Helper Phenotype Research Articles

Immune cell profiles in synovial fluid after anterior cruciate ligament and meniscus injuries

BackgroundAnterior cruciate ligament (ACL) and meniscus tears are common knee injuries. Despite the high rate of post-traumatic osteoarthritis (PTOA) following these injuries, the contributing factors remain unclear. In this study, we characterized the immune cell profiles of normal and injured joints at the time of ACL and meniscal surgeries.MethodsTwenty-nine patients (14 meniscus-injured and 15 ACL-injured) undergoing ACL and/or meniscus surgery but with a normal contralateral knee were recruited. During surgery, synovial fluid was aspirated from both normal and injured knees. Synovial fluid cells were pelleted, washed, and stained with an antibody cocktail consisting of fluorescent antibodies for cell surface proteins. Analysis of immune cells in the synovial fluid was performed by polychromatic flow cytometry. A broad spectrum immune cell panel was used in the first 10 subjects. Based on these results, a T cell-specific panel was used in the subsequent 19 subjects.ResultsUsing the broad spectrum immune cell panel, we detected significantly more total viable cells and CD3 T cells in the injured compared to the paired normal knees. In addition, there were significantly more injured knees with T cells above a 500-cell threshold. Within the injured knees, CD4 and CD8 T cells were able to be differentiated into subsets. The frequency of total CD4 T cells was significantly different among injury types, but no statistical differences were detected among CD4 and CD8 T cell subsets by injury type.ConclusionsOur findings provide foundational data showing that ACL and meniscus injuries induce an immune cell-rich microenvironment that consists primarily of T cells with multiple T helper phenotypes. Future studies investigating the relationship between immune cells and joint degeneration may provide an enhanced understanding of the pathophysiology of PTOA following joint injury.

Bio-P-16 - Peripheral T-cell lymphoma: molecular profiling distinguishes subclasses, recognizes the tumor architecture and identifies prognostic markers

<h3>Introduction:</h3> Peripheral T-cell lymphomas (PTCLs) are aggressive tumours with unfavourable prognosis, with around 30% OS after 5 years. Histological and molecular studies have revealed a striking degree of heterogeneity, with three major PTCL subtypes defined. Consistent diagnosis and prognostication are still difficult to achieve, because of the difficulty of reproducing results, and the dearth of clinically applicable prognostic biological markers. <h3>Methods:</h3> We have analyzed a series of 105 PTCL cases (66 angioimmunoblastic T-cell lymphoma, AITL; 21 PTCL-not otherwise specified, PTCL-NOS; and 18 PTCL-with T follicular helper phenotype, PTCL-TFH) patients using a customized NanoString platform that includes 208 genes associated with T-cell differentiation, oncogenes and tumor suppressor genes, deregulated pathways and stromal cell subpopulations. Specifically, the platform includes genes expressed by the multiple cell types present in PTCL specimens, together with normal T-cell populations. These are used to try and enable the deconvolution of the T-cell lymphoma microenvironment, and thereby develop an integrated perspective on the cell composition of PTCL tumor specimens. <h3>Results:</h3> A comparative analysis of the various histological types of PTCL (AITL, PTCL-NOS and PTCL-TFH) showed specific sets of genes to be associated with each of the diagnoses, including TFH markers, cytotoxic markers and genes whose expression was a surrogate for specific cellular subpopulations, including follicular dendritic cells, mast cells and genes belonging to specific cell-survival pathways (NF-κB). Unsupervised analysis of the expression of the genes here studied revealed clusters of co-regulated genes that identified the main cell components of the tumor. The analysis did not reveal any differences in survival probability associated with the histological sub-classification, but did identify specific genes and gene sets whose expression was associated with changes in survival probability for each of the PTCL subtypes, independently of the clinical variables included in the International Prognostic Index (IPI). These included a B-cell gene set in cases of AITL, the expression of proliferation markers in PTCL-NOS, and the expression of cytotoxic markers in cases with a diagnosis of PTCL-TFH. For each PTCL lymphoma type, a multivariate analysis identified genes that allow the series of cases to be stratified into different risk groups. This was validated for AITL in an independent series of 54 additional cases. <h3>Conclusions:</h3> In summary, our study supports the current division of PTCL into these three categories (AITL, PTCL-NOS and PTCL-TFH), identifies gene sets potentially useful for this classification and recognizes the expression of B-cell genes as an IPI-independent prognostic factor for AITL subtype.

