Large Granular Lymphocyte Leukemia Research Articles

Next-Generation Sequencing Analysis of the Human TCRγδ+ T-Cell Repertoire Reveals Shifts in Vγ- and Vδ-Usage in Memory Populations upon Aging.

Immunological aging remodels the immune system at several levels. This has been documented in particular for the T-cell receptor (TCR)αβ+ T-cell compartment, showing reduced naive T-cell outputs and an accumulation of terminally differentiated clonally expanding effector T-cells, leading to increased proneness to autoimmunity and cancer development at older age. Even though TCRαβ+ and TCRγδ+ T-cells follow similar paths of development involving V(D)J-recombination of TCR genes in the thymus, TCRγδ+ T-cells tend to be more subjected to peripheral rather than central selection. However, the impact of aging in shaping of the peripheral TRG/TRD repertoire remains largely elusive. Next-generation sequencing analysis methods were optimized based on a spike-in method using plasmid vector DNA-samples for accurate TRG/TRD receptor diversity quantification, resulting in optimally defined primer concentrations, annealing temperatures and cycle numbers. Next, TRG/TRD repertoire diversity was evaluated during TCRγδ+ T-cell ontogeny, showing a broad, diverse repertoire in thymic and cord blood samples with Gaussian CDR3-length distributions, in contrast to the more skewed repertoire in mature circulating TCRγδ+ T-cells in adult peripheral blood. During aging the naive repertoire maintained its diversity with Gaussian CDR3-length distributions, while in the central and effector memory populations a clear shift from young (Vγ9/Vδ2 dominance) to elderly (Vγ2/Vδ1 dominance) was observed. Together with less clear Gaussian CDR3-length distributions, this would be highly suggestive of differentially heavily selected repertoires. Despite the apparent age-related shift from Vγ9/Vδ2 to Vγ2/Vδ1, no clear aging effect was observed on the Vδ2 invariant T nucleotide and canonical Vγ9–Jγ1.2 selection determinants. A more detailed look into the healthy TRG/TRD repertoire revealed known cytomegalovirus-specific TRG/TRD clonotypes in a few donors, albeit without a significant aging-effect, while Mycobacterium tuberculosis-specific clonotypes were absent. Notably, in effector subsets of elderly individuals, we could identify reported TRG and TRD receptor chains from TCRγδ+ T-cell large granular lymphocyte leukemia proliferations, which typically present in the elderly population. Collectively, our results point to relatively subtle age-related changes in the human TRG/TRD repertoire, with a clear shift in Vγ/Vδ usage in memory cells upon aging.

Mutations in the signal transducer and activator of transcription family of genes in cancer.

In recent years, it has become clear that members of the signal transducer and activator of transcription (STAT) family of genes play an important role in cancer. The STAT family consists of seven genes, STAT1‐4,STAT5A, STAT5B and STAT6, that are involved in regulating cellular proliferation, apoptosis, angiogenesis and the immune system response. Constitutive activation of STAT3, via mutational changes, is important in oncogenesis in both solid and hematopoietic cancers. In the case of hematopoietic neoplasms, STAT3 driver mutations have been described in T‐cell large granular lymphocytic (T‐LGL) leukemia and chronic natural killer lymphoproliferative disorders (CLPD‐NK) and are seen in 30%‐40% of T‐LGL leukemia patients. STAT5B is also mutated in T‐LGL leukemia and CLPD‐NK, but in a much smaller proportion. Here we review past and current research on STAT genes in hematopoietic and solid cancers with emphasis on STAT3 and STAT5B and their roles in the pathogenesis of hematopoietic malignancies, particularly T‐LGL leukemia and CLPD‐NK.

Dysregulation of the IFN-γ-STAT1 signaling pathway in a cell line model of large granular lymphocyte leukemia.

