Beyond GLP-1: Preclinical hurdles and solutions for next-generation peptide therapeutics

The success of GLP-1 receptor agonists/analogues has transformed metabolic disease treatment while demonstrating the potential of engineered peptides to deliver long-acting, highly targeted therapies. This breakthrough has accelerated peptide drug development across multiple therapeutic areas, including cardiovascular disease, inflammation, and oncology. However, as many teams develop next-generation peptide candidates, they’re finding that applying GLP-1 design principles requires more sophisticated preclinical approaches than expected.

Peptide therapeutics offer distinct advantages, including high specificity, tunable pharmacokinetics, and modular structures, but present unique development challenges compared to small molecules or antibodies. These challenges particularly emerge in ADME (absorption, distribution, metabolism, excretion), where structural modifications for enhanced stability can unexpectedly alter metabolic pathways, clearance mechanisms, and tissue distribution patterns.

The development experience with GLP-1 analogues offers critical insights for navigating these ADME challenges in contemporary peptide programmes.

Half-life extension strategies and their in vitro ADME implications

The engineering innovations that paved the way for GLP-1 analogues to achieve weekly, and now potentially monthly, dosing have inspired a new generation of long-acting peptide therapeutics. Common approaches include lipidation or fatty-acid conjugation to enhance albumin binding, backbone modifications, or incorporation of non-natural amino acids to increase metabolic stability, cyclic structures to improve stability, and depot-like formulations to extend absorption following subcutaneous administration.

These strategies provide clear clinical benefits, but also necessitate deeper characterisation in in vitro ADME and early in vivo pharmacokinetics (PK), particularly with respect to:

• Species-dependent plasma stability
• Unexpected cleavage pathways introduced by linkers or modified residues
• Altered renal handling due to prolonged systemic residence
• Distribution patterns influenced by reversible albumin binding or tissue sequestration

Standard in vitro assays (plasma stability, hepatocyte/S9 stability, permeability, and protein binding) remain essential, but may need adaptation for long-acting peptides, which often generate more complex metabolite profiles. Early integration of metabolite identification is critical, as is ensuring bioanalytical methods can reliably detect both parent compounds and relevant fragments.

Advancing peptide in vivo ADME studies: Practical considerations

Current approaches to studying the in vivo ADME of peptides have evolved significantly since the development of GLP-1 therapeutics. Radiolabelling techniques now serve as a robust platform for comprehensive ADME investigations. Aligning with in vitro ADME characteristics, in vivo ADME studies present three major practical challenges:

1. Detection sensitivity: The high molecular weight and low doses of peptides can limit signal detection. Implementation of multi-carbon (≥4 atoms) labelling strategies focused on lipid side chains or linker regions can enhance sensitivity.
2. Incomplete recovery: Prolonged systemic exposure leads to slow elimination. Extending sampling schedules can ensure the complete collection of excretory matrices for accurate mass balance determination.
3. Metabolite identification: Low circulating concentrations complicate structural elucidation. Including parallel high-dose groups can help facilitate metabolite detection and characterisation.

Originally pioneered for GLP-1 analogues, these radiolabelling strategies now serve as a gold standard for peptide ADME characterisation. Their ability to track both parent compounds and metabolites at pharmacologically relevant concentrations makes them indispensable for translational prediction.

Integrating ADME insights into toxicology programmes

Peptide toxicology presents unique considerations, many of which stem from the candidate’s ADME profile. Changes in absorption kinetics, albumin binding, or metabolic stability can influence dose-range finding outcomes, exposure margin calculations, delayed onset or prolonged recovery trends, and interpretation of target-organ toxicity.

For long-acting peptide modalities, it is imperative to align early ADME results with toxicology study design. This can include:

• Ensuring the bioanalytical method used in DMPK continues into toxicology studies to enable clear parent/metabolite interpretation.
• Selecting dose-escalation schemes that reflect expected accumulation or depot release.
• Incorporating exposure-response analyses where feasible to distinguish pharmacology from toxicity.
• Determining whether metabolites of interest appear at meaningful levels in toxicology species.

While the GLP-1 analogue development experience provides valuable insights, particularly regarding renal elimination, sustained systemic exposure, and minimal CYP involvement, these characteristics cannot be universally extrapolated to other peptide classes. Emerging constructs featuring dual mechanisms, unconventional linker systems, or high hydrophobicity often exhibit distinct ADME behaviours, requiring tailored study designs for accurate preclinical assessment.

In short, effective peptide toxicology relies on a disciplined sequence that includes in vitro ADME, targeted in vivo DMPK, and focused toxicology with data continuity across the entire workflow.

Future directions in peptide development

Three key trends are shaping the future of preclinical strategies for peptide development. First, peptide engineering is outpacing traditional ADME workflows, resulting in the establishment of radiolabelled studies as essential for complex peptide modalities. In addition, comprehensive preclinical characterisation is becoming a key differentiator, helping teams advance candidates with safety and metabolism profiles that are better understood.

The next wave of peptide therapeutics will require earlier, more strategic application of rigorous preclinical science, particularly in understanding structure-disposition relationships, building on lessons from GLP-1 development while addressing new scientific frontiers.

About the author

Lingling Zhang, PhD, is site head and senior director, DMPK Department, at WuXi AppTec, Nanjing, where she oversees radiolabelled ADME research and site operations. She has more than a decade of experience in mass balance studies, tissue distribution, and metabolite identification, supporting global IND and NDA submissions.