Phospho-mimetic CD3ε variants prevent TCR and CAR signaling.

Introduction: Antigen binding to the T cell antigen receptor (TCR) leads to the phosphorylation of the immunoreceptor tyrosine-based activation motifs (ITAMs) of the CD3 complex, and thereby to T cell activation. The CD3ε subunit plays a unique role in TCR activation by recruiting the kinase LCK and the adaptor protein NCK prior to ITAM phosphorylation. Here, we aimed to investigate how phosphorylation of the individual CD3ε ITAM tyrosines impacts the CD3ε signalosome. Methods: We mimicked irreversible tyrosine phosphorylation by substituting glutamic acid for the tyrosine residues in the CD3ε ITAM. Results: Integrating CD3ε phospho-mimetic variants into the complete TCR-CD3 complex resulted in reduced TCR signal transduction, which was partially compensated by the involvement of the other TCR-CD3 ITAMs. By using novel CD3ε phospho-mimetic Chimeric Antigen Receptor (CAR) variants, we avoided any compensatory effects of other ITAMs in the TCR-CD3 complex. We demonstrated that irreversible CD3ε phosphorylation prevented signal transduction upon CAR engagement. Mechanistically, we demonstrated that glutamic acid substitution at the N-terminal tyrosine residue of the CD3ε ITAM (Y39E) significantly reduces NCK binding to the TCR. In contrast, mutation at the C-terminal tyrosine of the CD3ε ITAM (Y50E) abolished LCK recruitment to the TCR, while increasing NCK binding. Double mutation at the C- and N-terminal tyrosines (Y39/50E) allowed ZAP70 to bind, but reduced the interaction with LCK and NCK. Conclusions: The data demonstrate that the dynamic phosphorylation of the CD3ε ITAM tyrosines is essential for CD3ε to orchestrate optimal TCR and CAR signaling and highlights the key role of CD3ε signalosome to tune signal transduction.

Enhancing the Antitumor Immunity of T Cells by Engineering the Lipid-Regulatory Site of the TCR/CD3 Complex.

The engagement of the T-cell receptor (TCR) by a specific peptide-MHC ligand initiates transmembrane signaling to induce T-cell activation, a key step in most adaptive immune responses. Previous studies have indicated that TCR signaling is tightly regulated by cholesterol and its sulfate metabolite, cholesterol sulfate (CS), on the membrane. Here, we report a novel mechanism by which CS modulates TCR signaling through a conformational change of CD3 subunits. We found that the negatively charged CS interacted with the positively charged cytoplasmic domain of CD3ε (CD3εCD) to enhance its binding to the cell membrane and induce a stable secondary structure. This secondary structure suppressed the release of CD3εCD from the membrane in the presence of Ca2+, which in turn inhibited TCR phosphorylation and signaling. When a point mutation (I/A) was introduced to the intracellular immunoreceptor tyrosine-based activation motifs (YxxI-x6-8-YxxL) of CD3ε subunit, it reduced the stability of the secondary structure and regained sensitivity to Ca2+, which abolished CS-mediated inhibition and enhanced the signaling of the TCR complex. Notably, the I/A mutation could be applied to both murine and human TCR-T cell therapy to improve the antitumor efficacy. Our study reveals insights into the regulatory mechanism of TCR signaling and provides a strategy to functionally engineer the TCR/CD3 complex for T cell-based cancer immunotherapy.

Cholesterol inhibits TCR signaling by directly restricting TCR-CD3 core tunnel motility.

Cholesterol molecules specifically bind to the resting αßTCR to inhibit cytoplasmic CD3ζ ITAM phosphorylation through sequestering the TCR-CD3 complex in an inactive conformation. The mechanisms of cholesterol-mediated inhibition of TCR-CD3 and its activation remain unclear. Here, we present cryoelectron microscopy structures of cholesterol- and cholesterol sulfate (CS)-inhibited TCR-CD3 complexes and an auto-active TCR-CD3 variant. The structures reveal that cholesterol molecules act like a latch to lock CD3ζ into an inactive conformation in the membrane. Mutations impairing binding of cholesterol molecules to the tunnel result in the movement of the proximal C terminus of the CD3ζ transmembrane helix, thereby activating the TCR-CD3 complex in human cells. Together, our data reveal the structural basis of TCR inhibition by cholesterol, illustrate how the cholesterol-binding tunnel is allosterically coupled to TCR triggering, and lay a foundation for the development of immunotherapies through directly targeting the TCR-CD3 complex.

