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

Executive Summary: PF-562271 HCl is a hydrochloride salt of PF-562271, a potent, ATP-competitive, reversible inhibitor with IC₅₀ values of 1.5 nM for FAK and 14 nM for Pyk2, displaying >100-fold selectivity over other kinases except select CDKs (APExBIO). In xenograft and transgenic mouse models, it dose-dependently suppresses FAK phosphorylation with an EC₅₀ of 93 ng/mL, resulting in reduced tumor proliferation and metastasis (Wang et al., 2025). PF-562271 HCl is insoluble in water/ethanol but dissolves in DMSO at ≥26.35 mg/mL with gentle warming. It enables targeted dissection of FAK/Pyk2 signaling in cancer biology, aiding studies of cellular adhesion, migration, and survival. For optimal stability, storage at -20°C is recommended.

Biological Rationale

Focal adhesion kinase (FAK) and proline-rich tyrosine kinase 2 (Pyk2) are non-receptor tyrosine kinases. Both regulate cell adhesion, migration, proliferation, and survival in physiological and pathological contexts (APExBIO). Aberrant FAK/Pyk2 signaling is implicated in tumor growth, metastasis, and resistance to therapy in solid cancers (Signal-Transducer-Article). Selective inhibition of these kinases can disrupt tumor cell motility and survival, making PF-562271 HCl a valuable tool for unraveling tumor biology and informing new cancer therapies (Wang et al., 2025).

Mechanism of Action of PF-562271 HCl

PF-562271 HCl competitively binds to the ATP-binding site of FAK and Pyk2, inhibiting their kinase activities. It is reversible and highly selective, with a 10-fold preference for FAK over Pyk2 and >100-fold selectivity over most other kinases (APExBIO). The compound blocks FAK autophosphorylation at Tyr397, a critical step for downstream signaling. This leads to disruption of focal adhesion turnover, cell spreading, and migration, as well as inhibition of survival signaling cascades (e.g., PI3K/AKT, ERK/MAPK). The result is impaired tumor cell proliferation and reduced metastatic potential (Cyclin-D1 Article). In the tumor microenvironment, FAK inhibition can modulate immune cell recruitment and enhance the efficacy of immunotherapies (Signal-Transducer-Article).

Evidence & Benchmarks

• PF-562271 HCl inhibits FAK kinase activity with an IC₅₀ of 1.5 nM at 25°C in biochemical assays (APExBIO).
• It inhibits Pyk2 with an IC₅₀ of 14 nM under similar conditions, showing a ~10-fold selectivity for FAK (APExBIO).
• PF-562271 HCl demonstrates >100-fold selectivity over most other kinases except several cyclin-dependent kinases (APExBIO).
• In xenograft and transgenic mouse tumor models, PF-562271 HCl inhibits FAK phosphorylation with an EC₅₀ of 93 ng/mL and significantly reduces tumor volume over 2–4 weeks (Wang et al., 2025).
• The compound is insoluble in water and ethanol but dissolves in DMSO at ≥26.35 mg/mL with gentle warming (room temperature, <40°C) (APExBIO).
• Storage at -20°C ensures chemical stability for >12 months (APExBIO).

This article extends the mechanistic depth and practical integration guidance beyond MolecularBeacon.net, which focuses more on translational research context.

Applications, Limits & Misconceptions

PF-562271 HCl is used to interrogate FAK/Pyk2-driven processes in preclinical cancer models, including:

• Assessing tumor growth and metastasis in cell-based and animal models (Pelubiprofenchems.com).
• Deciphering roles of FAK/Pyk2 in tumor microenvironment modulation and immune cell infiltration (Signal-Transducer-Article).
• Combining with radiotherapy or immunotherapies to explore synergistic antitumor effects (Wang et al., 2025).
• Benchmarking kinase selectivity and off-target profiles in high-throughput screens.

Common Pitfalls or Misconceptions

• PF-562271 HCl is not effective against receptor tyrosine kinases or all serine/threonine kinases; its activity is specific to FAK, Pyk2, and a subset of CDKs.
• Compound is insoluble in aqueous buffers and ethanol; dissolution must be in DMSO for accurate dosing.
• FAK inhibition may not fully suppress tumor progression in cancers driven by redundant or compensatory pathways (e.g., Src, integrins).
• PF-562271 HCl does not directly activate immune cells; its immunomodulatory effects are indirect via tumor microenvironment remodeling.
• Storage above -20°C or repeated freeze-thaw cycles can degrade compound potency.

This article clarifies application scope and experimental caveats beyond what is covered in Cyclin-D1.com, which focuses on assay reproducibility.

Workflow Integration & Parameters

PF-562271 HCl (SKU A8345, APExBIO) is provided as a solid. Dissolve at ≥26.35 mg/mL in DMSO at room temperature or with gentle warming (<40°C). For cell-based assays, dilute DMSO stocks to final concentrations (1–1000 nM) in serum-free or complete media, ensuring final DMSO <0.1% (v/v) to minimize cytotoxicity. For in vivo studies, formulate in DMSO or compatible vehicle according to animal welfare guidelines. Store aliquots at -20°C protected from light. Monitor for precipitation before use. Experimental controls should include vehicle-only and, where relevant, alternative kinase inhibitors.

This article updates practical handling guidelines and experimental design considerations in light of recent advances discussed in Signal-Transducer Article 105, adding explicit storage and solubility benchmarks.

Conclusion & Outlook

PF-562271 HCl, as supplied by APExBIO, is a rigorously validated, selective, reversible ATP-competitive inhibitor of FAK and Pyk2. It enables high-fidelity dissection of focal adhesion kinase pathways, supports cancer proliferation and metastasis research, and informs innovative combination therapy strategies. Careful attention to solvent compatibility and storage extends compound utility and reproducibility. As mechanistic understanding of FAK/Pyk2 expands, PF-562271 HCl remains a cornerstone for translational research, especially in the context of evolving immunotherapy and radiotherapy paradigms (Wang et al., 2025).