Structure and Function of an Inflammatory Cytokine, Interleukin-2, Analyzed Using the Bioinformatic Approach.

The inflammatory cytokine, interleukin-2 (IL-2), is an important regulator of cellular functions. This relatively less studied member of the interleukin protein family is responsible for multiple immuno-modulatory and immuno-stimulatory tasks, like T cell activation, triggering of natural killer cells, inflammation, as well as proliferation and progression of autoimmune diseases and cancers. In this communication we report the temporally variant structural aspects of the IL-2 ligand and its receptor interfaces, based on the available crystal structures. The intended goal of this effort is to generate simulated results that could potentially aid the designs of novel structure based therapeutics.

Autotaxin and soluble IL-2 receptor concentrations in cerebrospinal fluids are useful for the diagnosis of central nervous system invasion caused by haematological malignancies.

BACKGROUND: Invasion of the central nervous system by haematological malignancies is diagnosed by cytological analyses of cerebrospinal fluid or diagnostic imaging, while quantitative biomarkers for central nervous system invasion are not available and needed to be developed. METHODS: In this study, we measured the concentrations of autotaxin and soluble IL-2 receptor in cerebrospinal fluid and evaluated their usefulness as biomarkers for central nervous system invasion. RESULTS: We observed that both the autotaxin and soluble IL-2 receptor concentrations in cerebrospinal fluid were higher in subjects with central nervous system invasion than in those without, and the cerebrospinal fluid concentrations were independent from the serum concentrations of these biomarkers. ROC analyses revealed that the soluble IL-2 receptor concentration in cerebrospinal fluid was a strong discriminator of central nervous system invasion in subjects with haematological malignancies, while the autotaxin concentration in cerebrospinal fluid also had a strong ability to discriminate central nervous system invasion when the subjects were limited to those with lymphoma. The combined measurement of autotaxin and soluble IL-2 receptor in cerebrospinal fluid improved the sensitivity without notably reducing the specificity for central nervous system invasion in subjects with lymphoma when central nervous system invasion was diagnosed in cases where either value was beyond the respective cut-off value. CONCLUSION: These results suggest the possible usefulness of soluble IL-2 receptor and autotaxin concentrations in cerebrospinal fluid for the diagnosis of central nervous system invasion.

YKL-40, Soluble IL-2 Receptor, Angiotensin Converting Enzyme and C-Reactive Protein: Comparison of Markers of Sarcoidosis Activity.

The aims of this study were to describe the clinical, radiological and immunological features of a population of sarcoidosis patients and to analyse chitinase-3-like protein 1 (YKL-40), soluble interleukin-2 receptor (sIL-2R), neopterin concentrations and adenosine deaminase (ADA) activity in serum of these patients in order to understand their potential as disease markers. Fifty-nine patients affected by chronic sarcoidosis, in active (20 patients) and inactive (39 patients) phase according to the clinical, radiological and laboratory criteria were studied. Serum YKL-40, sIL-2R, high-sensitive C-reactive protein (hs-CRP), neopterin levels and ADA activities were evaluated and compared with those of 25 healthy controls. Individuals with chronic sarcoidosis were significantly higher serum YKL-40, sIL-2R, neopterin, hs-CRP concentrations, angiotensin converting enzyme (ACE) and ADA activity than those of control subjects. Sarcoidosis patients in the active phase of the disease were significantly higher YKL-40, sIL-2R, hs-CRP levels and ACE activity than those in the inactive phase, while ADA activities and neopterin levels did not display any significant difference between the active and inactive disease groups. In comparison to the other parameters, as panel measurement of the serum YKL-40, sIL-2R, ACE and hs-CRP indicate a greater discrimination between active and inactive disease. The results indicate that serum YKL-40, sIL-2R, ACE and hs-CRP concentrations may be useful marker for monitoring sarcoidosis disease activity.

UBQLN4 recognizes mislocalized transmembrane domain proteins and targets these to proteasomal degradation.