Bio-O2-08 - Mutational landscape of nodal peripheral T-cell lymphoma subtypes

<h3>Introduction:</h3> Nodal peripheral T-cell lymphoma (PTCL) subclassification (Angioimmunoblastic TCL, PTCL-with T Follicular Helper phenotype and PTCL-NOS) and therapeutic targeting is still controversial. Overall survival (OS) is only around 30% after 5 years. <h3>Methods:</h3> We performed targeted next-generation sequencing of 61 selected genes in 76 PTCL patients. Among the 76cases, 44 were classified as AITL, 18 as PTCL-TFH and 14 as PTCL-NOS. <h3>Results:</h3> Our analysis revealed a wide variety of genetic variants that possibly drive the development of AITL, PTCL-TFH and/or PTCL-NOS. We identified an average of 4 variants per patient, mainly clustered in genes regulating chromatin conformation (TET2, DNMT3A) and T-cell differentiation (RHOA, MTOR, IDH2, VAV1 and NOTCH1). More frequently mutated genes were TET2 (76,31%) and DNMT3A (27,63%). Multiple mutations were found in TET2, VAV1, RHOA, NOTCH1, MTOR, JAK1, ZEB1, ATM, DNMT3A and ARID1B. Main difference among the three Nodal PTCL classes was the higher frequency of RHOAG17V mutations (p<0.0001), present approximately 2 times more frequently in AITL cases (34,09%) than in PTCL-TFH (16,66%), and they were not detected in PTCL-NOS. All three Nodal PTCL subtypes share finding suggesting Clonal Hematopoiesis (CH) in TET2 and DNMT3A genes. Cases with higher variant allele frequencies (VAF)s in TET2 and DNMT3A genes vs. other mutated genes were attributed to CH. In our study we found this phenomenon to take place in all Nodal PTCL subtypes (AITL: 31,8%; PTCL-TFH: 50%; PTCL-NOS: 35,71%). A correlation was not found between specific mutations, mutational index, mutations attributed to CH and clinical outcome. <h3>Conclusions:</h3> There is a common molecular basis for the three Nodal PTCL with very frequent mutations in genes regulating chromatin conformation associated with mutations in genes governing T-cell differentiation. RHOA G17V mutations were highly significant enriched in AITL. Findings suggesting Clonal Hematopoiesis were found in the three types of nodal PTCL.

Targeting T-Cell Receptor Signaling Pathway in Angioimmunoblastic T-Cell Lymphoma

[Backgrounds] Angioimmunoblastic T-cell lymphoma (AITL) belongs to nodal T-cell lymphoma with T follicular helper (TFH) phenotype. Both the G17V RHOA mutation and the loss-of-function TET2 mutation have been identified in 70% of AITLs. We previously demonstrated that G17V mutant RHOA augmented T-cell receptor (TCR) signaling though binding to VAV1, an essential component of the T-cell signaling pathway (Leukemia, in press). However, it remains elusive whether inhibition of TCR pathway serves as an effective treatment in AITL. [Methods] Activation of VAV1 and PLCgamma1 in response to TCR stimulation was examined in Jurkat cells inducibly expressing wild-type and G17V mutant RHOA under the treatment with dasatinib, a multi-kinase inhibitor. Nuclear factor of activated T cell (NFAT) activity was similarly examined in Jurkat cells transiently transfected with wild-type and G17V mutant RHOA. Mice expressing G17V RHOA under CD2 promoter (G17VRHOA mice) were crossed with MxCre Tet2f/f mice and pIpC was given to the offspring 4 weeks after birth to obtain Tet2-/- and Tet2-/-G17VRHOA mice. Whole transcriptome analysis was performed for CD4+ T cells purified from the spleen of tumor developed mice. Serum cytokines were examined by BD Cytometric Beads Assay. Cells derived from tumors developed in Tet2-/-G17VRHOA mice were transplanted into nude mice. Dasatinib was orally given at 5 mg/kg/day for 14 days. [Results] The G17V mutant RHOA enhanced phosphorylation of VAV1 and PLCgamma1, and NFAT reporter activity in Jurkat cells. The phosphorylation and the NFAT activity were efficiently blocked by dasatinib at 10 nM. Approximately 70% of Tet2-/-G17VRHOA mice died around 40 wo, while 14% of Tet2-/- and none of G17VRHOA mice died at the same age (p<0.05). All the moribund Tet2-/-G17VRHOA mice presented splenomegaly and lymphadenopathy. The lymph nodes and the spleens showed diffuse infiltration of tumor cells expressing TFH markers, i.e., CD4, PD1, and ICOS. The aberrant increase of high endothelial venules was observed in the swollen lymph nodes, mimicking human AITL. Whole transcriptome analysis followed by gene-set enrichment analysis revealed enrichment of Il-2/STAT5, Il-6/JAK-STAT3, and TNFα/NFκB signaling pathways in Tet2-/-G17VRHOA CD4+ spleen cells. The serum concentrations of Il-2, Il6, and TNFα in Tet2-/-G17VRHOA were significantly higher than control at the same age (p<0.05). The CD4+ cells in the enlarged spleen and the swollen lymph nodes in Tet2-/-G17VRHOA mice at 40 wo showed oligoclonal rearrangement of TCR genes. Nude mice transplanted with tumor-derived cells showed splenomegaly and lymphadenopathy preceded by elevation ofthe serum concentrations of Il-2, Il-6 and TNFα (before vs. after the transplantation, 4.33±0.14 vs. 4.80±0.37, 5.85±2.96 vs. 34.92±15.49, and 7.68±2.08 vs. 45.70±33.72, p<0.01, respectively). Mice given with dasatinib starting at 10 days after the transplantation showed higher overall survival compared to the vehicle control (p<0.05, Figure 1). The serum concentrations of Il-2, Il-6, and TNFα were decreased in dasatinib group (before vs. after the dasatinib treatment, 4.85±0.40 vs. 4.28±0.18, 38.23±18.74 vs. 10.01±7.38, and 48.37±34.6 vs. 18.67±12.37, p<0.05, respectively).Discussions and Conclusion: The combination of G17V mutantRHOAexpression and Tet2 deletion led to development of AITL-like tumors in mice, which were hughly effectively treated by dasatinib. This model may also be useful to examine novel treatment strategies for AITL, other than dasatinib. [Display omitted] DisclosuresChiba:Nippon Shinyaku: Honoraria, Research Funding.