CD137 (4-1BB) costimulation of CD8+ T cells is more potent when provided in cis than in trans with respect to CD3-TCR stimulation.

CD137 (4-1BB; TNFSR9) is an activation-induced surface receptor that through costimulation effects provide antigen-primed T cells with augmented survival, proliferation and effector functions as well as metabolic advantages. These immunobiological mechanisms are being utilised for cancer immunotherapy with agonist CD137-binding and crosslinking-inducing agents that elicit CD137 intracellular signaling. In this study, side-by-side comparisons show that provision of CD137 costimulation in-cis with regard to the TCR-CD3-ligating cell is superior to that provided in-trans in terms of T cell activation, proliferation, survival, cytokine secretion and mitochondrial fitness in mouse and human. Cis ligation of CD137 relative to the TCR-CD3 complex results in more intense canonical and non-canonical NF-κB signaling and provides a more robust induction of cell cycle and DNA damage repair gene expression programs. Here we report that the superiority of cis versus trans CD137-costimulation is readily observed in vivo and is relevant for understanding the immunotherapeutic effects of CAR T cells and CD137 agonistic therapies currently undergoing clinical trials, which may provide costimulation either in cis or in trans.

Predicting antigen specificity of single T cells based on TCR CDR3 regions.

It has recently become possible to simultaneously assay T-cell specificity with respect to large sets of antigens and the T-cell receptor sequence in high-throughput single-cell experiments. Leveraging this new type of data, we propose and benchmark a collection of deep learning architectures to model T-cell specificity in single cells. In agreement with previous results, we found that models that treat antigens as categorical outcome variables outperform those that model the TCR and antigen sequence jointly. Moreover, we show that variability in single-cell immune repertoire screens can be mitigated by modeling cell-specific covariates. Lastly, we demonstrate that the number of bound pMHC complexes can be predicted in a continuous fashion providing a gateway to disentangle cell-to-dextramer binding strength and receptor-to-pMHC affinity. We provide these models in the Python package TcellMatch to allow imputation of antigen specificities in single-cell RNA-seq studies on T cells without the need for MHC staining.

Lck bound to coreceptor is less active than free Lck.

Src family kinase Lck plays critical roles during T cell development and activation, as it phosphorylates the TCR/CD3 complex to initiate TCR signaling. Lck is present either in coreceptor-bound or coreceptor-unbound (free) forms, and we here present evidence that the two pools of Lck have different molecular properties. We discovered that the free Lck fraction exhibited higher mobility than CD8α-bound Lck in OT-I T hybridoma cells. The free Lck pool showed more activating Y394 phosphorylation than the coreceptor-bound Lck pool. Consistent with this, free Lck also had higher kinase activity, and free Lck mediated higher T cell activation as compared to coreceptor-bound Lck. Furthermore, the coreceptor-Lck coupling was independent of TCR activation. These findings give insights into the initiation of TCR signaling, suggesting that changes in coreceptor-Lck coupling constitute a mechanism for regulation of T cell sensitivity.

Actin polymerization regulates recruitment of Nck to CD3ε upon T-cell receptor triggering.

Following T-cell antigen receptor (TCR) engagement, rearrangement of the actin cytoskeleton supports intracellular signal transduction and T-cell activation. The non-catalytic region of the tyrosine kinase (Nck) molecule is an adapter protein implicated in TCR-induced actin polymerization. Further, Nck is recruited to the CD3ε subunit of the TCR upon TCR triggering. Here we examine the role of actin polymerization in the recruitment of Nck to the TCR. To this end, Nck binding to CD3ε was quantified in Jurkat cells using the proximity ligation assay. We show that inhibition of actin polymerization using cytochalasin D delayed the recruitment of Nck1 to the TCR upon TCR triggering. Interestingly, CD3ε phosphorylation was also delayed. These findings suggest that actin polymerization promotes the recruitment of Nck to the TCR, enhancing downstream signaling, such as phosphorylation of CD3ε.

A scoring system for the electrostatic complementarities of T-cell receptors and cancer-mutant amino acids: multi-cancer analyses of associated survival rates.