The majority of transmembrane proteins are integrated into the endoplasmic reticulum (ER) by virtue of a signal sequence-mediated co-translational process. However, a substantial portion of transmembrane proteins fails to reach the ER, leading to mislocalized cytosolic polypeptides. Their appropriate recognition and removal are of the utmost importance to avoid proteotoxic stress. Here, we identified UBQLN4 as a BAG6-binding factor that eliminates newly synthesized defective polypeptides. Using a truncated transmembrane domain protein whose degradation occurs during a pre-ER incorporation process as a model, we show that UBQLN4 recognizes misassembled proteins in the cytoplasm and targets these to the proteasome. We suggest that the exposed transmembrane segment of the defective polypeptides is essential for the UBQLN4-mediated substrate discrimination. Importantly, UBQLN4 recognizes not only the defective model substrate but also a pool of endogenous defective proteins that were induced by the depletion of the SRP54 subunit of the signal recognition particle. This study identifies a novel quality control mechanism for newly synthesized and defective transmembrane domain polypeptides that fail to reach their correct destination at the ER membrane.

NKTR-214, an Engineered Cytokine with Biased IL2 Receptor Binding, Increased Tumor Exposure, and Marked Efficacy in Mouse Tumor Models.

PURPOSE: Aldesleukin, recombinant human IL2, is an effective immunotherapy for metastatic melanoma and renal cancer, with durable responses in approximately 10% of patients; however, severe side effects limit maximal dosing and thus the number of patients able to receive treatment and potential cure. NKTR-214 is a prodrug of conjugated IL2, retaining the same amino acid sequence as aldesleukin. The IL2 core is conjugated to 6 releasable polyethylene glycol (PEG) chains. In vivo, the PEG chains slowly release to generate active IL2 conjugates. EXPERIMENTAL DESIGN: We evaluated the bioactivity and receptor binding of NKTR-214 and its active IL2 conjugates in vitro; the tumor immunology, tumor pharmacokinetics, and efficacy of NKTR-214 as a single agent and in combination with anti-CTLA-4 antibody in murine tumor models. Tolerability was evaluated in non-human primates. RESULTS: In a murine melanoma tumor model, the ratio of tumor-killing CD8(+) T cells to Foxp3(+) regulatory T cells was greater than 400 for NKTR-214 compared with 18 for aldesleukin, supporting preferential activation of the IL2 receptor beta over IL2 receptor alpha, due to the location of PEG molecules. NKTR-214 provides a 500-fold greater exposure of the tumor to conjugated IL2 compared with aldesleukin. NKTR-214 showed efficacy as a single agent and provided durable immunity that was resistant to tumor rechallenge in combination with anti-CTLA-4 antibody. NKTR-214 was well tolerated in non-human primates. CONCLUSIONS: These data support further evaluation of NKTR-214 in humans for a variety of tumor types, adding to the repertoire of potent and potentially curative cancer immunotherapies.

99mTc-anti-TNF-α antibody for the imaging of disease activity in pulmonary sarcoidosis.

Infliximab, a monoclonal antibody directed against tumour necrosis factor (TNF)-α, is used in the treatment of refractory sarcoidosis. However, the clinical response is variable and a tool to select responders beforehand is highly desirable. In this study we evaluated scintigraphy with technetium-99m ((99m)Tc)-labelled infliximab for the imaging of disease activity in patients with pulmonary sarcoidosis.10 patients were studied using single photon emission computed tomography/computed tomography (CT) 6 h and 20 h after intravenous administration of 370 MBq of(99m)Tc-infliximab. Correlation analysis was performed between tissue accumulation of(99m)Tc-infliximab and laboratory parameters (including soluble interleukin-2 receptor and angiotensin-converting enzyme), lung function parameters (including forced expiratory volume in 1 s and the diffusing capacity of the lung for carbon monoxide) and(18)F-fluorodeoxyglucose (FDG) positron emission tomography (PET)/CT.Analysis showed selective and variable accumulation of(99m)Tc-infliximab in the target tissue. Accumulation correlated positively with all four laboratory parameters and negatively with all four lung function parameters, yielding better correlations than serum TNF-α levels or(18)F-FDG PET/CT.(99m)Tc-infliximab accumulation reflects thein situTNF-α expression in an individual patient and therefore provides valuable information on the presence of the biological target for anti-TNF-α therapy.

Interleukin-2 activity can be fine tuned with engineered receptor signaling clamps.