T cell large granular lymphocyte leukemia (T-LGLL) is a rare incurable disease that is characterized by defective apoptosis of cytotoxic CD8+ T cells. Chronic activation of the Janus Kinase-Signal Transducer and Activator of Transcription (JAK-STAT) pathway is a hallmark of T-LGLL. One manifestation is the constitutive phosphorylation of tyrosine 701 of STAT1 (p-STAT1). T-LGLL patients also exhibit elevated serum levels of the STAT1 activator, interferon-γ (IFN-γ), thus contributing to an inflammatory environment. In normal cells, IFN-γ production is tightly controlled through induction of IFN-γ negative regulators. However, in T-LGLL, IFN-γ signaling lacks this negative feedback mechanism as evidenced by excessive IFN-γ production and decreased levels of suppressors of cytokine signaling 1 (SOCS1), a negative regulator of IFN-γ. Here we characterize the IFN-γ-STAT1 pathway in TL-1 cells, a cell line model of T-LGLL. TL-1 cells exhibited lower IFN-γ receptor protein and mRNA expression compared to an IFN-γ responsive cell line. Furthermore, IFN-γ treatment did not induce JAK2 or STAT1 activation or transcription of IFN-γ-inducible gene targets. However, IFN-β induced p-STAT1 and subsequent STAT1 gene transcription, demonstrating a specific IFN-γ signaling defect in TL-1 cells. We utilized siRNA targeting of STAT1, STAT3, and STAT5b to probe their role in IL-2-mediated IFN-γ regulation. These studies identified STAT5b as a positive regulator of IFN-γ production. We also characterized the relationship between STAT1, STAT3, and STAT5b proteins. Surprisingly, p-STAT1 was positively correlated with STAT3 levels while STAT5b suppressed the activation of both STAT1 and STAT3. Taken together, these results suggest that the dysregulation of the IFN-γ-STAT1 signaling pathway in TL-1 cells likely results from low levels of the IFN-γ receptor. The resulting inability to induce negative feedback regulators explains the observed elevated IL-2 driven IFN-γ production. Future work will elucidate the best way to target this pathway, with the ultimate goal to find a better therapeutic for T-LGLL.

Divergent roles for antigenic drive in the aetiology of primary versus dasatinib-associated CD8+ TCR-V\u03b2+ expansions

CD8+ T-cell expansions are the primary manifestation of T-cell large granular lymphocytic leukemia (T-LGLL), which is frequently accompanied by neutropenia and rheumatoid arthritis, and also occur as a secondary phenomenon in leukemia patients treated with dasatinib, notably in association with various drug-induced side-effects. However, the mechanisms that underlie the genesis and maintenance of expanded CD8+ T-cell receptor (TCR)-Vβ+ populations in these patient groups have yet to be fully defined. In this study, we performed a comprehensive phenotypic and clonotypic assessment of expanded (TCR-Vβ+) and residual (TCR-Vβ−) CD8+ T-cell populations in T-LGLL and dasatinib-treated chronic myelogenous leukemia (CML) patients. The dominant CD8+ TCR-Vβ+ expansions in T-LGLL patients were largely monoclonal and highly differentiated, whereas the dominant CD8+ TCR-Vβ+ expansions in dasatinib-treated CML patients were oligoclonal or polyclonal, and displayed a broad range of memory phenotypes. These contrasting features suggest divergent roles for antigenic drive in the immunopathogenesis of primary versus dasatinib-associated CD8+ TCR-Vβ+ expansions.

T-Cell Large Granular Lymphocytic Leukemia and Coexisting B-Cell Lymphomas: A Study From the Bone Marrow Pathology Group.

T-cell large granular lymphocytic (T-LGL) leukemia is associated with B-cell lymphomas (BCLs), especially small BCLs. We aimed to explore and expand upon its association with BCLs. We retrospectively studied clinicopathologic features of T-LGL leukemia patients with coexisting BCL from January 2001 to December 2016. Among 432 patients with T-LGL leukemia, 22 (5.1%) had an associated B-cell non-Hodgkin lymphoma. Thirteen (59%) patients had large and nine (41%) had small BCL. T-LGL leukemia occurred synchronously with BCL in five, preceded BCL in three, and followed BCL in 14 patients. Anemia was the most common cytopenia (68%). Only one patient had a history of rheumatoid arthritis. To our knowledge, this is the first multicenter study looking at the spectrum and incidence of BCLs in patients with T-LGL leukemia and highlights its association with large BCLs (3% of T-LGL leukemias).

