T-cell Receptor Gene Rearrangement Research Articles

Clinical and Pathological Features of Hydroa Vacciniforme-Like Lymphoproliferative Disorder Along with Risk Factors Indicating Poor Prognosis.

To examine the clinical and pathological features, laboratory markers, therapeutic options and risk factors indicating poor prognosis of hydroa vacciniforme-like lymphoproliferative disorder (HVLPD). Seven patients with HVLPD had their clinical and pathological data collected. Immunohistochemical staining, Epstein-Barr virus-encoded RNA (EBER) in situ hybridization experiments, T-cell receptor (TCR) gene rearrangement, RT-PCR tests and the Elisa assay were carried out. The main clinical manifestations were papulovesicular lesions and ulcers on the face, neck, or trunk. Five cases had systemic symptoms. Three of the deceased patients had significant facial edema, deep body necrosis, and ulceration. The pathological results demonstrated that lymphocytes infiltrated blood vessels and sweat glands in addition to the dermis and subcutaneous tissues. All patients tested positive for CD3 and EBER. Six cases tested positive for TCRβF1, but none tested positive for TCRδ. TCRγ monoclonal rearrangement, strongly positive expression of TIA-1 and a Ki67 proliferation index of 40% occurred in 3 fatal cases. When compared to the survival group, the plasma EBV DNA in the deceased group was considerably higher (P<0.05). IFN-γ and TNF-α cytokine levels in patients were higher than in the control group, particularly in the deceased group (P<0.05). The skin lesions on all patients recovered quickly underwent conservative care. Nonetheless, 3 patients passed away as the disease progressed in its latter stages. In our cases, the main infiltrating cells were T cells and the dominant lymphocyte subclass was αβT cells. A significant increase in lgE level, plasma EBV DNA, IFN-γ, and TNF-α cytokine levels, decreased hemoglobin level, strongly positive expression of TIA-1, high Ki67 proliferation index, and positive TCR gene rearrangement are all indicators of a poor prognosis.

Primary Cutaneous Multifocal Indolent CD8+ T-Cell Lymphoma: A Novel Primary Cutaneous CD8+ T-Cell Lymphoma

We report the case of a patient who was referred to our institution with a diagnosis of CD4+ small/medium-sized pleomorphic lymphoma. At the time, the patient showed a plethora of lesions mainly localizing to the legs; thus, we undertook studies to investigate the lineage and immunophenotype of the neoplastic clone. Immunohistochemistry (IHC) showed marked CD4 and CD8 positivity. Flow cytometry (FCM) showed two distinct T-cell populations, CD4+ and CD8+ (+/- PD1), with no CD4/CD8 co-expression and no loss of panT-cell markers in either T-cell subset. FCM, accompanied by cell-sorting (CS), permitted the physical separation of four populations, as follows: CD4+/PD1-, CD4+/PD1+, CD8+/PD1- and CD8+/PD1+. TCR gene rearrangement studies on each of the four populations (by next generation sequencing, NGS) showed that the neoplastic population was of T-cytotoxic cell lineage. IHC showed the CD8+ population to be TIA-1+, but perforin- and granzyme-negative. Moreover, histiocytic markers did not render the peculiar staining pattern, which is characteristic of acral CD8+ T-cell lymphoma (PCACD8). Compared to the entities described in the 2018 update of the WHO-EORTC classification for primary cutaneous lymphomas, we found that the indolent lymphoma described herein differed from all of them. We submit that this case represents a hitherto-undescribed type of CTCL.

Diagnostic spectrum of hypereosinophilia based on bone marrow pathology: 10 years' experience at a tertiary care hospital.

Hypereosinophilia (HE) is defined as peripheral blood (PB) eosinophil count exceeding 1.5 × 109 /L. As the causes of HE can be diverse, the work-up of patients was complicated. In this study, we aimed to categorize the underlying diseases associated with HE and demonstrate minimum diagnostic approach. Cases presenting with HE within 7 days of bone marrow (BM) examination conducted between 2008 and 2019 were selected. Cases were classified by the revised 2022 WHO and ICC classification. We also assessed morphologic features of unclassified persisting HE (>4 weeks) patients according to the morphologic criteria suggested a previous study by Wang et al. RESULTS: A total of 364 patients were included. The work-up confirmed primary HE in 38.7%, secondary HE in 48.9%, HE patients with insufficient evaluation in 13.7%. When conducted a slide review of HE patients with sustained HE more than 4 weeks among HE patients with insufficient evaluation, the morphological features showed abnormal eosinophils in PB/BM (69.0%/81.0%), hypercellularity (26.2%), myelofibrosis (7.1%), increased M:E ratio (5.3%), and dysmegakaryopoiesis (4.8%). Of these patients, 14 patients who met all morphologic criteria were suspected of CEL. This study demonstrates that HE is associated with variable conditions. BM morphological assessment based on a robust criterion can help to confirm a MN irrespective of the presence of clonal markers. The work-up of patients in whom ruled out the common secondary causes of HE requires a systematic but sufficient approach including at a minimum BM karyotyping, PDGFRA testing, lymphocyte immunophenotyping and TCR gene rearrangement.