Comprehensive Genomic Profiling (CGP) Informs Molecular Diagnosis, Prognosis, and Therapeutic Decision-Making for Patients with Angioimmunoblastic T-Cell Lymphoma (AITL)

Background: AITL, under the 2016 WHO umbrella classification of nodal peripheral T cell lymphomas (PTCL) with a T follicular helper (TFH) phenotype, accounts for ~20% of all PTCL cases. Genomic analysis has revealed recurrent somatic mutations, such as RhoA G17V, that are hallmarks of AITL. Overlapping genomics have been observed in other PTCL subgroups, such as PTCL not otherwise specified (PTCL-NOS). We and others have reported frequent co-occurrence of RhoA G17V with mutations in the epigenetic regulators TET2 and IDH2R172 in AITL (Morley Blood 2015). Since treatment for the disease remains unsatisfactory, efforts are underway to assess the clinical utility of genomic information to guide therapy. The current study aimed to comprehensively assess the genomic landscape of AITL further, including identification of diagnostically and therapeutically relevant alterations.Methods: We explored the FoundationCORE™ database (Foundation Medicine Inc, Cambridge, MA) for cases assigned as AITL or harboring RhoA G17V positivity irrespective of disease ontology. This led to the inclusion of 42 cases: confirmed AITL (concordant diagnoses by internal and external pathologists, 29), suspected AITL (discordant diagnoses, 9), lymph node lymphoma NOS (PTCL, 1), or other neoplasm [pleomorphic myxofibrosarcoma (MFS), 1; marginal zone lymphoma (MZL), 1; cutaneous T-cell lymphoma (CTCL), 1]. Median cohort age was 65.5 years, 65 years for the AITL-specific subset; range for both, 34-88y. Pathology reports indicated Epstein-Barr virus positivity in 16/38 of AITL cases and prior autologous stem cell transplant in 4/38. DNA and RNA were extracted from formalin-fixed paraffin embedded samples or peripheral blood, and CGP performed using the CLIA-certified, NYS-approved, CAP-accredited FoundationOne® Heme assay (He Blood 2016).Results: RhoAG17V was seen in 57% (24/42) of cases in the cohort, including 53% (20/38) of AITL samples and in 1 case each of MFS, MZL, CTCL, and PTCL. Confirmed AITL harbored RhoAG17V in 52% (15/29), and suspected AITL in 56% (5/9), of samples. TET2 mutations (m TET2) were found in 89% of AITLs (26/29, confirmed; 8/9, suspected), and in 1/1 PTCL; as reported previously, m TET2 was not identified in the MFS, MZL, or CTCL. IDH2 mutations (m IDH2), all occurring at R172, were seen in 16% (6/38) of AITL cases, with all 6 co-occurring with RhoAG17V plus m TET2 . DNMT3A mutations (m DNMT3A) were seen in 32% (12/38) of AITLs (9/29, confirmed; 3/9, suspected) and in the single PTCL. RhoAG17V co-occurred with m TET2 (but notm DNMT3A orm IDH2) in 26% (10/38), m TET2 plus m DNMT3A in 7.9% (3/38), and m TET2, mDNMT3A, plus m IDH2 in 7.9% (3/38) of cases; all RhoAG17V cases(20/20) co-occurred with m TET2 . TP53 mutation was seen in 21% (8/38) of AITL cases. Other recurrent somatic mutations included BRCA1 or BRCA2 in 7.9% (3/38), CTNNB1 in 7.9% (3/38), and KRAS, ATM or STAT3 in 2.6% (1/38), of AITL samples. Assessment of tumor mutational burden (TMB), a biomarker for checkpoint inhibitor efficacy in solid tumors, revealed low TMB (<6 mutations/Mb) in 8/10 AITL cases and intermediate TMB (TMB-Intermediate, 6-19 mutations/Mb) in 2/10.Conclusion: This study utilized CGP to explore alterations of diagnostic and therapeutic potential in AITL. Confirming prior studies, in a significant proportion of AITL cases RhoAG17V co-occurred with m TET2 and m IDH2 ; all AITL cases positive for RhoAG17V had concurrent m TET2. We also found RhoAG17V, but not m TET2 or m IDH2, in solitary cases of MZL, MCL, and CTCL.Taken together, co-occurrence of RhoAG17V and m TET2 may provide greater specificity for AITL diagnosis than RhoAG17V alone. We also detected putative actionable targets in AITL. While predictors of response to hypomethylating agents (e.g. m TET2, m IDH2) were seen in most cases, a subset were found to have mutations in BRCA1/2 (3 pts), KRAS (1 pt), CTNNB1 (3 pts), or STAT3 (1 pt). These findings suggest alternative therapeutic opportunities in AITL (e.g. inhibitors of PARP, MEK, mTOR, and STAT3). We further report that mutational burden in AITL is generally low; however, a subset of cases (2/10) were TMB-Intermediate. Although the predictive power of TMB in lymphoma is unclear, in some solid tumors TMB-Intermediate status predicts response to immune checkpoint inhibition. In total, our findings highlight the utility of CGP to identify alterations that may influence the clinical management of AITL. DisclosuresMorley:Foundation Medicine: Employment, Other: Stock. He:Foundation Medicine, Inc: Employment, Other: Stock. Bailey:Foundation Medicine: Employment, Other: stock. Nahas:Foundation Medicine inc: Employment, Other: stockholder. Heilmann:Foundation Medicine, Inc: Employment, Other: Stockholder. Chudnovsky:Foundation Medicine, Inc: Employment, Other: Stockholder. Fabrizio:Foundation Medicine, Inc: Employment, Other: Stockholder. Ali:Foundation Medicine, Inc: Employment, Other: Stock. Lipson:Foundation Medicine: Employment, Other: stock. Stephens:Foundation Medicine Inc.: Employment, Equity Ownership. Ross:Foundation Medicine Inc: Employment, Other: stockholder. Miller:Foundation Medicine: Employment, Other: Stock. Severson:Foundation Medicine: Employment, Other: Stock. Vergilio:Foundation Medicine: Employment, Other: Stock. Erlich:Foundation Medicine: Employment, Other: stock. Mughal:Foundation Medicine, Inc: Employment, Other: Stock.

