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    • Birinapant (TL32711): SMAC Mimetic IAP Antagonist for Apo...

    2025-10-30

    Birinapant (TL32711): SMAC Mimetic IAP Antagonist for Apoptosis Induction

    Executive Summary: Birinapant (TL32711) is a bivalent SMAC mimetic that antagonizes inhibitor of apoptosis proteins (IAPs) with high affinity, notably XIAP (Kd = 45 nM) and cIAP1 (Kd < 1 nM), leading to rapid degradation of cIAP1/2 and subsequent caspase-8-mediated apoptosis in cancer cells (ApexBio). It inhibits TNF-mediated NF-κB activation, enhances TRAIL-induced cell death, and shows efficacy in preclinical melanoma and inflammatory breast cancer models (Survivin.net). Solubility in DMSO (≥40.35 mg/mL) and ethanol (≥46.9 mg/mL) supports diverse laboratory workflows. Birinapant is widely used for apoptosis research, cancer biology, and probing IAP-related signaling. It is best employed promptly after solution preparation and stored at -20°C (ApexBio).

    Biological Rationale

    Apoptosis resistance is a hallmark of cancer, contributing to treatment failure and poor prognosis. Inhibitor of apoptosis proteins (IAPs) such as XIAP, cIAP1, and cIAP2 suppress caspase activity and downstream cell death, enabling tumor cell survival (Ren et al., 2025). SMAC (Second Mitochondria-Derived Activator of Caspases) mimetics, including Birinapant, are designed to antagonize IAPs and restore apoptotic signaling. Birinapant’s high-affinity binding to IAP BIR domains triggers their autoubiquitination and proteasomal degradation. The resulting reduction in IAP levels permits caspase activation and enhances the cytotoxicity of pro-apoptotic stimuli such as TNFα and TRAIL. In colorectal cancer, modulation of apoptosis via the p53 pathway has been shown to sensitize tumors to chemoradiotherapy, supporting the translational value of agents like Birinapant (Ren et al., 2025).

    Mechanism of Action of Birinapant (TL32711)

    Birinapant is a bivalent small molecule that structurally mimics endogenous SMAC, targeting the BIR3 domains of XIAP, cIAP1, cIAP2, and ML-IAP. It binds cIAP1 with a dissociation constant (Kd) below 1 nM and XIAP with a Kd of 45 nM (ApexBio). The compound induces rapid degradation of TRAF2-bound cIAP1 and cIAP2, leading to loss of IAP-mediated inhibition of caspases. Upon TNF stimulation, Birinapant promotes the formation of the caspase-8:RIPK1 complex, resulting in activation of executioner caspases and programmed cell death (Survivin.net). Birinapant also inhibits TNF-induced NF-κB activation by removing cIAP1/2, which are required for NIK ubiquitination and NF-κB pathway maintenance.

    Evidence & Benchmarks

    • Birinapant binds cIAP1 with Kd < 1 nM and XIAP with Kd = 45 nM, resulting in pan-IAP antagonism (ApexBio).
    • In inflammatory breast cancer cells, Birinapant enhances TRAIL potency and increases apoptotic population size (Survivin.net).
    • Birinapant administration in melanoma xenotransplantation models reduces cIAP1 protein levels and elevates apoptosis rates, as measured by TUNEL staining (TNFalphaInhibitors.com).
    • Birinapant inhibits TNF-induced NF-κB signaling, resulting in reduced inflammatory gene expression (TRAF2.com).
    • Birinapant’s pan-IAP antagonism leads to rapid cIAP1 degradation, PARP cleavage, and caspase activation in vitro (ApexBio).
    • Combining apoptosis-inducing inhibitors (such as SMAC mimetics) with chemoradiotherapy can restore sensitivity in colorectal cancer models with low pro-apoptotic drive (Ren et al., 2025).

    Compared to previous mechanistic overviews, this article synthesizes recent data on chemoradiotherapy synergy and the translational role of Birinapant in apoptosis-resistant tumors.

    Applications, Limits & Misconceptions

    Birinapant is primarily used for:

    • Apoptosis induction studies in cancer cell lines and primary tumor models.
    • Dissecting IAP-related signaling pathways (XIAP, cIAP1, cIAP2, ML-IAP).
    • Enhancing the efficacy of chemoradiotherapy and TRAIL-based therapies in preclinical settings.
    • Evaluating mechanisms of apoptosis resistance and therapeutic sensitization (Ren et al., 2025).

    For a strategic workflow guide, see this reference. This article extends previous guides by integrating actionable benchmarks and highlighting pitfalls from recent translational studies.

    Common Pitfalls or Misconceptions

    • Birinapant is not water-soluble: Attempting to dissolve in aqueous buffers leads to precipitation and loss of activity (ApexBio).
    • Solutions are unstable long-term: Prepared solutions in DMSO/ethanol should be used promptly and not stored for extended periods.
    • Birinapant alone may not overcome all forms of apoptosis resistance: Its efficacy depends on the presence of functional death receptor and caspase machinery (Ren et al., 2025).
    • In vivo dosing requires rigorous optimization: Animal studies must titrate dose and schedule to avoid off-target toxicity.
    • Not suitable for use as a direct therapeutic in humans without clinical protocol: Birinapant is for research use only.

    Workflow Integration & Parameters

    Birinapant (A4219) is supplied as a solid and should be stored at -20°C. For solution preparation, dissolve at concentrations up to 40.35 mg/mL in DMSO or 46.9 mg/mL in ethanol. Use warm (37°C) solvent and ultrasonic shaking to enhance solubility. Solutions should be freshly prepared and used immediately to ensure compound integrity. Birinapant is typically applied to cell cultures at nanomolar to low micromolar concentrations, with dosing guided by cell type sensitivity and experimental endpoint. For in vivo studies, animal dosing protocols vary and should be referenced from peer-reviewed literature. Birinapant is compatible with apoptosis assays (e.g., caspase activity, PARP cleavage, TUNEL) and can be combined with chemotherapeutic agents to assess synergy. See the A4219 kit page for detailed preparation tips. For troubleshooting and advanced workflows, this resource offers expanded protocols.

    Conclusion & Outlook

    Birinapant (TL32711) is a well-characterized SMAC mimetic IAP antagonist that enables precise interrogation of apoptosis pathways in cancer research. Its high affinity for XIAP and cIAPs, robust solubility profile, and proven efficacy in preclinical models make it a standard tool for apoptosis induction studies. As new biomarkers (e.g., MDM1, p53) emerge for predicting therapeutic response, Birinapant’s integration with chemoradiotherapy protocols may further enhance translational impact (Ren et al., 2025). Continued benchmarking, careful workflow integration, and awareness of solubility/stability limits will maximize research value.