Read the clonotype: Next-generation sequencing-based lymphocyte clonality analysis and perspectives for application in pathology.

Clonality assessment using the unique rearrangements of immunoglobulin (IG) and T-cell receptor (TR) genes in lymphocytes is a widely applied supplementary test for the diagnosis of B-cell and T-cell lymphoma. To enable a more sensitive detection and a more precise comparison of clones compared with conventional clonality analysis based on fragment analysis, the EuroClonality NGS Working Group developed and validated a next-generation sequencing (NGS)-based clonality assay for detection of the IG heavy and kappa light chain and TR gene rearrangements for formalin-fixed and paraffin-embedded tissues. We outline the features and advantages of NGS-based clonality detection and discuss potential applications for NGS-based clonality testing in pathology, including site specific lymphoproliferations, immunodeficiency and autoimmune disease and primary and relapsed lymphomas. Also, we briefly discuss the role of T-cell repertoire of reactive lymphocytic infiltrations in solid tumors and B-lymphoma.

Prevalence of STAT3 mutations in patients with rheumatoid arthritis-associated T-cell large granular lymphocytic leukaemia and Felty syndrome.

Neutropenia is a key presentation of Felty syndrome (FS) and rheumatoid arthritis (RA)-associated T-cell large granular lymphocytic (T-LGL) leukaemia. Clonal rearrangement of T-cell receptor (TCR) gene supports the diagnosis of T-LGL leukaemia but not FS. Mutations in the signal transducer and activator of transcription 3 (STAT3) gene are highly specific for T-LGL leukaemia, but their prevalence in FS remains poorly clarified. The study included 100 patients with RA and unexplained neutropenia. TCR rearrangements were examined in blood (100 cases), bone marrow (47 cases), and spleen (12 cases) using the BIOMED-2 protocol. Patients were stratified into RA-associated T-LGL leukaemia cohort if a clonal TCR rearrangement was identified in any of the tested patient samples, and into FS cohort in other cases. Mutations in the STAT3 were examined using next-generation sequencing (NGS) technology in blood (100 cases), bone marrow (37 cases), and spleen (7 cases). STAT3 mutations were identified in 71% (49/69) patients with RA-associated T-LGL leukaemia and in 10% (3/31) patients with FS (p=4.7×10-8). Three samples from the RA-associated T-LGL leukaemia cohort and 5 samples from the FS cohort had STAT3 mutations in the absence of clonal TCR rearrangement. The results suggest that STAT3 mutations are significantly less common in FS than in RA-associated T-LGL leukaemia. Moreover, NGS can detect clones undetectable by fragment analysis. We speculate that in patients with RA and neutropenia, the detection of STAT3 mutations can point to T-LGL leukaemia even in the absence of clonal TCR rearrangement.

Primary central nervous system T-cell lymphoma in children and adolescents: a clinicopathological analysis of five cases

Objective: To study the clinicopathological characteristics, and further understand primary central nervous system T-cell lymphoma (PCNSTCL) in children and adolescents. Methods: Five cases of PCNSTCL in children and adolescents were collected from December 2016 to December 2021 at the First Affiliated Hospital of Zhengzhou University. The clinicopathological characteristics, immunophenotypic, and molecular pathologic features were analyzed, and relevant literatures reviewed. Results: There were two male and three female patients with a median age of 14 years (range 11 to 18 years). There were two peripheral T-cell lymphomas, not otherwise specified, two anaplastic large cell lymphoma, ALK-positive and one NK/T cell lymphoma. Pathologically, the tumor cells showed a variable histomorphologic spectrum, including small, medium and large cells with diffuse growth pattern and perivascular accentuation. Immunohistochemistry and in situ hybridization showed CD3 expression in four cases, and CD3 was lost in one case. CD5 expression was lost in four cases and retained in one case. ALK and CD30 were expressed in two cases. One tumor expressed CD56 and Epstein-Barr virus-encoded RNA. All cases showed a cytotoxic phenotype with expression of TIA1 and granzyme B. Three cases had a high Ki-67 index (>50%). T-cell receptor (TCR) gene rearrangement was clonal in two cases. Conclusions: PCNSTCL is rare, especially in children and adolescents. The morphology of PCNSTCL is diverse. Immunohistochemistry and TCR gene rearrangement play important roles in the diagnosis.

