title: HPLC method development, validation and impurity characterization for an antitumor Hsp90 inhibitor-PU-H71 (NSC 750424).
abstract: An HPLC method for the assay of the heat shock protein 90 inhibitor, PU-H71 (NSC 750424), has been developed and validated. The stress testing of PU-H71 was carried out in accordance with ICH guidelines Q1A (R2) under aqueous, acidic, alkaline, oxidative, thermolytic and photolytic conditions. The separation of PU-H71 from its impurities and degradation products was achieved within 50min on a Mac-Mod ACE 3 C18 column (150mm×4.6mm i.d., 3μm) with a gradient mobile phase comprising 20-95% acetonitrile in water, with 0.1% trifluroacetic acid in both phases. LC-quadrupole TOF/MS was used to obtain accurate mass data on various components as well as on their fragments for characterization of impurities and degradation products. The proposed HPLC assay method was validated for specificity, linearity (concentration range 0.1-0.3mg/mL, r≥0.9998), accuracy (recovery 99.7-101.1%), precision (intra-lab RSD≤1.39%, inter-lab RSD≤0.91%), sensitivity (LOD 0.08μg/mL), and ruggedness. The developed method was suitable for the assay and stability monitoring of PU-H71 drug substance.
authors.affiliation: SRI International, 333 Ravenswood Avenue, Menlo Park, CA 94025, USA.
authors.forename: Mingtao
authors.initials: M
authors.lastname: Liu
authors.affiliation: SRI International, 333 Ravenswood Avenue, Menlo Park, CA 94025, USA. Electronic address: jennie.wang@sri.com.
authors.forename: Jennie
authors.initials: J
authors.lastname: Wang
authors.affiliation: SRI International, 333 Ravenswood Avenue, Menlo Park, CA 94025, USA.
authors.forename: Xiaogang
authors.initials: X
authors.lastname: Wu
authors.affiliation: SRI International, 333 Ravenswood Avenue, Menlo Park, CA 94025, USA.
authors.forename: Euphemia
authors.initials: E
authors.lastname: Wang
authors.affiliation: SRI International, 333 Ravenswood Avenue, Menlo Park, CA 94025, USA.
authors.forename: Diego
authors.initials: D
authors.lastname: Baptista
authors.affiliation: Midwest Research Institute, 425 Volker Boulevard, Kansas City, MO 64110, USA.
authors.forename: Brendan
authors.initials: B
authors.lastname: Scott
authors.affiliation: Pharmaceutical Resources Branch, DCTD, NCI, 6130 Executive Boulevard, Bethesda, MD 20892, USA.
authors.forename: Paul
authors.initials: P
authors.lastname: Liu
mesh_terms: D000970:Antineoplastic Agents / Q000737:chemistry*; D052117:Benzodioxoles / Q000737:chemistry* / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004340:Drug Contamination; D004355:Drug Stability; D018841:HSP90 Heat-Shock Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D013058:Mass Spectrometry / Q000379:methods; D010084:Oxidation-Reduction; D010782:Photolysis; D011687:Purines / Q000737:chemistry* / Q000494:pharmacology*; D012680:Sensitivity and Specificity
chemical_list: D000970:Antineoplastic Agents; D052117:Benzodioxoles; D018841:HSP90 Heat-Shock Proteins; D011687:Purines; C526550:9H-purine-9-propanamine, 6-amino-8-((6-iodo-1,3-benzodioxol-5-yl)thio)-N-(1-methylethyl)-
keywords: ACN; ADP; ATP; Forced degradation; HPLC validation; Hsp; Impurity characterization; NSC 750424; PDA; PU-H71; Q-TOF; RRT; S/N; Stability study; TFA; acetonitrile; adenosine diphosphate; adenosine triphosphate; heat shock protein; photodiode array; quadrupole time-of-flight; relative retention time; signal to noise ratio; trifluoroacetic acid