181 A Rare and Aggressive Gamma-Delta (γδ) T-Cell Lymphoma, NOS Presenting as Heart Failure Without Leukocytosis: Clinicopathologic Course and Autopsy Findings

A 54-year-old man presenting with acute onset dyspnea on exertion in the absence of leukocytosis was found with global hypokinesis and 20%-25% EF on echocardiogram, consistent with systolic and diastolic heart failure. No evidence of obstructive CAD to support ischemic etiology was noted on catheterization. Microscopy of peripheral blood identified a population of medium to large lymphocytes with high nuclear/cytoplasmic ratios and convoluted, irregular nuclei. Flow cytometry showed a γδ T-cell population brightly expressing CD45 with intermediate forward and side scatter, CD2, CD3, and CD5 expression with loss of CD7, negative CD4 and CD8, upregulated CD57, and negativity of CD10, CD25, CD30, and CD56. A γδ T-cell lymphoproliferative disorder involving peripheral blood was suspected. Three main types include γδ T-cell large granular lymphocyte leukemia (T-LGL), which is indolent, whereas hepatosplenic γδ T-cell lymphoma (HSTCL) and mucocutaneous γδ T-cell lymphoma are aggressive. CD57 expression is specific for γδ T-cell LGL over the aggressive entities, which could suggest an indolent course; however, morphologic characteristics of γδT-cells differ from LGLs and are more suggestive of a lymphoma. Thoracentesis, liver and bone marrow biopsies found a γδ T-cell population identical to those in peripheral blood. A perisinusoidal and periportal pattern of liver involvement argued against HSTCL in which purely intrasinusoidal growth is typical. Expression of D5+, CD7–, CD56–, CD57+, TIA–, granzyme negative, and perforin-negative contrasts with that typical of HSTCL (CD5–, CD7+, CD56+, CD57–, TIA+, granzyme negative, and perforin negative). Also, HSTCL cases usually demonstrate isochromosome 7q, which was not seen in the patient’s lymphoma despite a complex karyotype. Following a rapidly fatal course, autopsy found virtually all body sites except the central nervous system were diffusely involved by γδ T-cell lymphoma of a rare immunophenotype not previously described. Infiltration of cardiac muscle provides a potential underlying etiology for the presenting symptoms of heart failure.

T-Cell Large Granular Lymphocytic Leukemia – Case Report

SUMMARY – T-cell large granular lymphocytic leukemia (T-LGLL) is an uncommon but probably underdiagnosed disease caused by clonal proliferation of large granular lymphocytes. Diagnosis is typically based on the high number of morphologically characteristic lymphoid cells and finding of an abnormal immunophenotype by flow cytometry. Because of its relatively indolent clinical behavior, observation is often an appropriate therapy. Here we present a case of a 53-year-old male admitted to the hospital because of abdominal pain. Blood examination revealed mild mycrocitic anemia and multiplied lactate dehydrogenase level. Abdominal ultrasound showed splenomegaly of 16 cm, with no lymphadenopathy. Fine needle aspiration of bone marrow revealed hypocellular marrow with 50% of atypical lymphoid cells. There were 81% of atypical medium sized granular lymphocytes with irregularly shaped nuclei in peripheral blood, so the cytologic diagnosis was lymphoproliferative process. Bone marrow biopsy showed nodular and interstitial proliferation of small, partially atypical T lymphocytic cells positive for CD2, CD3, CD5, CD8, granzyme and TIA, and negative for hairy cell markers, CD10, MUM 1, bcl 1, CD4 and CD56. The finding was consistent with T-LGLL. Due to splenomegaly, the patient was treated with cyclosporine and gradually reduced dose of corticosteroids, leading to regression of splenomegaly and normalization of lactate dehydrogenase level.