T-NHL subtypes and clinical perspective

Peripheral T-cell lymphomas (PTCL) are a distinct pathological entity with clinical advancements lagging behind its B-cell lymphoma counterpart. A significant proportion (15–20%) of PTCL cases are misdiagnosed as reactive or a different lymphoma subtype. An accurate diagnosis routinely requires analysis of the immunophenotype in conjunction with cellular morphology, analysis of lymph node architecture and molecular genetic analysis. Recent major advances in genomic studies examining gene expression profiling (GEP) and the genetic landscape of T-cell and NK-cell neoplasms have resulted in diagnostic, prognostic and therapeutic implications and has consequently led to revisions in the classification of PTCLs and introduction of new provisional entities. Incorporating these molecular and genetic markers, the latest 2016 WHO classification, an update of the current fourth edition, has identified 25 definitive and 6 provisional entities and further groups these lymphomas according to their usual presentation into disseminated disease (leukaemias), predominantly nodal, predominantly extra-nodal or cutaneous lymphomas. Nodal PTCLs represent the most prevalent PTCL subtype and include peripheral T-cell lymphoma - not otherwise specified (PTCL-NOS), angioimmunoblastic T-cell lymphoma (AITL), follicular T-cell lymphoma (FTCL), nodal PTCL with T follicular helper (TFH) phenotype, and systemic anaplastic large cell lymphoma (sALCL) either ALK-positive or ALK-negative. Due to better understanding of the cell of origin in T-cell lymphomas, AITL, FTCL and PTCL with TFH phenotype have been recognised as being derived from TFH cells as these neoplastic cells express at least two or three TFH cell antigens which include CD279/PD1, CD10, BCL6, CXCL13, ICOS, SAP and CCR5. In addition to the common cell of origin, these lymphomas share pathological and clinical features as well as recurrent genetic abnormalities such as mutations in TET2, IDH2, DNMT3A, RHOA and CD28, and gene fusions such as ITK-SYK and CTLA-CD28. Such features suggest that these lymphomas belong to the same spectrum of disease and are therefore now unified under a common heading. PTCL-NOS encompasses those which do not match the classification of other PTCL categories. This subtype mostly has a CD4+/CD8– phenotype and shows extreme cytological and phenotypical heterogeneity but GEP have identified at least three subgroups characterised by overexpression of GATA3, TBX21 and cytotoxic genes associated with varied clinical behaviour. Systemic ALCLs have a primarily aggressive nodal presentation and are further subdivided to ALK-positive or ALK-negative based on whether they carry the nucleoplasmin (NPM) anaplastic lymphoma kinase (ALK) translocation between chromosome 2 and 5, designated as t(2;5). We now know that a subset of ALCL, ALK-negative cases harbour rearrangement at the locus containing DUSP22 and IRF4 in chromosome 6p25, which induces down regulation of DUSP22, a dual specific phosphatase that inhibits TCR signalling. Another subset of ALK-negative ALCL has rearrangements in TP63. Clear distinction of PTCL with high CD30 expression from ALCL, ALK-negative have also been possible through GEP and as such, the former is no longer considered a provisional entity. Breast implant associated ALCL, a unique form of ALCL, ALK-negative arising in association with breast implants, is increasingly being recognised and has been included as a provisional entity. Cutaneous ALCL (cALCL), ALK-negative, the fourth subtype, is an indolent form limited to the skin. Whilst notorious to have an aggressive clinical course and an extremely poor overall prognosis, clinicians are constantly challenged with low rates of complete remissions, early relapses and failures to achieve long-term remissions despite aggressive first-line chemotherapy, in the majority of patients. More recently, improved understanding of disease biology and the molecular profile of specific PTCL subtypes has opened the door to the potential of more targeted therapies with a number of new biological therapies approved by regulatory agencies in the last few years.