Monitoring T-Cell Kinetics in the Early Recovery Period of Lung Transplantation Cases by Copy Number Levels of T-Cell Receptor Excision Circle.

Lung transplantation is a life-saving procedure for patients with end-stage lung diseases. T-Cell receptor excision circle (TREC) is circular DNA produced during T-cell receptor gene rearrangement in the thymus and indicates naive T-cell migration from the thymus. Therefore, its levels represent thymic T-cell output. Post-transplant lymphocyte kinetics correlate with graft tolerance. The aim of this study was to investigate T-lymphocyte kinetics in the early recovery period after lung transplantation. For this purpose, copy numbers of TREC were determined in patients with a lung transplant. In addition, TREC copy numbers were evaluated according to age, diagnosis and the forced expiratory volume in 1 second (FEV1) of lung transplant patients. Peripheral blood samples were taken from patients aged 23 to 59 years who underwent lung transplantation at the Thoracic Surgery Clinic, Kartal-Koşuyolu High Specialization Educational and Research Hospital. This study included peripheral blood samples from 11 lung transplant patients (comprising four with chronic obstructive pulmonary disease, three with idiopathic pulmonary fibrosis, one with cystic fibrosis, one with silicosis and two with bronchiectasis; three females in total). Samples were taken at three different timepoints: Before transplant, and 24 hours and 7 days post transplant. TREC copy numbers were analyzed with real time reverse transcriptase-polymerase chain reaction. Post-transplant TREC numbers and density values were higher compared to pre-transplant values, although these differences were statistically insignificant. TREC copy numbers were found to be significantly higher in patients younger than 45 years compared to patients older than 45 years. At 24 hours after the transplant, the average TREC copy number/peripheral blood mononuclear cells of the cases with an FEV1 value of or below 50% was found to be statistically significantly higher than that of cases with an FEV1 value above 50% (p=0.046). There was no statistically significant difference in TREC copy numbers between male and female patients or by diagnostic group. TREC copy numbers can be evaluated as a prognostic marker for lung transplantation. There is a need for multicenter studies with more patients.

Primary spinal extranodal NK/T-cell lymphoma diagnosed by fine-needle aspiration and core biopsy.

We report an extremely rare case of primary spinal extranodal NK/T-cell lymphoma (PSENKTL). A 54-year-old man presented with fever of unknown etiology for 2 months, with clinical evidence of hemophagocytic lymphohistiocytosis. Imaging studies revealed multiple spinal lesions without evidence of disease in other body sites. Fine-needle aspiration and core biopsy (FNACB) of a paraspinal mass showed a monotonous population of intermediate lymphocytes with abundant cytoplasm with fine granules, round to slightly irregular nuclei, and inconspicuous nucleoli. Core biopsy revealed diffuse infiltration by cells with a NK cell phenotype, positive Epstein-Barr virus-encoded small RNA, and negative T-cell receptor gene rearrangement. Bone marrow biopsy showed the presence of hemophagocytosis without evidence of lymphoma. The disease disseminated to the small bowel late in the clinical course and the patient died shortly after admission. This unusual case was diagnosed by FNACB and raised awareness of ENKTL as a differential diagnosis in spinal lesions.

Staging lymph nodes and blood at diagnosis in mycosis fungoides identifies patients at increased risk of progression to advanced stage: A retrospective cohort study.