Hereditary Multiple Exostoses Misdiagnosed as Rheumatoid Arthritis

Making a diagnosis of rheumatoid arthritis is often tricky. With no single test that confirms or eliminates the disease, the diagnosis of RA is based on a physical exam, patient history, laboratory tests and often imaging. However, symptoms of RA, such as pain, swelling and fatigue, are not exclusive to the disease. And correct diagnosis is important in choosing an appropriate treatment plan. A number of diseases, such as lupus, fibromyalgia or Sjogren’s syndrome, may easily be confused with RA, or coexist in a patient. Arthritis symptoms might develop following certain infections, such as Lyme disease, tuberculosis, gastrointestinal infection or sexually-transmitted diseases. Patients with certain cancers, such as large granular lymphocyte (LGL) leukemia, have an increased incidence of RA, an acute leukemia in children may even be misdiagnosed as idiopathic juvenile arthritis. In this case report a 25 years male patient presented with arthralgia, fatigue, swelling of swelling of all MCPs, PIPs both wrists and knees, tender shoulders, Limited range of motion of right elbow, limitation of ROM of knees, severe cervical muscle spasm, was misdiagnosed as rheumatoid arthritis with 2ry Sjogren's syndrome with no response to treatment; imaging revealed multiple exostoses at distal end of Rt radius, ulna and distal femoral and tibial end.

STAT3 mutation and its clinical and histopathologic correlation in T-cell large granular lymphocytic leukemia

Although T-cell large granular lymphocytic leukemia (T-LGLL) is a clinically indolent disorder, patients with moderate to severe cytopenia require therapeutic intervention. The recent discovery of STAT3 mutations has shed light on the genetic basis of T-LGLL pathogenesis. However, the association of STAT3 mutational status with patients' clinical, histopathologic, and other laboratory features has not been thoroughly evaluated in T-LGLL. In this study, STAT3 mutations were identified in 18 of 36 patients with T-LGLL (50%), including Y640F (12/18, 66.7%), N647I (3/18, 16.7%), E638Q (1/18, 5.6%), I659L (1/18, 5.6%), and K657R (1/18, 5.6%). Interestingly, pure red cell aplasia was seen exclusively in T-LGLL patients without STAT3 mutations (6/15 in the wild-type STAT3 group versus 0/13 in the mutant STAT3 group; P = .02); these patients also were the only responders to T-LGLL therapy (mainly cyclophosphamide) in wild-type STAT3 group. Patients harboring STAT3 mutations were more prone to rheumatoid arthritis (4/13 versus 0/15 in the wild-type STAT3 group; P = .04), frequently requiring therapy for neutropenia/neutropenia-associated infections, and demonstrated good therapeutic responses to methotrexate. No significant differences were seen in complete blood count, flow cytometric immunophenotypic features, T-cell receptor γ V-J rearrangement repertoire, and bone marrow biopsy morphology among the STAT3-mutation and wild-type groups other than significantly larger tumor burden in patients with STAT3 mutations. The distinct disease association and therapeutic responses observed in patients with mutant and wild-type STAT3 warrant further investigation to elucidate the underlying mechanisms. They also highlight the importance of identifying STAT3 mutational status in patients with T-LGLL, which may aid in clinical therapeutic choice.

Chronic neutropenia in LGL leukemia and rheumatoid arthritis.

This section reviews the diagnostic criteria and pathogenesis of large granular lymphocyte (LGL) leukemia. There is a particular focus on the overlap of LGL leukemia and rheumatoid arthritis (Felty's syndrome). Current understanding of the mechanisms of neutropenia in these disorders is discussed. Finally, treatment indications and therapeutic recommendations are outlined.

Somatic STAT3 mutations in Felty syndrome: an implication for a common pathogenesis with large granular lymphocyte leukemia.