Comprehensive analysis of clinical, pathological, and genomic characteristics of follicular helper T-cell derived lymphomas

BackgroundThe 2016 World Health Organization (WHO) classification introduced nodal lymphomas of T follicular helper (Tfh) cell origin, such as angioimmunoblastic T-cell lymphoma (AITL), follicular peripheral T-cell lymphoma (F-PTCL), and nodal peripheral T-cell lymphoma with T follicular helper phenotype (nodal PTCL with TFH phenotype). However, the accurate incidence rate and clinical characteristics of F-PTCL and nodal PTCL with TFH are unstudied.MethodsBetween February 2012 to June 2020, a total of 207 cases diagnosed with nodal lymphomas of T follicular helper (Tfh) cell origin and PTCL-NOS were reviewed for clinical and histopathologic data. PTCL-NOS was defined to not correlate to any of the specific entities of mature T cell lymphoma in the WHO 2016 classification. We attempted to classify PTCL-GATA3 and PTCL-TBX21 by IHC staining. Target gene analysis was performed on a few patients with sufficient blood and tissue samples additionally.ResultsAmong 207 patients, 111 patients (53.6%) had AITL, 67 patients (32.4%) had PTCL-NOS, 19 patients (9.2%) had F-PTCL, and 10 patients (4.8%) had nodal PTCL with TFH phenotype. We re-defined and analyzed F-PTCL and nodal PTCL with TFH phenotype into other TFH lymphomas. AITL (N = 101/111, 91.0%) was found to have a higher frequency of stage III/IV cancers compared to other TFH lymphomas (N = 22/29, 75.0%) and PTCL-NOS (N = 53/67, 79.1%; p-value = 0.03). The OS of AITL and other TFH lymphomas was similarly superior to PTCL-NOS (p-value = 0.02). AITL and other TFH lymphomas showed the TBX21 subtype more commonly than the GATA3 subtype. Mutations related to the RAS family (RHOA) and those related to epigenetic regulators (IDH2, DNMT3A, and TET2) were shown mainly in AITL and other TFH lymphomas.ConclusionsOther TFH lymphomas appear to be a rare disease entity around one-quarter in nodal lymphomas of T follicular helper (Tfh) cell origin. Their less aggressive clinical feature than we did not expect is utterly different from PTCL-NOS and AITL. On the other hand, other TFH lymphomas share some characteristics, such as the cell of origin, a more common TBX21 subtype, and genetic variation such as RAS family mutation and epigenetic regulators, with AITL.

Normalization of B Cell Subsets but Not T Follicular Helper Phenotypes in Infants With Very Early Antiretroviral Treatment.

Introduction: Infant HIV-1-infection is associated with high morbidity and mortality if antiretroviral treatment (ART) is not initiated promptly. We characterized development of circulating T follicular helper cells (cTfh) and their relationship to naïve/memory B cell subsets in a cohort of neonates initiating ART within the first week of life.Methods: Infants were diagnosed within 48 hours of birth and started ART as soon as possible. The frequency and phenotype of cTfh and B cells were analyzed at enrollment (birth −19 days) and at 4, 12, and 72 weeks of age in blood of 27 HIV-1-intrauterine-infected and 25 HIV-1 exposed uninfected (HEU) infants as part of a study in Johannesburg, South Africa. cTfh cells were divided into Tfh1, Tfh2, and Tfh17 subsets. B cell phenotypes were defined as naïve, resting memory, activated memory and tissue-like memory cells.Results: HIV-1-infected infants had higher frequencies of cTfh cells than HEU infants up to 12 weeks of age and these cTfh cells were polarized toward the Tfh1 subset. Higher frequencies of Tfh1 and lower frequencies of Tfh2 and Tfh17 correlated with lower CD4+ T cell percentages. Lower frequencies of resting memory, with corresponding higher frequencies of activated memory B cells, were observed with HIV-1 infection. Importantly, dysregulations in B cell, but not cTfh cell, subsets were normalized by 72 weeks.Conclusion: Very early ART initiation in HIV-1-infected infants normalizes B cell subsets but does not fully normalize perturbations in cTfh cell subsets which remain Tfh1 polarized at 72 weeks. It remains to be determined if very early ART improves vaccine antibody responses despite the cTfh and B cell perturbations observed over the time course of this study.

Characterization of HIV-induced remodeling reveals differences in infection susceptibility of memory CD4+ Tcell subsets invivo.

