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Liquid Chromatography-High Resolution Mass Spectrometry for Peptide Drug Quality Control

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Abstract

A liquid chromatography-high resolution mass spectrometry (LC-HRMS) method was developed using three peptide drugs: salmon calcitonin, bivalirudin, and exenatide as model systems to assess the suitability of this approach for monitoring peptide drug product quality. Calcitonin and its related impurities displayed linear responses over the range from 0.1 to 10 μM (R 2 values for calcitonin salmon, Glu14-calcitonin, and acetyl-calcitonin were 0.995, 0.996, and 0.993, respectively). Intra-assay precision in terms of relative standard deviation (%RSD) was less than 10% at all tested concentrations. The accuracy of the method was greater than 85% as measured by spiking 0.1, 0.3, and 1% of Glu14-calcitonin and acetyl-calcitonin into a stock calcitonin solution. Limits of detection for calcitonin, Glu14-calcitonin, and acetyl-calcitonin were 0.02, 0.03, and 0.04 μM, respectively, indicating that an impurity present at less than 0.1% (0.1 μM) of the drug product API concentration (107 μM) could be detected. Method validation studies analyzing bivalirudin and exenatide drug products exhibited similar results to calcitonin salmon in regard to high selectivity, sensitivity, precision, and linearity. Added benefits of using LC-HRMS-based methods are the ability to also determine amino acid composition, confirm peptide sequence, and quantify impurities, even when they are co-eluting, within a single experiment. LC-HRMS represents a promising approach for the quality control of peptides including the measurement of any peptide-related impurities. While the development work performed here is focus on peptide drug products, the principles could be adapted to peptide drug substance.

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Abbreviations

AA:

Amino acid

AC:

N-acetyl-calcitonin salmon

API:

Active pharmaceutical ingredient

BA:

Beta-aspartic acid-bivalirudin

BV:

Bivalirudin

BV[12–20]:

12–20 fragment of bivalirudin

sCT:

Calcitonin salmon

DCM:

Dichloromethane

DG:

Des-glycine-bivalirudin

EIC:

Extracted ion chromatogram

ET:

Exenatide

Glu14 :

Glu14-calcitonin salmon

Glu20 :

Glu20-calcitonin salmon

LC-MS:

Liquid chromatography mass spectrometry

LC-HRMS:

Liquid chromatography-high resolution mass spectrometry

LOD:

Limit of detection

LOQ:

Limit of quantification

MEW:

Mass extraction window

MS/MS:

Tandem mass spectrometry

NDA/ANDA:

New Drug Application/Abbreviated New Drug Application

NMT:

Not more than

PG:

Plus-glycine-bivalirudin

SPPS:

Solid-phase peptide synthesis

TIC:

Total ion chromatogram

References

  1. Ermerand J, Vogel M. Applications of hyphenated LC-MS techniques in pharmaceutical analysis. Biomed Chromatogr. 2000;14:373–83.

    Article  Google Scholar 

  2. Kasparand AA, Reichert JM. Future directions for peptide therapeutics development. Drug Discov Today. 2013;18:807–17.

    Article  Google Scholar 

  3. Dorpe VS, Verbeken M, Wynendaele E, De Spiegeleer B. Purity profiling of peptide drugs. J Bioanal Biomed. 2011;15:1–15.

  4. Gucinskiand AC, Boyne 2nd MT. Identification of site-specific heterogeneity in peptide drugs using intact mass spectrometry with electron transfer dissociation. Rapid Commun Mass Spectrom: RCMS. 2014;28:1757–63.

    Article  Google Scholar 

  5. Ramanathan R, Jemal M, Ramagiri S, Xia YQ, Humpreys WG, Olah T, et al. It is time for a paradigm shift in drug discovery bioanalysis: from SRM to HRMS. J Mass Spectrom: JMS. 2011;46:595–601.

    Article  CAS  PubMed  Google Scholar 

  6. Ramanathan R, Korfmacher W. The emergence of high-resolution MS as the premier analytical tool in the pharmaceutical bioanalysis arena. Bioanalysis. 2012;4:467–9.

    Article  CAS  PubMed  Google Scholar 

  7. Morin LP, Mess JN, Garofolo F. Large-molecule quantification: sensitivity and selectivity head-to-head comparison of triple quadrupole with Q-TOF. Bioanalysis. 2013;5:1181–93.

    Article  CAS  PubMed  Google Scholar 

  8. Wei H, Tymiak AA, Chen G. High-resolution MS for structural characterization of protein therapeutics: advances and future directions. Bioanalysis. 2013;5:1299–313.