Risk factors for progression to advanced-stage mycosis fungoides (MF) are poorly defined. The authors performed a single-center, retrospective cohort study among patients with MF at an academic medical center from 1990 to 2020 to identify clinical variables associated with progression to advanced-stage MF (stage IIB-IVB), and 388 patients who had a clinicopathologic diagnosis of early stage (IA-IIA) MF were identified from their cutaneous lymphoma database. Baseline clinical characteristics, laboratory values, imaging, and blood flow cytometry or T-cell receptor gene rearrangement (TCR) data were collected. Logistic regression was used to assess risk factors associated with progression. Overall, 93 of 388 patients (24.0%) progressed to advanced stage. Patients who progressed had an increased risk of death (hazard ratio, 4.50; 95% CI, 2.89-7.00; p<.001). Progression was associated with a higher overall stage at diagnosis, tumor stage, lymph node stage, low-level blood involvement, as measured with TCR data and/or flow cytometry, and elevated lactate dehydrogenase (LDH). Limitations included missing data for LDH, imaging, peripheral blood TCR data, or flow cytometry assessed at diagnosis. Staging and baseline laboratory assessments with imaging, peripheral blood flow cytometry, TCR data, and LDH in patients who have newly diagnosed MF may identify those who are at risk for progression to advanced stage.

Indolent T-cell lymphoproliferative disorder of gastrointestinal tract with unusual clinical courses: report of 6 cases and literature review.

Indolent T-cell lymphoproliferative disorder of the gastrointestinal tract (iTLPD-GI) is a rare neoplasm usually having an indolent clinical course and easily misdiagnosed as inflammatory bowel disease or other T-cell lymphomas. A subset of the disorders that progressed to overt peripheral T-cell lymphoma have been reported, and the etiology and pathogenesis are poorly understood. The current study retrospectively examined the pathological, molecular, and clinical features of 6 cases of iTLPD-GI. Hematoxylin and eosin staining, immunohistochemistry, in situ hybridization, T-cell receptor gene rearrangement, and next-generation sequencing (NGS) were performed with the diseased tissues. All the 6 patients were immunocompetent Chinese men, who presented with recurrent abdominal pain and diarrhea for 4 to 13years. Histologically, the intestinal tissue was expanded by lymphoid infiltration, composed of small-to-medium-sized lymphocytes with gland intact. The neoplastic cells were CD4 - /CD8 + with expression of TIA1 and variable granzyme B in five cases, and the other one was CD4 + /CD8 - . Two of the 5 patients progressed to more aggressive T-cell lymphoma and died of disease with complications. NGS identified TET2 and DDX3X mutations in patient 1, and BIRC6 and REV3L mutations in patient 2. Literature review indicated that iTLPD-GI with CD4 - /CD8 + immunophenotype was more commonly reported in Chinese cases. Our limited data indicated CD4-/CD8 + iTLPD-GI have similar potential to progress to more aggressive T-cell lymphoma as that of CD4 + /CD8 - , and gradually increased expression of granzyme B and Ki-67 may be early signs of the disease progression. Gain of novel gene mutations may be indicators of the pathogenesis.

BCR::ABL1-Positive Acute Lymphoblastic Leukemia: Different Clinical Behavior and Relevance of Prognostic Features in Typical ALL and in CML-like Disease