The anti-tumor immune response is considered to be due to the T-cell receptor (TCR) binding to tumor antigens, which can be either wild-type, early stem cell proteins, presumably foreign to a developed immune system; or mutant peptides, foreign to the immune system because of a mutant amino acid (aa) or otherwise somatically altered aa sequence. Recently, very large numbers of TCR complementarity-determining region-3 (CDR3) aa sequences obtained from tumor specimens have become available. We developed a novel algorithm for assessing the complementarity of tumor mutant peptides and TCR CDR3s, based on the retrieval of TCR CDR3 aa sequences from both tumor specimen and patient blood exomes and by using an automated process of assessing CDR3 and mutant aa electrical charges. Results indicated many instances where high electrostatic complementarity was associated with a higher survival rate. In particular, our approach led to the identification of specific genes contributing significantly to the complementary, TCR CDR3-mutant aa. These results suggest a novel approach to tumor immunoscoring and may lead to the identification of high-priority neo-antigen, peptide vaccines; or to the identification of ex vivo stimulants of tumor-infiltrating lymphocytes.

Ultrasmall silica nanoparticles directly ligate the T cell receptor complex.

The impact of ultrasmall nanoparticles (<10-nm diameter) on the immune system is poorly understood. Recently, ultrasmall silica nanoparticles (USSN), which have gained increasing attention for therapeutic applications, were shown to stimulate T lymphocytes directly and at relatively low-exposure doses. Delineating underlying mechanisms and associated cell signaling will hasten therapeutic translation and is reported herein. Using competitive binding assays and molecular modeling, we established that the T cell receptor (TCR):CD3 complex is required for USSN-induced T cell activation, and that direct receptor complex-particle interactions are permitted both sterically and electrostatically. Activation is not limited to αß TCR-bearing T cells since those with γδ TCR showed similar responses, implying that USSN mediate their effect by binding to extracellular domains of the flanking CD3 regions of the TCR complex. We confirmed that USSN initiated the signaling pathway immediately downstream of the TCR with rapid phosphorylation of both ζ-chain-associated protein 70 and linker for activation of T cells protein. However, T cell proliferation or IL-2 secretion were only triggered by USSN when costimulatory anti-CD28 or phorbate esters were present, demonstrating that the specific impact of USSN is in initiation of the primary, nuclear factor of activated T cells-pathway signaling from the TCR complex. Hence, we have established that USSN are partial agonists for the TCR complex because of induction of the primary T cell activation signal. Their ability to bind the TCR complex rapidly, and then to dissolve into benign orthosilicic acid, makes them an appealing option for therapies targeted at transient TCR:CD3 receptor binding.

Compositional characteristics of human peripheral TRBV pseudogene rearrangements.

The diversity of the T cell receptor (TCR) complementarity determining region 3 (CDR3) repertoire is the result of random combinations, insertions and deletions during recombination of the germline V, D and J gene fragments. During evolution, some human TCR beta chain variable (TRBV) pseudogenes have been retained. Many previous studies have focused on functional TRBV genes, while little attention has been given to TRBV pseudogenes. To describe the compositional characteristics of TRBV pseudogene rearrangements, we compared and analysed TRBV pseudogenes, TRBV open reading frames (ORFs) and functional TRBV genes via high-throughput sequencing of DNA obtained from the peripheral blood of 4 healthy volunteers and 4 patients. Our results revealed several differences in J and D gene usage. The V deletion distribution profile of the pseudogenes was significantly different from that of the ORFs and functional genes. In addition, arginine, lysine and cysteine were more frequently used in putative CDR3 pseudogene rearrangements, while functional rearrangements used more leucine. This study presents a comprehensive description of the compositional characteristics of peripheral TRBV pseudogene rearrangements, which will provide a reference for further research on TRBV pseudogenes.

Adoptive Transfer of mRNA-Transfected T Cells Redirected against Diabetogenic CD8 T Cells Can Prevent Diabetes.