Interleukin-2 (IL-2) regulates lymphocyte function by signaling through heterodimerization of the IL-2Rß and γc receptor subunits. IL-2 is of considerable therapeutic interest, but harnessing its actions in a controllable manner remains a challenge. Previously, we have engineered an IL-2 "superkine" with enhanced affinity for IL-2Rß. Here, we describe next-generation IL-2 variants that function as "receptor signaling clamps." They retained high affinity for IL-2Rß, inhibiting binding of endogenous IL-2, but their interaction with γc was weakened, attenuating IL-2Rß-γc heterodimerization. These IL-2 analogs acted as partial agonists and differentially affected lymphocytes poised at distinct activation thresholds. Moreover, one variant, H9-RETR, antagonized IL-2 and IL-15 better than blocking antibodies against IL-2Rα or IL-2Rß. Furthermore, this mutein prolonged survival in a model of graft-versus-host disease and blocked spontaneous proliferation of smoldering adult T cell leukemia (ATL) T cells. This receptor-clamping approach might be a general mechanism-based strategy for engineering cytokine partial agonists for therapeutic immunomodulation.

Dendritic cell derived IL-2 inhibits survival of terminally mature cells via an autocrine signaling pathway.

DCs are crucial for sensing pathogens and triggering immune response. Upon activation by pathogen-associated molecular pattern (PAMP) ligands, GM-CSF myeloid DCs (GM-DCs) secrete several cytokines, including IL-2. DC IL-2 has been shown to be important for innate and adaptive immune responses; however, IL-2 importance in DC physiology has never been demonstrated. Here, we show that autocrine IL-2 signaling is functional in murine GM-DCs in an early time window after PAMPs stimulation. IL-2 signaling selectively activates the JAK/STAT5 pathway by assembling holo-receptor complexes at the cell surface. Using the sensitivity of targeted mass spectrometry, we show conclusively that GM-DCs express CD122, the IL-2 receptor ß-chain, at steady state. In myeloid DCs, this cytokine pathway inhibits survival of PAMP-matured GM-DCs which is crucial for maintaining immune tolerance and preventing autoimmunity. Our findings suggest that immune regulation by this novel autocrine signaling pathway can potentially be used in DC immunotherapy.

An agonistic anti-BTLA mAb (3C10) induced generation of IL-10-dependent regulatory CD4+ T cells and prolongation of murine cardiac allograft.

BACKGROUND: The co-inhibitory receptor B and T lymphocyte attenuator (BTLA) has been implicated in the regulation of autoimmunity and may potentially play an important role in allograft tolerance. This study investigated the effect of an agonistic anti-BTLA mAb (3C10) in the fully major histocompatibility complex-mismatched murine cardiac transplantation. METHODS: CBA mice underwent transplantation of C57BL/6 hearts and received one dose of 3C10 on the day of transplantation (day 0) or four doses of 3C10 on day 0, 3, 6, and 9. Adoptive transfer studies were performed to determine whether regulatory cells were generated. Moreover, to confirm the requirement for regulatory T cell and Th-2 cytokines, anti-interleukin (IL)-2 receptor alpha antibody (PC-61) or anti-IL-10 antibody (JES-2A5) was administered to a 3C10-treated CBA recipient. RESULTS: CBA mice treated with one and four doses of 3C10 prolonged allograft survival (median survival times [MSTs], 43 and >100 days, respectively). Secondary CBA recipients given whole splenocytes or CD4 cells from primary 3C10-treated CBA recipients had significantly prolonged survival of C57BL/6 hearts (MSTs, >100 in both). Also, flow cytometry studies showed an increased CD4CD25Foxp3 cell population in 3C10-treated mice. Additionally, IL-2 and interferon-γ production were suppressed in 3C10-treated mice, and IL-4 and IL-10 from 3C10-treated CBA mice increased. Moreover, 3C10 directly suppressed alloproliferation in a mixed leukocyte culture. However, administration of PC-61 or JES-2A5 clearly attenuated prolonged survival of 3C10-treated mice (MSTs, 15.5 and 13.5 days, respectively). CONCLUSION: 3C10 could control acute rejection by its suppressive effect on alloreactive T cells and induction of IL-10-dependent regulatory CD4 T cells.

Clarifying off-target effects for torcetrapib using network pharmacology and reverse docking approach.