Recently we showed that in 20-30% of children diagnosed as BCR::ABL1-positive (Ph+) acute lymphoblastic leukemia (ALL) the BCR::ABL1 fusion is present in a wider clone, involving myeloid cells, non-ALL B-cells and T-cells. This leads to discordant minimal residual disease (MRD) levels assessed by the quantification of two available MRD targets - immunoglobulin (IG)/T-cell receptor (TR) genes rearrangements and BCR::ABL1 fusion. As the cases with multilineage BCR::ABL1 involvement resemble lymphoid blast crisis (LBC) of chronic myeloid leukemia (CML) we named these leukemias "CML-like", as distinct from "typical Ph+ ALL". To reveal biological and clinical differences between the two subtypes, we retrospectively analyzed prognostic relevance of MRD and other features in 147 children with BCR::ABL1-positive ALL treated according to EsPhALL (n = 133) or other (n = 14) protocols. MRD was assessed by DNA-based monitoring of BCR::ABL1 genomic breakpoint and of clonal IG/TR rearrangements. Although overall prognosis of CML-like (n = 48) and typical Ph+ ALL (n = 99) was similar (5-year EFS 60% and 49%; 5-year OS 75% and 73%, respectively), typical Ph+ ALL presented more relapses (12/48 vs. 42/99; p = 0.046) while CML-like patients more often died in the first remission (6/48 vs. 3/99; p = 0.059 for CR1 deaths in total and p = 0.01 for CR1 deaths out of the total deaths). Prognostic role of MRD measured at standard treatment timepoints (TP1 - end of induction IA, day 33; TP2 - end of consolidation IB, week 12) was highly significant in the typical Ph+ ALL (p = 0.0005 for EFS). In contrast, in CML-like patients prognostic impact of MRD was not significant and thus inapplicable for therapy adjustment. Similarly, although diagnostic white blood cell counts were similar in the two subgroups, impact of the hyperleukocytosis ≥ 50 x 109/l on outcome was highly significant in the typical Ph+ ALL (and remained significant in multivariate analyses, being apparent even in patients with fast MRD response), but no prognostic impact was found in CML-like patients. While in the typical Ph+ ALL the NCI risk tended to impact 5-year OS (89% vs. 67%; p = 0.03), there was no impact on prognosis in the CML-like patients. The BCR::ABL1 transcript type (minor/p190 vs. Major/p210), IKZF1 deletion, sex, or the age at diagnosis (< 10 vs. ≥ 10 years) did not show any prognostic relevance. Earlier start of continuous tyrosine kinase inhibitors (TKI) treatment (day 15 vs. day 33) was associated with lower MRD levels in the typical Ph+ ALL but not in CML-like patients. However, earlier TKI start tended to result in better 5-year OS in both subtypes (78% vs. 67%; p = 0.082 and 83% vs. 60%; p = 0.066 for typical Ph+ ALL and CML-like, respectively). To shed more light on the nature of the two subtypes, we now collect additional data (including transcriptome profiles, single cell characteristics, genomic breakpoint features) in order to reveal biological background, cell of origin and further potential differences between the subtypes. Such data could help to find optimal treatment strategies, and also to answer the attractive question whether CML-like disease is in fact a regular CML in LBC or whether there is a wider spectrum of BCR::ABL1-positive leukemias. If these studies lead to a conclusion that CML-like is indeed indistinguishable from LBC-CML, it will be necessary to re-define CML as a disease manifesting - at least in childhood - very often in LBC and with the minor-BCR::ABL1/p190 fusion variant. In conclusion, our data unequivocally confirm that although clinically presenting at diagnosis as one entity, there are two biologically distinct subtypes of BCR::ABL1-positive ALL - typical Ph+ ALL and CML-like disease. The different biology is reflected in treatment response and although the overall survival rates of both subtypes are similar on current protocol, the key reasons of treatment failure are different - toxicity of intensive ALL treatment in CML-like patients and relapses in typical Ph+ ALL. Thus, early distinguishing of the two subtypes is essential to enable optimal treatment approach and therapy adjustments in upcoming trials. For the typical Ph+ ALL, TKI and concurrent chemotherapy with risk-directed intensity should be recommended; in the CML-like disease, representing one-fourth to one-third of childhood patients diagnosed as BCR::ABL1-positive ALL, no relevant risk factor applicable for therapy tailoring was found so far.

Methods for Study of Mouse T Cell Receptor α and β Gene Rearrangements.

Quantitative real-time PCR and next-generation sequencing (NGS) are invaluable techniques to analyze T cell receptor (Tcr) gene rearrangements in mouse lymphocyte populations. Although these approaches are powerful, they also have limitations that must be accounted for in experimental design and data interpretation. Here, we provide relevant background required for understanding these limitations and then outline established quantitative real-time PCR and NGS methods that can be used for analysis of mouse Tcra and Tcrb gene rearrangements in mice.

