TL1A reporter cell line

Notification:  Reference #hkb-tl1a-av can only be ordered together with reference #hkb-tl1a.

TL1A responsive NF-κB/AP1-SEAP reporter assay

Signaling pathway in HEK-Blue™ TL1A cells

HEK-Blue™ TL1A cells are designed to monitor human TL1A-induced NF-κB/AP1 stimulation or inhibition. This colorimetric cytokine bioassay can be used to screen activatory or inhibitory molecules, such as engineered cytokines and neutralizing antibodies, respectively.

TL1A reporter cells respond specifically to recombinant human (h) and mouse (m) TL1A. Their reliable and consistent performance makes them suitable for release assays of activatory and inhibitory molecules such as Tulisokibart, a monoclonal antibody that targets TL1A and prevents its binding to its receptor (see figures).

Key features

• Readily assessable NF-κB/AP1 reporter activity
• Convenient readout using QUANTI-Blue™ Solution
• High sensitivity to human (h)TL1A and mouse (m)TL1A activity
• Stability guaranteed for 20 passages

Applications

• Therapeutic development
• Drug screening
• Release assay

Tumor necrosis factor (TNF)-like 1A (TL1A) drives the production of pro-inflammatory cytokines and differentiation of T helper subsets. Abnormal TL1A expression is linked to enteric autoimmune diseases (e.g. Crohn’s disease) and extends to other diseases, including allergic airway inflammation.

 More details

InvivoGen’s products are for internal research use only and not for clinical or veterinary use.

Figures

Dose-response of HEK-Blue™ TL1A cells to recombinant TL1A. Cells were stimulated with increasing concentrations of recombinant human (h) or mouse (m) TL1A. After overnight incubation, the NF-κB/AP1-induced SEAP activity was determined using QUANTI-Blue™, a SEAP detection reagent. Data are shown as optical density (OD) at 650 nm (mean ± SEM).

 

Response of HEK-Blue™ TL1A cells to a panel of cytokines. Cells were stimulated with various human and murine recombinant cytokines: 10 ng/ml of hTL1A, mTL1A, hAPRIL, hBAFF, hCD40L, hRANKL, hTNF-α, hIFN-γ, hIL-1β, hIL-6, hIL-27 and 1000 U/ml hIFN-α. After overnight incubation, SEAP activity was assessed using QUANTI‑Blue™ The OD at 630 nm is shown as mean ± SEM.

Dose-dependent inhibition of HEK-Blue™ TL1A cell response using Tulisokibart biosimilar. Increasing concentrations of Anti-hTL1A-hIgG1 (0.01 ng/ml - 1 µg/ml) were incubated with recombinant human TL1A (3 ng/ml) for 1 h before the addition of HEK-Blue™ TL1A cells. After overnight incubation, SEAP activity in the cell culture supernatant was assessed using QUANTI-Blue™ Solution. Data are shown in percentage of activity (mean ± SEM).

Specifications

Cell type: Epithelial

Tissue origin: Human Embryonic Kidney

Target: TL1A

Specificity: Human, Mouse

Reporter gene: SEAP

Antibiotic resistance: Blasticidin, Zeocin®

Detection range: 300 pg/ml - 30 ng/ml (hTL1A and mTL1A)

Growth medium: Complete DMEM (see TDS)

Growth properties: Adherent

Mycoplasma-free: Verified using Plasmotest™

Quality control: Each lot is functionally tested and validated.

Contents

• 1 vial containing 3-7 x 10⁶ cells
• 1 ml of Blasticidin (10 mg/ml)
• 1 ml of Zeocin® (100 mg/ml)
• 1 ml Normocin® (50 mg/ml)
• 1 ml of QB reagent and 1 ml of QB buffer (sufficient to prepare 100 ml of QUANTI-Blue™ Solution, a SEAP detection reagent).