Felty syndrome is a rare disease defined by neutropenia, splenomegaly, and rheumatoid arthritis. Sometimes the differential diagnosis between Felty syndrome and large granular lymphocyte leukemia is problematic. Recently, somatic STAT3 and STAT5B mutations were discovered in 30–40% of patients with large granular lymphocyte leukemia. Herein, we aimed to study whether these mutations can also be detected in Felty syndrome, which would imply the existence of a common pathogenic mechanism between these two disease entities. We collected samples and clinical information from 14 Felty syndrome patients who were monitored at the rheumatology outpatient clinic for Felty syndrome. Somatic STAT3 mutations were discovered in 43% (6/14) of Felty syndrome patients with deep amplicon sequencing targeting all STAT3 exons. Mutations were located in the SH2 domain of STAT3, which is a known mutational hotspot. No STAT5B mutations were found. In blood smears, overrepresentation of large granular lymphocytes was observed, and in the majority of cases the CD8+ T-cell receptor repertoire was skewed when analyzed by flow cytometry. In bone marrow biopsies, an increased amount of phospho-STAT3 positive cells was discovered. Plasma cytokine profiling showed that ten of the 92 assayed cytokines were elevated both in Felty syndrome and large granular lymphocyte leukemia, and three of these cytokines were also increased in patients with uncomplicated rheumatoid arthritis. In conclusion, somatic STAT3 mutations and STAT3 activation are as frequent in Felty syndrome as they are in large granular lymphocyte leukemia. Considering that the symptoms and treatment modalities are also similar, a unified reclassification of these two syndromes is warranted.

Rational management approach to pure red cell aplasia.

Pure red cell aplasia is an orphan disease, and as such lacks rationally established standard therapies. Most cases are idiopathic; a subset is antibody-mediated. There is overlap between idiopathic cases and those with T-cell large granular lymphocytic leukemia, hypogammaglobulinemia, and low-grade lymphomas. In each of the aforementioned, the pathogenetic mechanisms may involve autoreactive cytotoxic responses. We selected 62 uniformly diagnosed pure red cell aplasia patients and analyzed their pathophysiologic features and responsiveness to rationally applied first-line and salvage therapies in order to propose diagnostic and therapeutic algorithms that may be helpful in guiding the management of prospective patients, 52% of whom were idiopathic, while the others involved large granular lymphocytic leukemia, thymoma, and B-cell dyscrasia. T-cell-mediated responses ranged between a continuum from polyclonal to monoclonal (as seen in large granular lymphocytic leukemia). During a median observation period of 40 months, patients received a median of two different therapies to achieve remission. Frequently used therapy included calcineurin-inhibitors with a steroid taper yielding a first-line overall response rate of 76% (53/70). Oral cyclophosphamide showed activity, albeit lower than that produced by cyclosporine. Intravenous immunoglobulins were effective both in parvovirus patients and in hypogammaglobulinemia cases. In salvage settings, alemtuzumab is active, particularly in large granular lymphocytic leukemia-associated cases. Other potentially useful salvage options include rituximab, anti-thymocyte globulin and bortezomib. The workup of acquired pure red cell aplasia should include investigations of common pathological associations. Most effective therapies are directed against T-cell-mediated immunity, and therapeutic choices need to account for associated conditions that may help in choosing alternative salvage agents, such as intravenous immunoglobulin, alemtuzumab and bortezomib.

Results from a First-in-Human Study with Bnz-1: A Novel Peptide Inhibitor of IL-2, IL-9 and IL-15 for the Treatment of T-Cell Malignancies That Safely and Selectively Decreases Regulatory T-Cells, Natural Killer Cells, and CD8+ Central Memory T-Cells