Cyclic Thrombocytopenia: A Case Report

Introduction Cyclic thrombocytopenia (CTP) is a rare disorder characterized by episodes of oscillating platelet counts. It is usually associated with another primary hematologic disorder but there are idiopathic cases in the literature. Patients with CTP have similar clinical features and symptoms with idiopathic thrombocytopenic purpura (ITP), so they are often misdiagnosed as ITP and treated accordingly. In general, CTP does not respond to ITP therapies, so it should always be considered when treatments are not effective. It is more common in females. However, in our case report we present a 93-year-old male with CTP who was initially mistaken for ITP. Case description A 93-year-old male patient with a past medical history of hypertension was initially thought to have idiopathic thrombocytopenia (ITP) presenting with mucosal gum bleeding and a petechial rash. He was found to have isolated thrombocytopenia, and no evidence of other cytopenias. He did trials of multiple medications such as corticosteroids and intravenous immunoglobulin (IVIG) with no response. He was not a surgical candidate for splenectomy. He also deferred a bone marrow biopsy, though peripheral blood testing with flow cytometry showed a small monoclonal B cell population. Peripheral blood myeloid sequencing panel showing ASXL1 and EZH2. He was started on Eltrombopag, a thrombopoietin (TPO) agonist with significant lability in the platelet count, from 8,000 per microliter (mcL) to over 1,000,000 per mcL. Romiplostim, another TPO agonist also gave similar results. A diagnosis of cyclic thrombocytopenia was suspected. A Helicobacter pylori antibody was positive, and treated with antibiotics with no improvement in platelet count oscillations. Thyroid function was within normal limits. T-cell gene rearrangement was negative. Thrombopoetin (TPO) levels were checked and these were found to be in discordance with the platelet count, consistent with a CTP diagnosis. He was started on cyclosporine 150 milligrams twice daily with significant improvement in fluctuations of platelet counts. Discussion Cyclic hematopoietic disorders are rare and have regular predictable oscillations of one or more cellular components of the blood. Physiologically, there is a complex interplay between thrombopoietic stimulation and feedback inhibition in order to maintain the circulating platelets within the normal range. An error in any of these steps results in oscillation of the platelet counts (cyclic thrombocytopenia). Many mechanisms are behind the pathophysiology of this disease, such as: autoimmune destruction of platelets, hypoplasia of megakaryocytes, a defect in regulating platelet formation and an infectious or hormonal etiology. There are many reports linking T-lymphocytes proliferation and clonal TCR gene rearrangement to cyclic thrombocytopenia. We noticed in the literature that there can be variable responses to TPO agonists, but often can result in very high platelet counts. Due to its rarity, there are only case reports which show reports of remission after hormone therapy with danazol and immune suppressant therapies, such as cyclosporine. Further studies are needed to determine the mechanism responsible for cycling phenomena and optimal treatments for this disorder. In conclusion, any presumed ITP case that does not respond well to the standard therapy of ITP should be reevaluated for the possibility of cyclic thrombocytopenia.

Focal Myositis with CD8+T-Cell predominance: an Inflammatory Myositis Mimicking a Soft Tissue Neoplasm

Abstract Introduction/Objective FM is a rare self-limiting T-cell rich lesion arising within muscles of young adults as a solitary lesion and can be confused with a variety of neoplastic/inflammatory conditions or lymphoma. Here we describe a rare T-cell rich variant of FM with CD8 predominance. Methods/Case Report A 51-year-old female presented with two-month history of left trapezius swelling. MRI showed an enhancing tumor within the muscle, suspicious of sarcoma, less likely myeloma. Results (if a Case Study enter NA) Biopsy showed diffuse infiltration of small lymphocytes in a fibrotic background and atrophic skeletal muscle. These lymphocytes stained positive for CD2, CD3, CD7, CD8, CD5 (partial), TIA1 (partial), CD43, CD57 and negative for CD4, CD30, CD56, CD57, granzyme B, perforin, BCL-6, BCL-2, Pax5, TCL1, and CD56. Ki-67 was (&amp;lt;10%) with few background CD20+ B cells. By flow cytometry, the CD8+ T-cells co-express CD2, CD3, CD7, with dim CD5 expression. TCR gene rearrangement study by PCR showed no clonal TCR Gamma gene rearrangement. Conclusion FM is extremely rare with only 22 cases well-described in the literature, all predominantly composed of CD4+ T-cells, clinically concerning for low-grade sarcomas, as inflammatory myofibroblastic tumor, inflammatory leiomyosarcoma, or liposarcoma. Careful analysis is essential for correct diagnosis. Mimics include other causes of myositis, such as polymyositis and inclusion body myositis. Depending on the extent of lymphocytic infiltrate, gene rearrangement studies might be necessary to rule out clonality. Recognition of this rare entity with excellent prognosis is crucial to provide appropriate management and avoid unnecessary, aggressive procedures.

MYCOSIS FUNGOIDES (MF): THE MASTER OF MIMICRY

<h3>Introduction</h3> Mycosis fungoides (MF) is the most common form of primary cutaneous T-cell lymphomas, characterized by a long clinical course and malignant T-cell proliferation. The early stages of MF can mimic atopic dermatitis (AD), and psoriasis and thus, diagnosis can be challenging and oftentimes delayed. <h3>Case Description</h3> A 39-year-old African American female with developmental delay was referred for a mildly pruritic, generalized eczematous dermatitis, hyperkeratosis of the palms and soles, and contractures with fingernail clubbing. Skin biopsy two years prior was reportedly unremarkable. Treatment for an initial diagnosis of fungal dermatitis with mupirocin, clotrimazole, and lotrisone was started with regular debridement of the feet. Rheumatological workup was negative, including x-rays of the hands and chest. Clobetasol wraps provided short-term relief. Dupixent for AD was discussed but not started. A second skin biopsy ultimately revealed erythrodermic MF. Flow cytometry was compatible with Sezary syndrome and T-cell receptor gene rearrangement study was positive for monoclonal gene rearrangement. <h3>Discussion</h3> MF diagnosis is not easy, primarily due to the atypical clinical presentation of the disease at an early stage, and non-specific changes on histopathology. As such, multiple biopsies may be needed to make a diagnosis. This case highlights important points for the allergist. 1) Erythema in skin of color is hard to appreciate 2) MF can often look like AD and 3) Palmoplantar Keratoderma may be associated with MF.