BACKGROUND: Torcetrapib, a cholesteryl ester transfer protein (CETP) inhibitor which raises high-density lipoprotein (HDL) cholesterol and reduces low-density lipoprotein (LDL) cholesterol level, has been documented to increase mortality and cardiac events associated with adverse effects. However, it is still unclear the underlying mechanisms of the off-target effects of torcetrapib. RESULTS: In the present study, we developed a systems biology approach by combining a human reassembled signaling network with the publicly available microarray gene expression data to provide unique insights into the off-target adverse effects for torcetrapib. Cytoscape with three plugins including BisoGenet, NetworkAnalyzer and ClusterONE was utilized to establish a context-specific drug-gene interaction network. The DAVID functional annotation tool was applied for gene ontology (GO) analysis, while pathway enrichment analysis was clustered by ToppFun. Furthermore, potential off-targets of torcetrapib were predicted by a reverse docking approach. In general, 10503 nodes were retrieved from the integrative signaling network and 47660 inter-connected relations were obtained from the BisoGenet plugin. In addition, 388 significantly up-regulated genes were detected by Significance Analysis of Microarray (SAM) in adrenal carcinoma cells treated with torcetrapib. After constructing the human signaling network, the over-expressed microarray genes were mapped to illustrate the context-specific network. Subsequently, three conspicuous gene regulatory networks (GRNs) modules were unearthed, which contributed to the off-target effects of torcetrapib. GO analysis reflected dramatically over-represented biological processes associated with torcetrapib including activation of cell death, apoptosis and regulation of RNA metabolic process. Enriched signaling pathways uncovered that IL-2 Receptor Beta Chain in T cell Activation, Platelet-Derived Growth Factor Receptor (PDGFR) beta signaling pathway, IL2-mediated signaling events, ErbB signaling pathway and signaling events mediated by Hepatocyte Growth Factor Receptor (HGFR, c-Met) might play decisive characters in the adverse cardiovascular effects associated with torcetrapib. Finally, a reverse docking algorithm in silico between torcetrapib and transmembrane receptors was conducted to identify the potential off-targets. This screening was carried out based on the enriched signaling network analysis. CONCLUSIONS: Our study provided unique insights into the biological processes of torcetrapib-associated off-target adverse effects in a systems biology visual angle. In particular, we highlighted the importance of PDGFR, HGFR, IL-2 Receptor and ErbB1tyrosine kinase might be direct off-targets, which were highly related to the unfavorable adverse effects of torcetrapib and worthy of further experimental validation.

Central role for interleukin-2 in type 1 diabetes.

Type 1 diabetes presents clinically with overt hyperglycemia resulting from progressive immune-mediated destruction of pancreatic ß-cells and associated metabolic dysfunction. Combined genetic and immunological studies now highlight deficiencies in both the interleukin-2 (IL-2) receptor and its downstream signaling pathway as a central defect in the pathogenesis of type 1 diabetes. Prior intervention studies in animal models indicate that augmenting IL-2 signaling can prevent and reverse disease, with protection conferred primarily by restoration of regulatory T-cell (Treg) function. In this article, we will focus on studies of type 1 diabetes noting deficient IL-2 signaling and build what we believe forms the molecular framework for their contribution to the disease. This activity results in the identification of a series of potentially novel therapeutic targets that could restore proper immune regulation in type 1 diabetes by augmenting the IL-2 pathway.

Hot spots and transient pockets: predicting the determinants of small-molecule binding to a protein-protein interface.

Protein-protein interfaces are considered difficult targets for small-molecule protein-protein interaction modulators (PPIMs ). Here, we present for the first time a computational strategy that simultaneously considers aspects of energetics and plasticity in the context of PPIM binding to a protein interface. The strategy aims at identifying the determinants of small-molecule binding, hot spots, and transient pockets, in a protein-protein interface in order to make use of this knowledge for predicting binding modes of and ranking PPIMs with respect to their affinity. When applied to interleukin-2 (IL-2), the computationally inexpensive constrained geometric simulation method FRODA outperforms molecular dynamics simulations in sampling hydrophobic transient pockets. We introduce the PPIAnalyzer approach for identifying transient pockets on the basis of geometrical criteria only. A sequence of docking to identified transient pockets, starting structure selection based on hot spot information, RMSD clustering and intermolecular docking energies, and MM-PBSA calculations allows one to enrich IL-2 PPIMs from a set of decoys and to discriminate between subgroups of IL-2 PPIMs with low and high affinity. Our strategy will be applicable in a prospective manner where nothing else than a protein-protein complex structure is known; hence, it can well be the first step in a structure-based endeavor to identify PPIMs.

Focus on composition and interaction potential of single-pass transmembrane domains.

