene); abnormalities of chromosomes 8 and 12p; and deletions of the long arm of chromosomes 5, 6, 11, and 13.⁵² In routine practice, a TPLL diagnosis should be confirmed with inv(14) (paracentric inversion of chromosome 14) or t(14;14) by conventional cytogenetic studies and/or rearrangement of the TCL1 gene by FISH. In addition, ATM and JAK3 mutations are highly recurrent in TPLL and may aid in the diagnosis of challenging cases.⁵³

Anaplastic Large Cell Lymphoma

The World Health Organization recognizes 3 distinct types of anaplastic large cell lymphoma (ALCL): systemic anaplastic lymphoma kinase (ALK)–positive ALCL, systemic ALK-negative ALCL, and primary cutaneous ALCL. Systemic ALK-positive ALCLs consistently have ALK gene rearrangements and favorable outcomes. The most common translocation is the t(2;5) rearrangement of NPM1 and ALK, though other ALK partners are also possible. In contrast, systemic ALK-negative ALCLs lack ALK gene rearrangements and as a whole have outcomes inferior to those of systemic ALK-positive ALCLs. However, studies have found systemic ALK-negative ALCL to be a genetically and clinically heterogeneous entity.⁵⁴ About 30% of cases have rearrangements of the DUSP22-IRF4 locus on 6p25.3 (DUSP22 rearrangement), and these cases have favorable outcomes similar to those of systemic ALK-positive ALCL.⁵⁵ Only 8% of patients have TP63 rearrangements and very poor outcomes. The remaining cases lack ALK, DUSP22, and TP63 rearrangements and have intermediate outcomes. The HMG subcommittee recommends considering DUSP22 rearrangement by FISH in the evaluation of systemic ALK-negative ALCL.

Conclusion

The pathologic diagnosis, classification, and risk stratification of lymphoma and leukemia require an approach that integrates morphology, flow cytometry, cytogenetics, and molecular pathology. Rapidly evolving molecular techniques currently allow for detailed description of the molecular defects in lymphoma and leukemia, including driver mutations, amplification/deletion events, and clonal evolution. Unfortunately, the technical ability to catalogue the molecular defects in lymphoma and leukemia, often at great expense, is outpacing the ability to use this detailed information in treating patients with hematologic malignancies. The challenge, then, is to identify best practices for the diagnosis and classification of lymphoma and leukemia in VHA hospitals that incorporate the most useful molecular tests without wasting financial resources.

In this report, the HMG subcommittee of the MGPW has presented its recommendations for molecular testing in AML, MPN, MDS, and lymphomas in the context of standard morphologic and immunophenotypic approaches to hematopathology diagnosis and classification. Adoption of these recommendations by VHA hospitals and clinics should help ensure that all VA patients with hematologic malignancies benefit from the latest advances in precision medicine.

Within the vast and comprehensive national VHA health care system are multiple centers of expertise in hematopathology. In addition, multiple VA clinical molecular diagnostic laboratories are performing state-of-the-art testing. The HMG subcommittee proposes that, to make best use of these expert resources, the VHA should establish an interfacility hematopathology consultation service. This service would allow any VA pathologist to consult a board-certified hematopathologist regarding use of ancillary molecular genetic testing in the diagnosis of hematologic malignancy.

In addition, the HMG subcommittee recommends consolidating VA molecular diagnostic reference laboratories and having them perform molecular testing for other VA hospitals rather than using commercial reference laboratories, where testing standards are not uniform and results may be difficult to interpret. Several well-established VA clinical laboratories with technical expertise and informatics support are already performing selected molecular diagnostic testing. These laboratories’ resources should be expanded, where practical, to cost-effectively provide VA expertise to all veterans and to improve access to appropriate molecular diagnostic testing.

Author disclosures
The authors report no actual or potential conflicts of interest with regard to this article.

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