Primary Vitreoretinal Lymphoma: Current Diagnostic Laboratory Tests and New Emerging Molecular Tools.

Primary vitreoretinal lymphoma (PVRL), a rare aggressive malignancy primarily involving the retina and/or the vitreous, is a major diagnostic challenge for clinicians (who commonly misdiagnose it as chronic uveitis) as well as for pathologists (for biological and technical reasons). Delays in diagnosis and treatment are responsible for visual impairments and life-threatening consequences, usually related to central nervous system involvement. The identification of lymphoma cells in vitreous fluid, obtained by vitrectomy, is required for diagnosis. Of note, the scarcity of neoplastic cells in small volumes of vitreous sample, and the fragility of lymphoma cells with degenerative changes caused by previous steroid use for presumed uveitis makes diagnosis based on cytology plus immunophenotyping difficult. Interleukin levels, immunoglobulin heavy chain or T-cell receptor gene rearrangements, and MYD88 mutation are applied in combination with cytology to support diagnosis. We aim to describe the current laboratory technologies for PVRL diagnosis, focusing on the main issues that these methods have. In addition, new emerging diagnostic strategies, such as next-generation sequencing analysis, are discussed. The genetic profile of PVRL remains largely unexplored. Better knowledge of genetic alterations is critical for precision medicine interventions with target-based treatments of this lymphoma for which no standardised treatment protocol currently exists.

Intraepithelial cytotoxic lymphocytes are associated with a poor prognosis in feline intestinal T-cell lymphoma.

The expression of cytotoxic molecules in feline intestinal T-cell lymphoma cells was examined immunohistochemically using endoscopic samples of 50 cases. Cases included 14 large-cell lymphomas (LCLs) and 36 small-cell lymphomas (SCLs). Most LCL and some SCL exhibited marked erosion and villous atrophy. Clonal T-cell receptor (TCR) gene rearrangement was detected in 10/14 (71%) LCL cases and 33/36 (92%) SCL cases. No clonal immunoglobulin heavy chain (IgH) gene rearrangement was detected. Immunohistochemically, all cases were positive for CD3 and negative for CD79α, CD30, CD56, and Foxp3. LCLs were positive for CD8 in 13/14 cases (93%), T-cell intracellular antigen 1 (TIA1) in 14/14 cases (100%), and granzyme B in 6/14 cases (43%). SCLs were positive for CD8 in 28/36 cases (78%), TIA1 in 33/36 cases (92%), and granzyme B in 2/36 cases (6%). TIA1- and granzyme B-positive neoplastic lymphocytes were predominantly observed in the mucosal epithelium of 10/50 cases (20%) and 6/50 cases (12%), respectively. No significant differences in survival time were found based on cell size or epitheliotropism. However, cases with TIA1+ and/or granzyme B+ neoplastic lymphocytes predominantly in the mucosal epithelium had significantly shorter survival times (P < .05), suggesting that mucosal epithelium infiltration of neoplastic cells with a cytotoxic immunophenotype is a negative prognostic factor. Therefore, intraepithelial cytotoxic lymphocytes may be associated with mucosal injury and impaired intestinal function, leading to a poor prognosis in cats with intestinal T-cell lymphoma.

STAT3 mutations in "gray-zone" cases of T-cell large granular lymphocytic leukemia associated with autoimmune rheumatic diseases.

