PF-562271 HCl: Potent ATP-Competitive FAK/Pyk2 Inhibitor for Cancer Research

Executive Summary: PF-562271 HCl is a hydrochloride salt form of a small molecule that acts as a potent, reversible ATP-competitive inhibitor of focal adhesion kinase (FAK, IC50 = 1.5 nM) and proline-rich tyrosine kinase 2 (Pyk2, IC50 = 14 nM). This compound offers >10-fold selectivity for FAK over Pyk2 and >100-fold selectivity against most other kinases except certain cyclin-dependent kinases, enabling highly specific pathway interrogation in cancer models (Keller et al. 2023). PF-562271 HCl inhibits FAK phosphorylation in vivo (EC50 = 93 ng/mL), leading to the suppression of tumor growth and metastasis in mouse models. The compound is supplied by APExBIO as SKU A8345 and is widely adopted for mechanistic studies of FAK/Pyk2 signaling, tumor microenvironment modulation, and anti-cancer drug development [product details]. Its defined solubility and stability characteristics further support robust experimental workflows.

Biological Rationale

Focal adhesion kinase (FAK, also known as PTK2) is a non-receptor tyrosine kinase. It orchestrates cell adhesion, migration, survival, and proliferation. FAK is upregulated or hyperactivated in many solid tumors, correlating with metastatic progression and poor prognosis (Keller et al. 2023). Proline-rich tyrosine kinase 2 (Pyk2, PTK2B) is structurally similar to FAK, sharing 48% amino acid identity, and also mediates integrin and growth factor receptor signaling. Both kinases regulate the tumor microenvironment, influencing stromal interactions, immune cell infiltration, and extracellular matrix remodeling [Related: FAK/Pyk2 selectivity review]. Aberrant FAK/Pyk2 activity is implicated in resistance to targeted therapies such as HER2 inhibitors in breast cancer, highlighting the need for novel interventions (Keller et al. 2023).

Mechanism of Action of PF-562271 HCl

PF-562271 HCl is a reversible, ATP-competitive inhibitor. It binds to the kinase domain of FAK and Pyk2, blocking autophosphorylation at critical tyrosine residues (e.g., Y397 in FAK). This inhibition prevents downstream activation of signaling pathways implicated in cell migration, survival, and metastasis. The compound demonstrates IC50 values of 1.5 nM for FAK and 14 nM for Pyk2, with approximately 10-fold selectivity for FAK and >100-fold selectivity over other kinases except some cyclin-dependent kinases (CDKs). PF-562271 HCl has been shown to inhibit FAK phosphorylation in tumor-bearing mice at an EC50 of 93 ng/mL, resulting in reduced tumor growth and metastatic spread (Keller et al. 2023). These quantitative benchmarks position it as a reference tool for dissecting the focal adhesion kinase signaling pathway.

Evidence & Benchmarks

• PF-562271 HCl inhibits FAK kinase activity with an IC50 of 1.5 nM, and Pyk2 with an IC50 of 14 nM, demonstrating 10-fold selectivity (Keller et al. 2023, DOI).
• The compound exhibits >100-fold selectivity against other kinases, except for certain cyclin-dependent kinases, minimizing off-target effects (Keller et al. 2023, DOI).
• In mouse models, PF-562271 HCl suppresses FAK phosphorylation with an EC50 of 93 ng/mL, leading to reduction of tumor volume and metastasis (Keller et al. 2023, DOI).
• PF-562271 HCl is soluble in DMSO at ≥26.35 mg/mL at room temperature with gentle warming but insoluble in water or ethanol (APExBIO product page).
• Optimized for use in studies of FAK/Pyk2 signaling, tumor microenvironment modulation, and anti-cancer drug development (Precision inhibitor review).

Applications, Limits & Misconceptions

PF-562271 HCl is widely used in:

• Dissecting FAK/Pyk2 signaling pathways in cancer and stromal cells.
• Investigating tumor microenvironment modulation, including effects on immune cell infiltration and matrix remodeling.
• Preclinical testing of anti-cancer strategies targeting focal adhesion signaling.
• Exploring resistance mechanisms to targeted therapies, such as HER2 inhibitors in breast cancer (Keller et al. 2023).

Recent literature highlights its unique value for precision experiments due to its nanomolar potency and reversible inhibition profile. See this review for a focused workflow guide; the present article extends that discussion with updated selectivity and in vivo benchmarks.

Common Pitfalls or Misconceptions

• Not a pan-kinase inhibitor: PF-562271 HCl does not broadly inhibit all kinases; selectivity is >100-fold over most non-FAK/Pyk2 kinases except some CDKs.
• Limited solubility: The compound is insoluble in water and ethanol; use DMSO at ≥26.35 mg/mL with gentle warming (APExBIO).
• Not suitable for long-term solution storage: Stability is optimal as a solid at -20°C; prepared solutions should be used promptly.
• Potential off-target effects at high concentrations: Some cyclin-dependent kinases may be inhibited if concentrations exceed recommended ranges.
• Not a direct anti-viral or anti-bacterial agent: Activity is specific to kinase targets relevant to mammalian cell signaling, not infectious pathogens.

Workflow Integration & Parameters

PF-562271 HCl (SKU A8345) is provided by APExBIO in solid form. Recommended storage is at -20°C, protected from light and moisture. For use, dissolve in DMSO at ≥26.35 mg/mL; gentle warming may aid dissolution. Avoid aqueous or ethanol-based solvents. Prepare working solutions fresh and use promptly to maintain activity. Typical experimental concentrations range from 10 nM to 10 μM, depending on cell line and assay design. For in vivo studies, dosing should be referenced against published EC50 values (e.g., 93 ng/mL for FAK phosphorylation inhibition in mice) (Keller et al. 2023). For troubleshooting cell viability or matrix modulation assays, researchers may consult this protocol review, which this article updates with the latest selectivity and in vivo efficacy data.

For extended mechanistic or immunomodulatory applications, see this analysis; the current overview supplements that resource with contemporary benchmarks and workflow integration guidance.

Conclusion & Outlook

PF-562271 HCl is a powerful, well-characterized ATP-competitive, reversible FAK/Pyk2 inhibitor with validated selectivity and in vivo efficacy benchmarks. Its application extends from basic mechanistic studies to preclinical cancer research, with a growing role in dissecting resistance mechanisms and tumor microenvironment dynamics. As new signaling crosstalk and resistance pathways are elucidated, PF-562271 HCl will remain a cornerstone pharmacological tool for the cancer research community. For detailed product specifications and ordering, see the APExBIO PF-562271 HCl product page.