Chimeric major histocompatibility complex (MHC) molecules supplemented with T cell receptor (TCR) signaling motifs function as activation receptors and can redirect gene-modified T cells against pathogenic CD8 T cells. We have shown that ß2 microglobulin (ß2m) operates as a universal signaling component of MHC-I molecules when fused with the CD3-ζ chain. Linking the H-2Kd-binding insulin B chain peptide insulin B chain, amino acids 15-23 (InsB15-23) to the N terminus of ß2m/CD3-ζ, redirected polyclonal CD8 T cells against pathogenic CD8 T cells in a peptide-specific manner in the non-obese diabetic (NOD) mouse. Here, we describe mRNA electroporation for delivering peptide/ß2m/CD3-ζ genes to a reporter T cell line and purified primary mouse CD8 T cells. The peptide/ß2m/CD3-ζ products paired with endogenous MHC-I chains and transmitted strong activation signals upon MHC-I cross-linking. The reporter T cell line transfected with InsB15-23/ß2m/CD3-ζ mRNA was activated by an InsB15-23-H-2Kd-specific CD8 T cell hybrid only when the transfected T cells expressed H-2Kd. Primary NOD CD8 T cells expressing either InsB15-23/ß2m/CD3-ζ or islet-specific glucose-6-phosphatase catalytic subunit-related protein, amino acids 206-214 (IGRP206-214)/ß2m/CD3-ζ killed their respective autoreactive CD8 T cell targets in vitro. Furthermore, transfer of primary CD8 T cells transfected with InsB15-23/ß2m/CD3-ζ mRNA significantly reduced insulitis and protected NOD mice from diabetes. Our results demonstrate that mRNA encoding chimeric MHC-I receptors can redirect effector CD8 against diabetogenic CD8 T cells, offering a new approach for the treatment of type 1 diabetes.

Phase 1 Results of ZUMA-1: A Multicenter Study of KTE-C19 Anti-CD19 CAR T Cell Therapy in Refractory Aggressive Lymphoma.

Outcomes for patients with refractory diffuse large B cell lymphoma (DLBCL) are poor. In the multicenter ZUMA-1 phase 1 study, we evaluated KTE-C19, an autologous CD3ζ/CD28-based chimeric antigen receptor (CAR) T cell therapy, in patients with refractory DLBCL. Patients received low-dose conditioning chemotherapy with concurrent cyclophosphamide (500 mg/m2) and fludarabine (30 mg/m2) for 3 days followed by KTE-C19 at a target dose of 2 × 106 CAR T cells/kg. The incidence of dose-limiting toxicity (DLT) was the primary endpoint. Seven patients were treated with KTE-C19 and one patient experienced a DLT of grade 4 cytokine release syndrome (CRS) and neurotoxicity. Grade ≥3 CRS and neurotoxicity were observed in 14% (n = 1/7) and 57% (n = 4/7) of patients, respectively. All other KTE-C19-related grade ≥3 events resolved within 1 month. The overall response rate was 71% (n = 5/7) and complete response (CR) rate was 57% (n = 4/7). Three patients have ongoing CR (all at 12+ months). CAR T cells demonstrated peak expansion within 2 weeks and continued to be detectable at 12+ months in patients with ongoing CR. This regimen of KTE-C19 was safe for further study in phase 2 and induced durable remissions in patients with refractory DLBCL.

A conserved αß transmembrane interface forms the core of a compact T-cell receptor-CD3 structure within the membrane.

The T-cell antigen receptor (TCR) is an assembly of eight type I single-pass membrane proteins that occupies a central position in adaptive immunity. Many TCR-triggering models invoke an alteration in receptor complex structure as the initiating event, but both the precise subunit organization and the pathway by which ligand-induced alterations are transferred to the cytoplasmic signaling domains are unknown. Here, we show that the receptor complex transmembrane (TM) domains form an intimately associated eight-helix bundle organized by a specific interhelical TCR TM interface. The salient features of this core structure are absolutely conserved between αß and γδ TCR sequences and throughout vertebrate evolution, and mutations at key interface residues caused defects in the formation of stable TCRαß:CD3δε:CD3γε:ζζ complexes. These findings demonstrate that the eight TCR-CD3 subunits form a compact and precisely organized structure within the membrane and provide a structural basis for further investigation of conformationally regulated models of transbilayer TCR signaling.

A novel Dock8 gene mutation confers diabetogenic susceptibility in the LEW.1AR1/Ztm-iddm rat, an animal model of human type 1 diabetes.