Background: The γc-family of cytokines (IL-2, IL-4, IL-7, IL-9, IL-15, IL-21) control major immune responses and lymphocyte development. However, aberrant γc cytokine activity contributes to or pathologically drives human diseases including T-cell malignancies, graft-versus-host disease, and numerous autoimmune diseases (e.g., alopecia areata), where IL-2, IL-9, and IL-15 have been specifically implicated. Importantly, these multi-cytokine diseases are not effectively and safely treated by the currently available anti-cytokine approaches.Methods: BNZ-1 is the first of a novel class of rationally-designed, multi-cytokine inhibitors that selectively inhibits IL-2 and IL-15, and to a lesser degree, IL-9 signaling through the γc receptor, without affecting IL-4, IL-7, or IL-21. Preclinical proof-of-concept has been demonstrated in several animal models of T-cell malignancies (Nata et al., 2015 J Biol Chem), including Large Granular Lymphocyte Leukemia (LGL) and Adult T-cell Leukemia. In this open-label, single-dose, dose-escalation, first-in-human study conducted at a single center in the United States (NCT03046459), 18 healthy adults (n=3/cohort) received a single intravenous dose of 0.2, 0.4, 0.8, 1.6, 3.2 or 6.4 mg/kg infused over ≤5 minutes on Day 1 and were followed for safety and sample collection for 30 days.Results: All subjects completed the study. BNZ-1 was considered well-tolerated with a good safety profile with no serious or severe adverse events (AEs), no dose-limiting toxicities, and no clinically-significant changes on clinical labs (serum biochemistry, hematology, liver enzymes), vital signs or electrocardiograms. Headache was the only AE reported in ≥1 subject (n=3).BNZ-1 exposure was generally dose proportional with an elimination half-life of ~5 days across the range of doses tested, which is consistent with other PEGylated peptides and supports weekly or every other week dosing.The pharmacodynamic activity of BNZ-1 was characterized by flow cytometry of PBMCs obtained on Days 4, 15 and 31 and used to calculate the maximum change from baseline (Emax) for regulatory T-cells (Tregs; IL-2 effect), NK cells (IL-15 effect) and CD8+ central memory T-cells (Tcm; IL-2 & IL-15 effect). For Tregs, a ~50-60% decrease was observed on Day 4 after doses of 0.4 to 1.6mg/kg, while doses of 3.2 and 6.4mg/kg produced an 80-93% decrease from baseline that peaked on Day 15. NK cells were decreased from baseline on Day 4 by 20%, 40% and 60% at 0.2, 0.4 and 0.8mg/kg, respectively, and plateaued at 70 to 80% decrease at doses ≥1.6 mg/kg. Tcm were decreased at Day 4 for the three highest dose cohorts that continued to decline to Day 15 when all doses, except 0.2 mg/kg, showed a mean decrease ranging from 10 to 81% that generally trended with dose. Tregs, NK cells, and Tcm returned to/toward baseline by 30 days post dose. Post-dose counts of CD4+ and CD8+ T-cells, B-cells, and monocytes were unchanged at all time points studied.Conclusions: These preliminary clinical data suggest that BNZ-1 is a highly-active, selective immunomodulator that safely decreases Tregs, NK cells and Tcm, while leaving the major leukocyte populations unaffected. These data support the planned Phase 1/2 dose-ranging study in LGL and Cutaneous T-cell Lymphoma. DisclosuresFrohna:Bioniz Therapeutics: Employment. Tagaya:Bioniz Therapeutics: Equity Ownership. Ratnayake:Bioniz Therapeutics: Consultancy. Doerr:Bioniz Therapeutics: Employment. Basheer:Bioniz Therapeutics: Employment. Al-Mawsawi:Bioniz Therapeutics: Employment. Kim:Bioniz Therapeutics: Employment. Azimi:Bioniz Therapeutics: Employment.

Successful treatment of cerebral aspergillosis: case report of a patient with T-cell large granular lymphocytic leukemia (T-LGL)

BackgroundInvasive aspergillosis involving patients with neutropenia or severe immunosuppression, such as patients with hematologic malignancies is associated with high mortality. Patients with T-cell large granular lymphocytic leukemia (T-LGL) on the other hand are considered to be less vulnerable for severe opportunistic fungal infection as their course of disease is chronic and marked by less violent cytopenia then in e.g. Aplastic Anemia. Only neutropenia is regarded as independent risk factor for severe opportunistic infection in T-LGL patients.Case presentationWe report a case of a 53 year old patient with T-LGL, Immune-Thrombocytopenia (ITP) and combined antibody deficiency, who presented with fever and reduced general condition. The patient revealed a complicated infection involving the lungs and later the brain, with the presentation of vomiting and seizures. Broad microbiological testing of blood-, lung- and cerebrospinal fluid samples was inconclusive. In the absence of mycological proof, Aspergillus infection was confirmed by pathological examination of a brain specimen and finally successfully treated with liposomal amphotericin B and voriconazole, adopting a long-term treatment scheme.ConclusionsBeyond typical problems in the clinical practice involving fungal infections and hematologic malignancies, this case of invasive aspergillosis in a patient with T-LGL illustrates caveats in diagnosis, therapy and follow-up. Our data support careful ambulatory monitoring for patients with T-LGL, even in the absence of neutropenia. Especially those patients with combined hematologic malignancies and immune defects are at risk. Long-term treatment adhesion for 12 months with sufficient drug levels was necessary for sustained clearance from infection.