SUMMARYRelatively little is known about features of T cells targeted by HIV in vivo. By applying bioinformatics analysis to mass cytometry (CyTOF)-phenotyped specimens from individuals with viremia and in-vitro-infected cells from uninfected donors, we provide an atlas of the phenotypes of in vivo and in vitro HIV-susceptible cells. T helper 17 (Th17) and α4β1+ cells are preferentially targeted in vivo, whereas T effector memory (Tem), T transitional memory (Ttm), Th1, and Th1/Th17 subsets are targeted in vitro. Multiple proteins—including chemokine and cytokine receptors—are remodeled by HIV in vivo, and these changes are mostly recapitulated in vitro. HIV remodels cells to a T follicular helper (Tfh) phenotype. Using clustering, we uncover a subset of CD29-expressing, Tem-like cells that are highly susceptible to infection in vivo and in vitro and experimentally confirm that susceptibility. These studies provide an in-depth look at features of HIV-susceptible cells in individuals with viremia and demonstrate that some—but not all—HIV-susceptible cells identified in vitro effectively model in vivo susceptibility.

Early detection of T-cell lymphoma with T follicular helper phenotype by RHOA mutation analysis.

Angioimmunoblastic T-cell lymphoma (AITL) and peripheral T-cell lymphoma with T follicular helper phenotype (PTCL-TFH) are a group of complex clinicopathological entities that originate from T follicular helper cells and share a similar mutation profile. Their diagnosis is often a challenge, particularly at an early stage, because of a lack of specific histological and immunophenotypic features, paucity of neoplastic T cells and prominent polymorphous infiltrate. We investigated whether the lymphoma-associated RHOA Gly17Val (c.50G>T) mutation, occurring in 60% of cases, is present in the early “reactive” lesions, and whether mutation analysis could help to advance the early diagnosis of lymphoma. The RHOA mutation was detected by quantitative polymerase chain reaction with a locked nucleic acid probe specific to the mutation, and a further peptide nucleic acid clamp oligonucleotide to suppress the amplification of the wild-type allele. The quantitative polymerase chain reaction assay was highly sensitive and specific, detecting RHOA Gly17Val at an allele frequency of 0.03%, but not other changes in Gly17, nor in 61 controls. Among the 37 cases of AITL and PTCL-TFH investigated, RHOA Gly17Val was detected in 62.2% (23/37) of which 19 had multiple biopsies including preceding biopsies in ten and follow-up biopsies in 11 cases. RHOA Gly17Val was present in each of these preceding or follow-up biopsies including 18 specimens that showed no evidence of lymphoma by combined histological, immunophenotypic and clonality analyses. The mutation was seen in biopsies 0-26.5 months (mean 7.87 months) prior to the lymphoma diagnosis. Our results show that RHOA Gly17Val mutation analysis is valuable in the early detection of AITL and PTCL-TFH.

Clinicopathologic Features of Peripheral T-Cell Lymphoma in Sub-Saharan Africa.

Peripheral T-cell lymphomas (PTCLs) are heterogeneous, clinically aggressive, and rare. Subtype distribution varies by geographic location; however, data from sub-Saharan Africa (SSA) are lacking. We sought to elucidate clinicopathologic features of PTCL in SSA. We reviewed PTCL consultation cases from three SSA countries. PTCL subtype was determined per 2017 World Health Organization classification. Cases with sufficient material were evaluated by polymerase chain reaction for human T-cell leukemia virus type 1 (HTLV-1) and T-cell receptor γ (TCRG) rearrangement. Among 32 cases, median age was 45 years and male-to-female ratio was 1.7. Thirty (94%) of 32 cases required additional workup for subclassification. PTCL, not otherwise specified (PTCL-NOS) was the most common subtype (13/32, 41%), followed by PTCL with T-follicular helper phenotype (6/32, 19%) and systemic anaplastic large cell lymphoma (6/32, 19%). Four (16%) of 25 cases were Epstein-Barr virus positive (EBV+) (2/2 extranodal natural killer/T-cell lymphoma, 1/13 PTCL-NOS, and 1/4 angioimmunoblastic T-cell lymphoma with EBV+ immunoblasts). Two (15%) of 13 patients with PTCL-NOS were human immunodeficiency virus positive. No cases with evaluable DNA (0/15) were HTLV-1 positive, and 9 of 10 showed clonal TCRG rearrangements. In comparison to Western studies, PTCLs from SSA show similar subtype distribution and male predominance but a younger age at diagnosis. Appropriate diagnosis of PTCL requires extensive ancillary testing not readily available in low-income countries, including much of SSA.

Multi-Center Phase II Study of Oral Azacitidine (CC-486) Plus CHOP As Initial Treatment for Peripheral T-Cell Lymphoma (PTCL)

Multi-Center Phase II Study of Oral Azacitidine (CC-486) Plus CHOP As Initial Treatment for Peripheral T-Cell Lymphoma (PTCL)

T follicular helper phenotype predicts response to histone deacetylase inhibitors in relapsed/refractory peripheral T-cell lymphoma