AIMS/HYPOTHESIS: The LEW.1AR1-iddm rat, an animal model of human type 1 diabetes, arose through a spontaneous mutation within the inbred strain LEW.1AR1. A susceptibility locus (Iddm8) on rat chromosome 1 (RNO1) has been identified previously, which is accompanied by autoimmune diabetes and the additional phenotype of a variable CD3(+) T cell frequency. METHODS: In the present study we characterised the Iddm8 region on RNO1 in backcross strains using the genetically divergent Brown Norway (BN) and Paris (PAR) rats. Candidate genes of the Iddm8 region were sequenced for mutation analysis. RESULTS: The Iddm8 region could be subdivided by single nucleotide polymorphism (SNP) analyses. In the first region, a mutation in exon 44 of the Dock8 gene was identified resulting in an amino acid exchange in the protein from glutamine to glutamate. This exchange is unique for the LEW.1AR1-iddm rat. In the second region, a SNP was detected in exon 11 of the Vwa2 gene with an exchange from arginine to tryptophan. This SNP is also present in other rat strains. CONCLUSIONS/INTERPRETATION: The Dock8 mutation gave rise to a new type 1 diabetes rat model with very close similarity to type 1 diabetes in humans, providing a deepened insight into the impact of genes involved in diabetes development.

High-throughput sequencing reveals an altered T cell repertoire in X-linked agammaglobulinemia.

To examine the T cell receptor structure in the absence of B cells, the TCR ß CDR3 was sequenced from DNA of 15 X-linked agammaglobulinemia (XLA) subjects and 18 male controls, using the Illumina HiSeq platform and the ImmunoSEQ analyzer. V gene usage and the V-J combinations, derived from both productive and non-productive sequences, were significantly different between XLA samples and controls. Although the CDR3 length was similar for XLA and control samples, the CDR3 region of the XLA T cell receptor contained significantly fewer deletions and insertions in V, D, and J gene segments, differences intrinsic to the V(D)J recombination process and not due to peripheral T cell selection. XLA CDR3s demonstrated fewer charged amino acid residues, more sharing of CDR3 sequences, and almost completely lacked a population of highly modified Vß gene segments found in control DNA, suggesting both a skewed and contracted T cell repertoire in XLA.

Identification of the Docking Site for CD3 on the T Cell Receptor ß Chain by Solution NMR.

The T cell receptor (TCR)-CD3 complex is composed of a genetically diverse αß TCR heterodimer associated noncovalently with the invariant CD3 dimers CD3ϵγ, CD3ϵδ, and CD3ζζ. The TCR mediates peptide-MHC recognition, whereas the CD3 molecules transduce activation signals to the T cell. Although much is known about downstream T cell signaling pathways, the mechanism whereby TCR engagement by peptide-MHC initiates signaling is poorly understood. A key to solving this problem is defining the spatial organization of the TCR-CD3 complex and the interactions between its subunits. We have applied solution NMR methods to identify the docking site for CD3 on the ß chain of a human autoimmune TCR. We demonstrate a low affinity but highly specific interaction between the extracellular domains of CD3 and the TCR constant ß (Cß) domain that requires both CD3ϵγ and CD3ϵδ subunits. The mainly hydrophilic docking site, comprising 9-11 solvent-accessible Cß residues, is relatively small (∼400 Å(2)), consistent with the weak interaction between TCR and CD3 extracellular domains, and devoid of glycosylation sites. The docking site is centered on the αA and αB helices of Cß, which are located at the base of the TCR. This positions CD3ϵγ and CD3ϵδ between the TCR and the T cell membrane, permitting us to distinguish among several possible models of TCR-CD3 association. We further correlate structural results from NMR with mutational data on TCR-CD3 interactions from cell-based assays.

Dendritic cells promote expansion and survival of aberrant TCR-negative intraepithelial lymphocyte lines from refractory celiac disease type II patients.