Case report on a rare pulmonary bacterial infection by Actinomyces odontolyticus in a T-cell large granular lymphocyte leukemia patient

Case report on a rare pulmonary bacterial infection by Actinomyces odontolyticus in a T-cell large granular lymphocyte leukemia patient

T-Cell Large Granular Lymphocyte Leukemia Diagnosis in a Patient with Felty Syndrome

T-Cell Large Granular Lymphocyte Leukemia Diagnosis in a Patient with Felty Syndrome

Drug sensitivity profiling identifies potential therapies for lymphoproliferative disorders with overactive JAK/STAT3 signaling.

Constitutive JAK/STAT3 signaling contributes to disease progression in many lymphoproliferative disorders. Recent genetic analyses have revealed gain-of-function STAT3 mutations in lymphoid cancers leading to hyperactivation of STAT3, which may represent a potential therapeutic target. Using a functional reporter assay, we screened 306 compounds with selective activity against various target molecules to identify drugs capable of inhibiting the cellular activity of STAT3. Top hits were further validated with additional models including STAT3-mutated natural killer (NK)-cell leukemia/lymphoma cell lines and primary large granular lymphocytic (LGL) leukemia cells to assess their ability to inhibit STAT3 phosphorylation and STAT3 dependent cell viability. We identified JAK, mTOR, Hsp90 and CDK inhibitors as potent inhibitors of both WT and mutant STAT3 activity. The Hsp90 inhibitor luminespib was highly effective at reducing the viability of mutant STAT3 NK cell lines and LGL leukemia patient samples. Luminespib decreased the phosphorylation of mutant STAT3 at Y705, whereas JAK1/JAK2 inhibitor ruxolitinib had reduced efficacy on mutant STAT3 phosphorylation. Additionally, combinations involving Hsp90, JAK and mTOR inhibitors were more effective at reducing cell viability than single agents. Our findings show alternative approaches to inhibit STAT3 activity and suggest Hsp90 as a therapeutic target in lymphoproliferative disorders with constitutively active STAT3.

Recurrent Lactobacillus Bacteremia in a Patient With Leukemia.

Lactobacillus species are a commensal flora of the human gastrointestinal and the female genitourinary tract. Lactobacilli especially the rhamnosus species, are common components of commercial probiotics. They are rarely associated with pathology in immunocompetent people, but they have been known to cause dental caries, bacteremia, and endocarditis in patients with suppressed immune function. Cases of Lactobacillus bacteremia have been reported in patients with acute myeloid leukemia, large granular lymphocytic leukemia, and in transplant recipients. In this article, we report a strange case of recurrent Lactobacillus bacteremia causing multiple episodes of fever of unknown origin in a patient with leukemia. This report is unique as Lactobacillus is not recognized as a common source of bacteremia. Moreover, the source of the bacillus continued to elude us even after extensive investigation.

Systematic STAT3 sequencing in patients with unexplained cytopenias identifies unsuspected large granular lymphocytic leukemia.

TO THE EDITOR: Large granular lymphocytic (LGL) leukemia and myelodysplastic syndromes (MDSs) both typically present with unexplained cytopenias, yet these diseases are pathobiologically distinct and associated with substantial differences in prognosis and therapy. LGL leukemias, including both T-

Livedoid eruption in a patient affected by T-γδ large granular lymphocyte leukaemia

Livedo is an ischaemic dermopathy characterised by a reddish-blue to violaceous mottling of the skin with a net-like reticular appearance. Livedo has been described in association with several medical conditions...