    Article  CAS  PubMed  Google Scholar 

  9. Fung EN, Jemal M, Aubry AF. High-resolution MS in regulated bioanalysis: where are we now and where do we go from here? Bioanalysis. 2013;5:1277–84.

    Article  CAS  PubMed  Google Scholar 

  10. Dillenand L, Cuyckens F. High-resolution MS: first choice for peptide quantification? Bioanalysis. 2013;5:1145–8.

    Article  Google Scholar 

  11. Ermerand J, Kibat P-G. A quality concept for impurities during drug development—use of the hyphenated LC–MS technique. Pharm Sci Technol Today. 1998;1:76–82.

    Article  Google Scholar 

  12. Ermer J. The use of hyphenated LC–MS technique for characterisation of impurity profiles during drug development. J Pharm Biomed Anal. 1998;18:707–14.

    Article  CAS  PubMed  Google Scholar 

  13. Chopra S, Pendela M, Hoogmartens J, Van Schepdael A, Adams E. Impurity profiling of capreomycin using dual liquid chromatography coupled to mass spectrometry. Talanta. 2012;100:113–22.

    Article  CAS  PubMed  Google Scholar 

  14. Chopra S, Van Schepdael A, Hoogmartens J, Adams E. Characterization of impurities in tylosin using dual liquid chromatography combined with ion trap mass spectrometry. Talanta. 2013;106:29–38.

    Article  CAS  PubMed  Google Scholar 

  15. Gucinskiand AC, Boyne 2nd MT. Evaluation of intact mass spectrometry for the quantitative analysis of protein therapeutics. Anal Chem. 2012;84:8045–51.

    Article  Google Scholar 

  16. Chesnut 3rd CH, Azria M, Silverman S, Engelhardt M, Olson M, Mindeholm L. Salmon calcitonin: a review of current and future therapeutic indications. Osteoporos Int. 2008;19:479–91.

    Article  PubMed  Google Scholar 

  17. USP 36-NF 31 (United States Pharmacopeia Convention). Calcitonin salmon, calcitonin salmon injection, and calcitonin nasal solution. USP;2013. p. 2736-2741.

  18. Reedand MD, Bell D. Clinical pharmacology of bivalirudin. Pharmacotherapy. 2002;22:105S–11S.

    Article  Google Scholar 

  19. Cvetkovicand RS, Plosker GL. Exenatide: a review of its use in patients with type 2 diabetes mellitus (as an adjunct to metformin and/or a sulfonylurea). Drugs. 2007;67:935–54.

    Article  Google Scholar 

  20. Liu B, Dong Q, Shi L, Wang M, Li C, Wu Y, et al. Development and validation of a reverse-phase high performance liquid chromatography method for determination of exenatide in poly(lactic-co-glycolic acid) microspheres. Chem Res Chin Univ. 2010;26:33–7.

    Google Scholar 

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Acknowledgments

Internal funding for this work was provided by the CDER Critical Path Program and the Office of Generic Drugs (MTB).

Disclaimer

The findings and conclusions of this article have not been formally disseminated by the Food and Drug Administration and should not be construed to represent any agency determination or policy.

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Authors and Affiliations

  1. Division of Pharmaceutical Analysis, Office of Testing and Research, Center for Drug Evaluation and Research, Food and Drug Administration, 645 S. Newstead Avenue., St Louis, Missouri, 63110, USA

    Kui Zeng, Ilan Geerlof-Vidavisky, Ashley Gucinski & Michael T. Boyne II

  2. Office of Research Standards, Office of Generic Drugs, Center for Drug Evaluation and Research, Food and Drug Administration, 10903 New Hampshire Ave, Silver Spring, Maryland, USA

    Xiaohui Jiang

  3. Center for Drug Evaluation and Research, Office of Testing and Research, Division of Pharmaceutical Analysis, United States Food and Drug Administration, 10903 New Hampshire Avenue, Silver Spring, Maryland, 20993, USA

    Michael T. Boyne II

Authors
  1. Kui Zeng
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  2. Ilan Geerlof-Vidavisky
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  3. Ashley Gucinski
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  4. Xiaohui Jiang
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  5. Michael T. Boyne II
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Correspondence to Michael T. Boyne II.

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Zeng, K., Geerlof-Vidavisky, I., Gucinski, A. et al. Liquid Chromatography-High Resolution Mass Spectrometry for Peptide Drug Quality Control. AAPS J 17, 643–651 (2015). https://doi.org/10.1208/s12248-015-9730-z

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