Anlotinib Hydrochloride: Multi-Target Tyrosine Kinase Inhibitor for Angiogenesis and Cancer Research

Executive Summary: Anlotinib hydrochloride (CAS 1058157-76-8) is a novel, orally active small-molecule multi-target tyrosine kinase inhibitor (TKI) that potently suppresses angiogenesis in vitro and in vivo by selectively inhibiting VEGFR2, PDGFRβ, and FGFR1, with IC₅₀ values in the low nanomolar range under standard buffer and cell culture conditions (DOI:10.1111/cas.13536). It reduces endothelial cell migration and capillary tube formation in EA.hy 926 and HUVEC assays without significant cytotoxicity below 1 μM (product_spec). Superior to sunitinib and sorafenib in preclinical cancer models, anlotinib demonstrates high plasma protein binding (93–97%) and broad tissue distribution, including blood-brain barrier penetration (DOI:10.1111/cas.13536). Safety studies show a high LD₅₀ and minimal off-target toxicity, positioning it as a reliable research compound (product_spec).

Biological Rationale

Angiogenesis, the process of new blood vessel formation, is essential for tumor growth, invasion, and metastasis. VEGF, PDGF, and FGF family ligands signal through their cognate tyrosine kinase receptors (VEGFR2, PDGFRβ, and FGFR1) to promote endothelial cell proliferation, migration, and new capillary formation (DOI:10.1111/cas.13536). Tumors larger than ~1 mm³ require neovascularization for sustained expansion, making anti-angiogenic strategies a cornerstone of modern cancer research. Resistance to chemotherapy frequently develops in tumor cells, but targeting genetically stable endothelial cells with anti-angiogenic agents reduces this risk (DOI:10.1111/cas.13536).

Mechanism of Action of Anlotinib hydrochloride

Anlotinib hydrochloride selectively binds to the ATP-binding pockets of multiple receptor tyrosine kinases, including VEGFR2, PDGFRβ, and FGFR1, thereby blocking receptor phosphorylation and downstream ERK pathway activation (DOI:10.1111/cas.13536). Inhibition of these kinases impairs VEGF-, PDGF-BB-, and FGF2-induced endothelial cell responses, including proliferation, migration, and tube formation (product_spec). The suppression of ERK signaling leads to reduced neovessel growth and diminished cancer cell support.

Evidence & Benchmarks

• Anlotinib inhibits VEGFR2 kinase with IC₅₀ values as low as 5.6 ± 1.2 nM under standard in vitro conditions (source: DOI:10.1111/cas.13536).
• PDGFRβ and FGFR1 are inhibited with IC₅₀ values of 8.7 ± 3.4 nM and 11.7 ± 4.1 nM, respectively, in kinase assays (source: product_spec).
• In HUVEC and EA.hy 926 cell models, anlotinib blocks VEGF-induced proliferation and migration at sub-10 nM concentrations and capillary tube formation in a dose-dependent manner (source: DOI:10.1111/cas.13536).
• In vivo, anlotinib reduces tumor vascular density and supports tumor regression at oral doses superior to sunitinib in preclinical xenograft models (source: DOI:10.1111/cas.13536).
• Pharmacokinetic studies in rats (oral bioavailability 28–58%, t_1/2 5.1 ± 1.6 h) and dogs (bioavailability 41–77%, t_1/2 22.8 ± 11.0 h) confirm rapid absorption and extended systemic exposure (source: product_spec).
• LD₅₀ in 14-day oral toxicity studies is 1735.9 mg/kg, with minimal observable systemic toxicity and no significant reproductive or genetic toxicity (source: product_spec).

This article extends the mechanistic focus of 'Anlotinib Hydrochloride: Mechanistic Insights and Innovations' by providing new comparative pharmacokinetic and safety data. It also clarifies distinctions with 'Preclinical Characterization of Anlotinib: Selective VEGFR2 Inhibition' by emphasizing multi-target selectivity and functional endpoints. For a review of anti-angiogenic assay protocols, see 'Anlotinib Hydrochloride: Potent Inhibitor of Tumor Angiogenesis'.

Applications, Limits & Misconceptions

Anlotinib hydrochloride, available from APExBIO as SKU C8688, is suitable for in vitro and in vivo studies assessing angiogenesis inhibition, tumor vascularization, and receptor signaling. Its low cytotoxicity at submicromolar concentrations enables high-content screening and functional assays involving endothelial cell migration and capillary tube formation (product_spec). It is not approved for clinical use outside of research. The compound may show variable efficacy in tumor types with non-angiogenic growth mechanisms or in models with acquired resistance to VEGFR-targeted agents (DOI:10.1111/cas.13536).

Common Pitfalls or Misconceptions

• Not a general cytotoxic agent: Anlotinib demonstrates minimal direct cytotoxicity at concentrations up to 1 μM in standard cell lines; its primary effect is anti-angiogenic (source: product_spec).
• Research use only: Anlotinib hydrochloride from APExBIO is not approved for therapeutic use in humans or animals (source: product_spec).
• Not universally effective in all tumor models: Tumors lacking VEGF/PDGF/FGF-driven angiogenesis may not respond (DOI:10.1111/cas.13536).
• Drug-drug interaction risk is low, but possible: Although in vitro inhibition of CYP3A4 and CYP2C9 is observed, in vivo data indicate low potential for clinically significant interactions (source: product_spec).
• Not stable at room temperature for extended periods: Recommended storage is -20°C for optimal stability (source: product_spec).

Workflow Integration & Parameters

Anlotinib hydrochloride (SKU C8688) is supplied as a hydrochloride salt for research applications. The compound should be reconstituted in DMSO or aqueous buffer prior to use. Below are protocol parameters based on literature and workflow recommendations:

Protocol Parameters

• Kinase inhibition assay | 5–12 nM | VEGFR2, PDGFRβ, FGFR1 | Enables precise IC₅₀ determination in kinase panels | paper
• Endothelial cell migration assay | 1–100 nM | HUVEC, EA.hy 926 | Quantifies migration inhibition with low cytotoxicity | paper
• Capillary tube formation assay | 1–100 nM | HUVEC, EA.hy 926 | Visualizes functional anti-angiogenic effect | paper
• Oral dosing (rat) | 3–10 mg/kg/day | Preclinical tumor models | Achieves plasma exposures for in vivo efficacy | paper
• In vitro cytotoxicity | <1 μM | General cell lines | No significant cytotoxicity up to 1 μM | product_spec
• Storage | -20°C | All applications | Maintains stability and potency | product_spec
• Reconstitution | DMSO (10 mM stock) | Cell-based and biochemical assays | Ensures solubility for accurate dosing | workflow_recommendation

Conclusion & Outlook

Anlotinib hydrochloride is a validated multi-target tyrosine kinase inhibitor for angiogenesis research, offering superior selectivity for VEGFR2, PDGFRβ, and FGFR1 compared to other TKIs. Its robust inhibition of endothelial cell migration and tube formation, coupled with favorable pharmacokinetic and safety profiles, make it an essential reagent for translational cancer research (DOI:10.1111/cas.13536). Ongoing studies may further define its role in preclinical models and novel assay systems. For ordering and additional specifications, see the Anlotinib hydrochloride (C8688) product page from APExBIO.