AbstractHistone deacetylase inhibitors (HDACi) are active agents for peripheral T-cell lymphoma (PTCL). Anecdotally angioimmunoblastic T-cell lymphoma (AITL) appears to respond better than PTCL–not otherwise specified (NOS) to HDACi. The new World Health Organization classification shows that a subgroup of PTCL carries similarities in phenotype and gene expression profiling to AITL, comparable to T follicular helper (TFH) cells. The disease might behave similarly to AITL when treated with HDACi. We analyzed 127 patients with AITL or PTCL-NOS treated with HDACi at relapse as a single agent or in combination. We re-reviewed the pathology of all PTCL-NOS to identify the TFH phenotype. Patients received HDACi at relapse as a single agent in 97 cases (76%, 59 TFH, 38 non-TFH) or in combination in 30 cases (24%, 18 TFH, 12 non-TFH) including duvelisib, lenalidomide, lenalidomide plus carfilzomib, and pralatrexate. Seven PTCL-NOS had TFH phenotype; 2 PTCL-NOS were reclassified as AITL. Overall response rate (ORR) was 56.5% (28.9% complete response [CR]) in TFH and 29.4% (19.6% CR) in non-TFH phenotype patients (P = .0035), with TFH phenotype being an independent predictor of ORR (P = .009). Sixteen patients sufficiently responded to HDACi or HDACi in combination with another agent to proceed directly to allogeneic transplantation; 1 of 16 responded to donor lymphocyte infusion (12 TFH, 4 non-TFH). Our results, although retrospective, support that HDACi, as a single agent or in combination, may have superior activity in TFH-PTCL compared with non-TFH PTCL. This differential efficacy could help inform subtype-specific therapy and guide interpretation of HDACi trials.

Cytokines and transcription factors in the differentiation of CD4+ T helper cell subsets and induction of tissue inflammation and autoimmunity.

CD4+T helper (Th) cells are critical in homeostasis and host defense but are also central to the development of various autoimmune diseases if they become dysregulated. Specifically, pathogenic Th1 and Th17 cells contribute to autoimmune inflammation whereas Treg and Tr1 cells are important for maintaining immune tolerance and resolution of inflammation, respectively. Cytokines trigger signaling pathways in naive T cells that induce lineage-defining transcription factors that direct their differentiation into the distinct T helper cell subsets. It has become clear that the differentiation of T helper cells is not only influenced by the cytokine milieu but also by their metabolic state, cues from the microbiota and the tissue they reside in. A comprehensive understanding how these various stimuli contribute to T helper cell differentiation and phenotype could potentially provide novel ways for therapeutic intervention in autoimmunity and tissue inflammation.

American Registry of Pathology Expert Opinions: Recommendations for the diagnostic workup of mature T cell neoplasms

The diagnosis of T-cell lymphomas is highly challenging and requires an integrated approach in which clinical, morphologic, immunophenotypic and molecular data are incorporated into the diagnosis. Under the auspices of the American Registry of Pathology, the authors met to discuss this topic with the goal to provide practical and useful recommendations for pathologists when evaluating T-cell lymphomas. In this review, we discuss the diagnostic findings and workup for the various types of nodal T-cell lymphoma including anaplastic large cell lymphoma, nodal peripheral T-cell lymphoma not otherwise specified (PTCL-NOS), and PTCL with a T follicular helper (TFH) phenotype. We review clinicopathologic and immunophenotypic features (including flow cytometry panels) helpful in the differential diagnosis of mature T-cell lymphomas presenting in the peripheral blood and bone marrow, and we discuss some of the more common extranodal-based T-cell lymphomas including extranodal natural killer/T-cell lymphoma of nasal and non-nasal type, gamma delta T cell lymphomas, and aggressive and indolent T- and NK-lymphoproliferative disorders involving the gastrointestinal tract. Mycosis fungoides and most other cutaneous T-cell lymphomas are not the focus of this review, although the differential diagnosis of Sezary syndrome from mycosis fungoides is covered. We do not intend for these recommendations to be anything other than suggestions that will hopefully spur on additional discussion, and perhaps eventually evolve into a consensus approach for the workup of T-cell lymphomas.

IL-10-producing Tfh cells accumulate with age and link inflammation with age-related immune suppression.

Aging results in profound immune dysfunction, resulting in the decline of vaccine responsiveness previously attributed to irreversible defects in the immune system. In addition to increased interleukin-6 (IL-6), we found aged mice exhibit increased systemic IL-10 that requires forkhead box P3-negative (FoxP3-), but not FoxP3+, CD4+T cells. Most IL-10-producing cells manifested a T follicular helper (Tfh) phenotype and required the Tfh cytokines IL-6 and IL-21 for their accrual, so we refer to them as Tfh10 cells. IL-21 was also required to maintain normal serum levels of IL-6 and IL-10. Notably, antigen-specific Tfh10 cells arose after immunization of aged mice, and neutralization of IL-10 receptor signaling significantly restored Tfh-dependent antibody responses, whereas depletion of FoxP3+ regulatory and follicular regulatory cells did not. Thus, these data demonstrate that immune suppression with age is reversible and implicate Tfh10 cells as an intriguing link between "inflammaging" and impaired immune responses with age.