Celiac disease (CD) patients who fail to respond to a gluten-free diet suffer from refractory celiac disease (RCD). A marked expansion of intraepithelial lymphocytes (IEL) lacking surface TCR/CD3 expression characterizes the RCD subtype II. In up to 50% of RCDII patients these so-called aberrant IEL (a-IEL) develop into lymphoma and can disseminate into other tissues. Elevated levels of Interleukin-15 (IL-15) in the intestine of CD and RCD patients likely contribute to the expansion of a-IEL. Here, we investigated if interactions with other cells might also influence a-IEL expansion. Similar to IL-15, cells from the monocyte lineage, particularly mature dendritic cells (DCs), promoted proliferation, prevented apoptosis and induced IFNγ secretion of a-IEL derived from RCDII biopsies (RCDII cell lines), which in turn induced CXCL10. In contrast to IL-15, mature DCs did not induce proliferation of regular TCR(+)IEL lines, generated from CD biopsies and IL-15-blocking antibodies did not inhibit DC-induced proliferation of RCDII cell lines. Furthermore, proliferation was dependent on cell-cell contact, but independent of the HLA-genotype of the stimulating cells. Our results suggest that contact with DC, either in the epithelium or upon dissemination, contributes to uncontrolled expansion of a-IEL in RCDII, independent of HLA-genotype and IL-15.

Guanylate binding protein 1-mediated interaction of T cell antigen receptor signaling with the cytoskeleton.

GTPases act as important switches in many signaling events in cells. Although small and heterotrimeric G proteins are subjects of intensive studies, little is known about the large IFN-inducible GTPases. In this article, we show that the IFN-γ-inducible guanylate binding protein 1 (GBP-1) is a regulator of T cell activation. Silencing of GBP-1 leads to enhanced activation of early T cell Ag receptor/CD3 signaling molecules, including Lck, that is translated to higher IL-2 production. Mass spectrometry analyses showed that regulatory cytoskeletal proteins, like plastin-2 that bundles actin fibers and spectrin ß-chain, brain 1 that links the plasma membrane to the actin cytoskeleton, are binding partners of GBP-1. The spectrin cytoskeleton influences cell spreading and surface expression of TCR/CD3 and the leukocyte phosphatase CD45. We found higher cell spreading and enhanced surface expression of TCR/CD3 and CD45 in GBP-1 silenced T cells that explain their enhanced TCR/CD3 signaling. We conclude that GBP-1 is a downstream processor of IFN-γ via which T cells regulate cytoskeleton-dependent cell functions.

T cells redirected by a CD3ζ chimeric antigen receptor can establish self-antigen-specific tumour protection in the long term.

A majority of cancer deaths are because of an uncontrolled relapse of the disease despite initial remission after therapy, asking for strategies to control tumour cells in the long term. Adoptive therapy with chimeric antigen receptor (CAR)-redirected T cells showed promising success in primary tumour elimination; the capacity of such engineered T cells to establish enduring tumour protection is currently a matter of discussion, in particular as most targeted 'tumour-associated antigens' are self-antigens. To address the issue in a clinically relevant model that closely mimics the human situation, we recorded rejection of carcinoembryonic antigen (CEA)-positive pancreatic tumours in the CEA transgenic mouse that expressed CEA as self-antigen in healthy cells of the gastrointestinal tract. Adoptive therapy with CD8(+) T cells, which were redirected by a CEA-specific, low-affinity CAR with CD3ζ endodomain, eliminated CEA(+) tumours in a primary response; cured mice produced an efficient recall response in the long term towards CEA(+) tumour cells upon rechallenge. Secondary tumour rejection was CEA specific, mediated by engineered T cells and did not require host T cells. No toxicity towards healthy tissues with CEA expression was recorded. Data indicate that adoptive therapy with engineered T cells can establish self-antigen-specific tumour protection in the long term without autoimmunity.

CD3-T cell receptor co-stimulation through SLAMF3 and SLAMF6 receptors enhances RORγt recruitment to the IL17A promoter in human T lymphocytes.

Th17 lymphocytes play a key role during immune responses against bacteria and fungi and are involved in the pathophysiology of multiple autoimmune disorders. The co-stimulatory molecules SLAMF3 and SLAMF6 have been implicated in the formation of Th17 phenotypes and IL-17A expression. Increased surface expression of SLAMF3 and SLAMF6 has been linked with disease activity in systemic lupus erythematosus. Here we demonstrate that in human total T lymphocytes the canonical CD28 and the non-canonical SLAMF3/SLAMF6 co-stimulatory pathways cooperate in the recruitment of the transcription factor NFAT1 to the IL17A promoter. Furthermore, the dominance of the SLAMF3/SLAMF6 pathway in inducing IL-17A production can be attributed to an increased nuclear abundance and recruitment of RORγt to the IL17A promoter. Thus, we have identified an additional mechanism that may be central for the specific control of IL17A gene regulation in systemic lupus erythematosus T lymphocytes.