 Shipped on dry ice (Europe, USA, Canada)

Details

Cell line description

HEK-Blue™ TL1A cells were generated by stable transfection of the human embryonic kidney HEK293 cell line with the gene encoding the human Death Receptor 3 (DR3) and a SEAP (secreted embryonic alkaline phosphatase) reporter gene under the control of the IFN-β minimal promoter fused to five AP-1 and five NF-κB binding sites.

The binding of TL1A to its receptor triggers a signaling cascade leading to AP-1/NF-κB activation and the subsequent production of SEAP.  This can be readily assessed in the supernatant using QUANTI-Blue™ Solution, a SEAP detection reagent. HEK-Blue™ TL1A cells detect human (h) and mouse (m) TL1A. Of note, these cells also respond to two other human AP-1/ NF-κB-signaling cytokines, TNF-α and IL-1 β. However, they do not respond to other cytokines of the TNF superfamily: APRIL, BAFF, RANKL, and CD40L (see figures).

TL1A background

Tumor necrosis factor (TNF)-like 1A (TL1A), also called TNFSF15, is a cytokine that belongs to the TNF superfamily. TL1A is produced by endothelial cells, activated dendritic cells, and macrophages. It is synthesized as a membrane-bound trimeric molecule. Alternative splicing or cleavage by the tumor necrosis factor-alpha converting enzyme results in soluble TL1A [1]. Both forms bind the homotrimeric transmembrane death receptor 3 (DR3), triggering TRADD/TRAF2/RIP signaling, and ultimately leading to NF-κB and MAPKs activation. Subsequent gene expression in target cells drives the production of pro-inflammatory cytokines and differentiation of T helper subsets [1].

Relevance for therapeutics development

TL1A is normally responsible for mediating protective immune responses. However, it can also play a role in chronic inflammatory disorders and have harmful impacts. Abnormal TL1A expression or TL1A overactivity promotes excessive inflammation and immune cell infiltration, contributing to tissue damage and further inflammation amplification [1].

Pharmaceutical companies have developed potent monoclonal antibodies to block TL1A signaling and stop the downstream inflammatory cascade and tissue damage, primarily in the onset of asthma and Crohn’s disease. Anti-TL1A inhibitors such as Tulisokibart (MK-7240 or PRA023, Merck), RVT-3101 (PF-06480605, Roche), and Duvakitug (TEV-48574, Sanofi & Teva Pharmaceuticals) have demonstrated "best-in-class" potential for treating inflammation and scarring in IBD [2-4]. Duvakitug is currently being studied for its safety as an asthma medication [5-6].

References:

1. Xu W.D., et al., 2022. Role of TL1A in Inflammatory Autoimmune Diseases: A Comprehensive Review. Front. Immunol. 13: 891328.
2. Sands, B.E., et al., 2024. Phase 2 Trial of Anti-TL1A Monoclonal Antibody Tulisokibart for Ulcerative Colitis. N Engl J Med, 391(12): p. 1119-1129.
3. Danese, S., et al., 2021. Anti-TL1A Antibody PF-06480605 Safety and Efficacy for Ulcerative Colitis: A Phase 2a Single-Arm Study. Clinical Gastroenterology and Hepatology. 19(11): p. 2324-2332.e6.
4. Solitano, V., et al., 2024. TL1A inhibition for inflammatory bowel disease treatment: From inflammation to fibrosis. Med. 5(5): p. 386-400.
5. Balyan, R., et al.,, 2024. P633 First-in-Human Pharmacokinetic and Safety Study of an Anti-TL1A Antibody, TEV-48574, in Healthy Volunteers and Asthma Patients. Journal of Crohn's and Colitis. 18(Supplement_1): p. i1215-i1216.
6. Raphael, G., et al., 2024. P1061 TEV-48574, an anti-TL1A antibody in development for use in IBD, is safe and well tolerated following 16 weeks of subcutaneous treatment in adults with severe uncontrolled T2-low/non T2 asthma. Journal of Crohn's and Colitis. 18(Supplement_1): p. i1908-i1908.

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