A persistently increased T-cell large granular lymphocyte (T-LGL) count in the blood of more than 2 × 109/L for at least 6 months is necessary for a reliable diagnosis of T-LGL leukemia. In cases with LGL counts of approximately 0.5–2 × 109/L, a diagnosis of T-LGL leukemia can be made if clonal rearrangement of T-cell receptor (TCR) genes is present and if the patient shows typical manifestations of T-LGL leukemia, such as cytopenia, splenomegaly, or concomitant autoimmune disease. However, in cases with LGL counts of less than 0.5 × 109/L, the diagnosis of T-LGL leukemia is questionable (termed as “gray-zone” cases). Although mutations in signal transducer and activator of transcription 3 (STAT3) gene are the molecular hallmark of T-LGL leukemia, their diagnostic value in the “gray-zone” cases of T-LGL leukemia has not been evaluated – our study has been aimed to examine the prevalence of STAT3 mutations in these cases. Herein, we describe 25 patients with autoimmune rheumatic diseases, neutropenia, clonal rearrangement of TCR genes, and circulating LGL count of less than 0.5 × 109/L. Splenomegaly was observed in 19 (76%) patients. Mutations in the STAT3 were detected in 56% of patients using next-generation sequencing. Importantly, in 3 patients, no involvement of the blood and bone marrow by malignant LGLs was noted, but examination of splenic tissue revealed infiltration by clonal cytotoxic T-lymphocytes within the red pulp, with greater prominence in the cords. We suggest using the term “splenic variant of T-LGL leukemia” for such cases.

T-cell clonality testing for the diagnosis of T-cell large granular lymphocytic leukemia: Are we identifying pathology or incidental clones?

T-cell clonality testing by T-cell receptor (TCR) gene rearrangement is key to the diagnosis of T-cell lymphoproliferative disorders such as T-cell large granular lymphocytic (T-LGL) leukemia. Benign clonal T-cell expansions, however, are commonly found in patients without identifiable disease, a condition referred to as T-cell clones of uncertain significance (T-CUS). In practice, T-cell clonality testing is performed for a range of reasons and results are often challenging to interpret given the overlap between benign and malignant clonal T-cell proliferations and uncertainties in the management of T-CUS. We conducted a 5-year retrospective cohort study of 211 consecutive patients who underwent PCR-based T-cell clonality testing for suspected T-LGL leukemia at our institution to characterize the use of T-cell clonality testing and its impact on patient management. Overall, 46.4% (n=98) of individuals tested had a clonal T-cell population identified. Patients with a monoclonal T-cell population were more likely to be older, have rheumatoid arthritis and have higher lymphocyte counts compared to patients with polyclonal populations. The majority of patients eventually diagnosed and treated for T-LGL leukemia had rheumatoid arthritis and lower neutrophil counts compared to untreated patients with monoclonal T-cell populations. A diagnosis of T-LGL leukemia was made in only a minority of patients (n=48, 22.7%), and only a small proportion were treated (n=17, 8.1%). Our study suggests that T-cell clonality testing most commonly identifies incidental T-cell clones with only a minority of patients receiving a diagnosis of T-LGL leukemia and fewer requiring active treatment. These finding indicate an opportunity to improve utilization of T-cell clonality testing in clinical practice to better target patients where the results of testing would impact clinical management.

The expressional characteristics and diagnostic values of TRBC1 in mature T-cell lymphoma

目的分析TCRβ链恒定区结构域蛋白TRBC1在成熟T细胞淋巴瘤（TCL）中的表达特点，并与TCRVβ分析和TCR基因重排结果比较，探索TRBC1在TCL诊断中的价值。方法通过多参数流式细胞术（FCM）检测南京医科大学第一附属医院血液科收治的30例TCL患者（TCL组）、40名正常对照和50例无T淋巴细胞增殖性疾病患者（非TCL组）TRBC1的表达特点，同时分析TCRVβ受体库检测、TCR基因重排与TRBC1限制性表达检测在TCL中的诊断价值。结果正常对照组CD4+T和CD8+T细胞亚群TRBC1阳性表达率分别为（39.6±6.5）％和（39.3±4.4）％，非TCL组患者CD4+T和CD8+T细胞亚群TRBC1阳性表达率分别为（39.1±3.8）％和（36.0±8.4）％，TRBC1表达在两组中均呈双相表达模式。TCL组患者均为CD3+TCRγδ−，TRBC1阳性表达率>92.3％或<12.7％，所有病例均呈限制性表达模式（单克隆表达），与正常对照和非TCL组相比差异有统计学意义（P<0.001）。在T细胞克隆性检测效能方面，TRBC1灵敏性为100％，TCR基因重排阳性检出率为92.8％，TCRVβ流式试剂盒敏感性为94.1％，Kappa检验显示，三种检测方法一致性较高。结论多参数FCM检测TRBC1表达水平能够快速高效地诊断TCL，具有较好的临床应用价值。