The FDA has posted a request for information and comments in the Federal Register on the subject Evaluating the Immunogenicity Risk of Host Cell Proteins in Follow-On Recombinant Peptide Products. BEBPA strongly encourages participation in this process. Comments will close on September 23, 2024.

This effort originates out of CDER and they are requesting comments on “suitable methods to detect, identify, and quantify HCPs and the minimal residual amounts of HCPs achievable in commercial lots of rPeptide products.” The FDA has expressed interest specifically in these questions:

1. What is the lowest and routinely achievable level of total HCPs across your well-controlled rPeptide manufacturing process(es), and how are they calculated/established?
2. What are the challenges in reducing HCP levels?
3. What analytical methods are currently being used to detect, identify, and quantify HCPs in a rPeptide product?
4. What is the generally achievable percent coverage of the HCP spectrum for your HCP quantification assay?
5. Are there any qualitative or quantitative characteristics of HCPs associated with a higher likelihood of adverse clinical sequelae?
6. What tools (in silico, in vitro or in vivo studies) do you currently use or plan to use to compare the potential immunogenicity risk of two products with different HCP profiles? What is your approach to risk assessment of HCPs based upon such data?

While rPeptides are smaller than the recombinant proteins that have dominated our BEBPA HCP conferences, the fundamental challenges of HCP detection and risk assessment will be similar. Just as biologics are more complex in structure and function than small-molecule pharmaceuticals, HCPs are more complex than small-molecule impurities. There is no perfect method for detecting and quantifying HCPs. The HCP ELISA is based on an immunological response that is heavily influenced by the antigenicity of the HCPs in the animal used to generate the ELISA antibodies. Mass spectrometry relies on the ability of an HCP to be digested into smaller peptides, getting these peptides to chromatograph, and then ionize and fragment in the mass spectrometer, and mapping the MS/MS data to a protein in a proteome database for the host organism. ELISA and mass spectrometry have very little in common with respect to their underlying biochemical/biophysical mechanisms, and they are usually run in multi-analyte format, which calls into question what is really being quantitated by each method and how quantitative results can be correlated between the methods. Finally, as has been repeatedly emphasized at our conferences –  the HCP ELISA is not a purity assay – it should be viewed as a control system assay. Despite all these problems, HCP ELISAs and mass spectrometry are being used successfully to characterize HCPs in biologics. The USP has published <1132> Residual Host Cell Protein Measurement in Biopharmaceuticals, which focuses on HCP ELISA methods, and USP will release <1132.1> Residual Host Cell Protein Measurement in Biopharmaceuticals by Mass Spectrometry in May 2025.   Both of these documents provide a foundation for answering the questions raised by the FDA about HCP analytics.

Regarding risk assessment, this request from the FDA focuses on immunogenicity of HCPs. In 2015 de Zafra et al. published a risk assessment framework that accounts for HCP identity, clinical experience and exposure multiples, clinical indication, patient population, phase of development, route of administration, duration of treatment, and frequency of dosing. de Zafra et al. concluded that weaknesses in the HCP analytics (i.e., multi-analyte ELISAs and MS methods, where quantitation is imperfect) lead to the case that “it is therefore not feasible to establish a common HCP threshold for all biotherapeutic products below which all products will be safe.” Risk assessment of HCPs is therefore not a straightforward task with clear linear pathways between HCP detection and patient safety.

If your company is involved in HCP analysis/assessment in rPeptides, we encourage you to respond to the FDA’s request for comments. Also, we encourage you to submit an abstract to the 2025 BEBPA Host Cell Protein Conference to discuss any case studies you may have on this subject.