Skin manifestation of angioimmunoblastic T-cell lymphoma mimicking Hansen’s disease on histology

&lt;p&gt;Angioimmunoblastic T-cell lymphoma (AITL) is an uncommon form of peripheral T-cell lymphoma now put under the category of nodal T-cell lymphoma with follicular T helper phenotype. 50% cases of AITL have varied cutaneous manifestations including macules, papules, petechiae, purpura, nodules, non-specific rash and urticaria. Herein we present a case of AITL initially presented as maculopapular rash which on histology was mimicking as Hansen’s disease with perineural and perivascular inflammation; however, an infective organism could not be demonstrated. Later on, a delayed diagnosis was made on lymph node biopsy and immunohistochemistry and patient responded well with chemotherapy.&lt;/p&gt;

Genomics of Peripheral T-Cell Lymphoma and Its Implications for Personalized Medicine.

Peripheral T-cell lymphoma (PTCL) is a rare, heterogenous group of mature T-cell neoplasms that comprise 10–15% of non-Hodgkin lymphoma cases in the United States. All subtypes of PTCL, except for ALK+ anaplastic T-cell lymphoma, are associated with poor prognosis, with median overall survival (OS) rates of 1–3 years. The diagnosis of PTCL is mainly based on clinical presentation, morphologic features, and immunophenotypes. Recent advances in genome sequencing and gene expression profiling have given new insights into the pathogenesis and molecular biology of PTCL. An enhanced understanding of its genomic landscape holds the promise of refining the diagnosis, prognosis, and management of PTCL. In this review, we examine recently discovered genetic abnormalities identified by molecular profiling in 3 of the most common types of PTCL: RHOAG17V and epigenetic regulator mutations in angioimmunoblastic T-cell lymphoma, ALK expression and JAK/STAT3 pathway mutations in anaplastic T-cell lymphoma, and T-follicular helper phenotype and GATA3/TBX21 expression in PTCL-not otherwise specified. We also discuss the implications of these abnormalities for clinical practice, new/potential targeted therapies, and the role of personalized medicine in the management of PTCL.

Dissecting the mechanisms responsible for the generation of regulatory versus pathogenic human CD4+ T cells by TLR9-activated plasmacytoid dendritic cells

Abstract Understanding the mechanisms underlying how regulatory and pathogenic CD4+ T cells are generated will accelerate therapeutic target identification in autoimmune diseases such as Systemic Lupus Erythematosus (SLE), where this balance is altered. We recently described that in vitro priming of naïve CD4+ T cells with plasmacytoid DCs (pDCs) activated with either CpGA or Oxidized mitochondrial DNA (Ox mtDNA) leads to the generation of Type 1 regulatory T cells (Tr1) or of a novel T helper subset (Th10 cells), respectively. Th10 cells are expanded in the blood of SLE patients and accumulate within the tubuloinsterstitial areas of proliferative nephritis (PLN) lesions. These cells produce IL10 and mitochondrial ROS (mtROS) as the result of reverse electron transport (RET) fueled by the tricarboxylic acid (TCA) cycle intermediate succinate. Functionally, Th10 cells are not suppressive, but they provide B cell help through the synergistic effect of IL10 and succinate. The mechanisms responsible for the acquisition of a regulatory versus a helper phenotype remain elusive. We now show that naïve CD4+ T cells require the activation of the Delta Like Canonical Notch Ligand 4 (Dll4)-Notch pathway in order to acquire a TH10 phenotype. Conversely, the TLR7-IRF7 pathway is necessary for the generation of Tr1 cells. These pathways might be exploited for therapeutic intervention in human autoimmune diseases.

Phenotype and functionality of follicular helper T cells in patients with acute dengue infection

BackgroundThe association of functionality and phenotype of follicular helper T cells (Tfh) with dengue virus (DENV) specific antibody responses and clinical disease severity has not been well studied.MethodsWe investigated the phenotype and functionality of Tfh cells and plasmablasts in adult patients (DF = 18, DHF = 22) with acute dengue (day 4 to 8 since onset of fever) of varying severity using multiparametric flowcytometry. The properties of Tfh cells were correlated with viraemia, disease severity, plasmablast responses and DENV-specific serum antibody responses. We further evaluated the kinetics of neutralizing antibodies (Neut50) throughout the course of illness in order to evaluate their association with clinical disease severity and viraemia.ResultsTfh cells (especially those producing IL-21 and co-expressing PD-1 and ICOS) were found to be significantly expanded (p < 0.0001) and highly activated in patients with DHF compared to those with DF. The frequency of Tfh cells significantly correlated with DENV-specific IgG, NS1-specific antibodies and Neut50 antibody titres in patients with DHF but not in those with DF. Although the Neut50 titres increased during the course of acute secondary DENV infection, they showed differences based on serotype. For instance, the Neut50 titres were significantly higher during the latter part of illness in patients with DF compared to DHF in DENV1 infection, while in DENV2, patients with DHF had significantly higher titres. The viral loads during early illness did not correlate with the subsequent rise in the Neut50 antibody titres during any time point of illness.ConclusionsThe expansion of Tfh cells is associated with DHF and DENV-specific IgG, NS1-specific and neutralizing antibodies. Neut50 titres did not associate with disease severity or viraemia at the point of first presentation during the febrile phase, but later titres do show differential association with severity in patients with DENV1 compared to DENV2.