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Differential stability of antigenic MHC class I-restricted synthetic peptides
C Widmann1, J L Maryanski, P Romero
1Institute of Biochemistry, University of Lausanne, Epalinges, Switzerland.
Insights
Synthetic peptide stability varies significantly, impacting immune responses. Serum proteolysis is a key factor in peptide inactivation, influencing their effectiveness in vivo for immunotherapy applications.
Area of Science:
- Immunology
- Molecular Biology
- Biochemistry
Background:
- Synthetic peptides are crucial for studying T-cell responses.
- Understanding peptide stability is vital for immunotherapy development.
Purpose of the Study:
- To assess the in vitro and in vivo stability of synthetic antigenic peptides.
- To investigate factors influencing peptide inactivation, particularly serum proteolysis.
Main Methods:
- Quantifying peptide inactivation by titrating sensitization of target cells for CTL lysis.
- Assessing peptide stability at different temperatures and in the presence/absence of serum.
- Utilizing protease inhibitors and radiolabeled peptides to identify mechanisms of inactivation.
Main Results:
- Significant variation in peptide inactivation observed, ranging from minimal to over 10,000-fold loss of activity.
- Serum inactivation was identified as a major factor, inhibited by protease inhibitors.
- In vivo peptide persistence correlated directly with in vitro stability findings.
Conclusions:
- Differential stability of synthetic peptides is a critical consideration for immunotherapy.
- Serum proteolysis significantly impacts peptide efficacy in vivo.
- Peptide stability directly influences the manipulation of immune responses.
Abstract:
Various synthetic peptides recognized as Ag by CTL in the context of MHC class I molecules were tested for stability in vitro and in vivo. Peptide inactivation in vitro was quantitated by titrating the amount of peptide required to sensitize target cells for lysis by specific CTL clones. The degree of inactivation after overnight incubation at 37 degrees C varied widely among a series of antigenic peptides. Some were nearly unaffected, whereas others lost activity by more than 100-fold or even 10,000-fold. However, no correlation was found between susceptibility to serum inactivation and antigenic potency as measured in short term cytolytic assays. No inactivation occurred at 4 degrees C, or at 37 degrees C in the absence of serum, under the conditions used. Serum inactivation most likely involved proteolysis because it could be inhibited by protease inhibitors. Moreover, presumed cleavage products of a radiolabeled susceptible peptide could be visualized by TLC. In vivo, the persistence of the antigenic activity of the injected peptides, either in extracellular fluids or on tumor target cells growing in an ascites form, correlated with the degree of stability found for the peptides in vitro. The differential stability of synthetic peptides may have important consequences for attempts to manipulate the development of an immune response in vivo.