Transmembrane domains (TMD) connect the inner with the outer world of a living cell. Single TMD containing (bitopic) receptors are of particular interest, because their oligomerization seems to be a common activation mechanism in cell signaling. We analyzed the composition of TMDs in bitopic proteins within the proteomes of 12 model organisms. The average number of strongly polar and charged residues decreases during evolution, while the occurrence of a dimerization motif, GxxxG, remains unchanged. This may reflect the avoidance of unspecific binding within a growing receptor interaction network. In addition, we propose a new experimental approach for studying helix-helix interactions in giant plasma membrane vesicles using scanning fluorescence cross-correlation spectroscopy. Measuring eGFP/mRFP tagged versions of cytokine receptors confirms the homotypic interactions of the erythropoietin receptor in contrast to the Interleukin-4 receptor chains. As a proof of principle, by swapping the TMDs, the interaction potential of erythropoietin receptor was partially transferred to Interleukin-4 receptor α and vice versa. Non-interacting receptors can therefore serve as host molecules for TMDs whose oligomerization capability must be assessed. Computational analysis of the free energy gain resulting from TMD dimer formation strongly corroborates the experimental findings, potentially allowing in silico pre-screening of interacting pairs.

IL-2 induces conformational changes in its preassembled receptor core, which then migrates in lipid raft and binds to the cytoskeleton meshwork.

While interleukin (IL)-2 clearly initiates the sequential assembly of its soluble receptor fragments (sIL-2R) in vitro (with sIL-2Rα first, sIL-2Rß second, and sγc last), the assembly mechanism of full-length subunits (IL-2R) at the surface of living lymphocytes remains to be elucidated. Here we demonstrate by fluorescence cross-correlated spectroscopy that native IL-2Rß and γc assemble spontaneously at the surface of living human leukemia T cells (Kit-225 cell line) in the absence of IL-2 and with 1:1 stoichiometry. The dissociation constant of the membrane-embedded IL-2Rß/γc complex is measured in situ. Förster fluorescence resonance energy transfer analyzed by confocal microscopy of transfected COS-7 cells between combination pairs of various-length receptor chain constructions, using green fluorescent protein derivatives as cytoplasmic carboxy-terminal extensions, showed that IL-2Rß:ECFP and γc:EYFP bind each other through their extracellular domains, and that IL-2 binding brings their transmembrane domains 30 Å closer together. These observations demonstrate that IL-2Rß/γc heterodimers are preformed and that their cytoplasmic domains, carrying Janus kinase (Jak) 1 and Jak3, are pulled and tethered together on cytokine binding, triggering signaling transduction. IL-2 binding stabilizes IL-2/IL-2R complexes in membrane nanodomains that promote Jak1/Jak3 phosphorylation. The complexes then interact with the cytoskeleton, which slows receptor diffusion (as measured by fluorescence cross-correlated spectroscopy) and promotes STAT (signal transducer and activator of transcription) 5 phosphorylation. Separation of IL-2-activated receptors from Triton-lysed cells in detergent-resistant membrane nanodomains by ultracentrifugation on a sucrose gradient confirmed their presence in lipid rafts. The release of the IL-2-activated receptor from cytochalasin-treated cells and the IL-2-induced recruitment of actin and tubulin, analyzed by immunoprecipitation, confirmed that the activated receptor interacts with the cytoskeleton. Although IL-2Rα (the third chain that gives the IL-2Rß/γc receptor core its high affinity for IL-2) is highly expressed at the cell surface and mainly clustered in membrane microdomains at the surface of Kit-225 cells, the few free IL-2Rα present bind last to the IL-2/IL-2Rß/γc complex and lock IL-2 to its binding site for prolonged action, promoting signal amplification.

Interleukin 2 in cancer therapy.

Cancer immunotherapy with interleukin-2 (IL-2) has demonstrated long term disease control in metastatic renal cell carcinoma and malignant melanoma. With introduction of novel kinase inhibitors, immunomodulatory molecules, cytokines, and vaccines for treatment of cancer there is an increased interest in combining these therapeutic strategies with IL-2. Here we discuss toxicity and established activity of IL-2 in the management of advanced malignancies, and speculate on future use of this cytokine for treatment of cancer.

HotPoint: hot spot prediction server for protein interfaces.

The energy distribution along the protein-protein interface is not homogenous; certain residues contribute more to the binding free energy, called 'hot spots'. Here, we present a web server, HotPoint, which predicts hot spots in protein interfaces using an empirical model. The empirical model incorporates a few simple rules consisting of occlusion from solvent and total knowledge-based pair potentials of residues. The prediction model is computationally efficient and achieves high accuracy of 70%. The input to the HotPoint server is a protein complex and two chain identifiers that form an interface. The server provides the hot spot prediction results, a table of residue properties and an interactive 3D visualization of the complex with hot spots highlighted. Results are also downloadable as text files. This web server can be used for analysis of any protein-protein interface which can be utilized by researchers working on binding sites characterization and rational design of small molecules for protein interactions. HotPoint is accessible at http://prism.ccbb.ku.edu.tr/hotpoint.

Immunological synapses within context: patterns of cell-cell communication and their application in T-T interactions.

The cell-biology of intercellular communication between T cells and their partners has been greatly advanced over the past 10 years. The key morphological and motility features of cell contact-based communication between T cells and APCs can now be seen as a collection of patterns for cell-cell interactions amongst immune cells more generally, each serving to contribute to the outcome of the contact both locally and globally. Here we review the conservation of these patterns, amongst which is the emergent "immunological synapse," and describe a newly defined example, formed between the adjacent activating T cells. We subsequently seek to put these and the pattern more generally into the framework of system-wide behavior of the immune system. We postulate that the patterns are fine-tuned to provide quorum-like decisions by collections of activating and activated cells that interact over time and space.

18F-FDG PET, genotype-corrected ACE and sIL-2R in newly diagnosed sarcoidosis.

PURPOSE: Angiotensin-converting enzyme (ACE) and soluble interleukin-2 receptor (sIL-2R) are serological markers, widely used for determining sarcoidosis activity. (18)F-FDG PET has proven to be a sensitive technique in the imaging of sarcoidosis. The aim of this study was to determine sensitivity of (18)F-FDG PET, genotype-corrected ACE and sIL-2R in active sarcoidosis as well as their correlation. METHODS: This retrospective study included 36 newly diagnosed, symptomatic sarcoidosis patients. ACE and sIL-2R levels were simultaneously obtained within 4 weeks of (18)F-FDG PET. ACE was corrected for genotype and expressed as Z-score. (18)F-FDG PET was visually evaluated and scored as positive or negative. Maximum and average standardized uptake values (SUV(max) and SUV(avg)) were compared with ACE and sIL-2R. RESULTS: (18)F-FDG PET was found positive in 34 of 36 patients (94%). Thirteen patients (36%) showed an increased ACE with the highest sensitivity found in patients with the I/I genotype (67%). Seventeen patients (47%) showed an increased sIL-2R. No correlation was found between SUV and ACE or sIL-2R. Increased ACE and sIL-2R correlated with a positive (18)F-FDG PET in 12 patients (92%) and 16 patients (94%), respectively. CONCLUSION: (18)F-FDG PET is a very sensitive technique to assess active sarcoidosis, in contrast with ACE and sIL-2R, suggesting a pivotal role for (18)F-FDG PET in future sarcoidosis assessment.

T cell growth control using hapten-specific antibody/interleukin-2 receptor chimera.

IL-2 is a cytokine that is essential for the expansion and survival of activated T cells. Although adoptive transfer of tumor-specific T cells with IL-2 is one of strategies for cancer immunotherapy, it is essential to replace IL-2 that exerts severe side effects in vivo. To solve this problem, we propose to use an antibody/IL-2R chimera, which can transduce a growth signal in response to a cognate antigen. We constructed two chimeras, in which ScFv of anti-fluorescein antibody was tethered to extracellular D2 domain of erythropoietin receptor and transmembrane/cytoplasmic domains of IL-2Rbeta or gamma chain. When the chimeras were co-expressed in IL-3-dependent pro-B cell line Ba/F3 and IL-2-dependent T cell line CTLL-2, gene-modified cells were selectively expanded in the absence of IL-3 and IL-2, respectively, by adding fluorescein-conjugated BSA (BSA-FL) as a cognate antigen. Growth assay revealed that the cells with the chimeras transduced a growth signal in a BSA-FL dose-dependent manner. Furthermore, STAT3, STAT5, ERK1/2 and Akt, which are hallmarks for IL-2R signaling, were all activated by the chimeras in CTLL-2 transfectant. We also demonstrated that the chimeras were functional in murine primary T cells. These results demonstrate that the antibody/IL-2R chimeras could substantially mimic the wild-type IL-2R and could specifically expand gene-modified T cells in the presence of the cognate antigen.