Formulation development and analytical evaluation of α-CGRP 8-37 for a potential subcutaneous psoriasis therapy

Abstract

α-CGRP 8-37 is a peptide fragment of calcitonin gene-related peptide and is commonly used as a CGRP antagonist in pharmacological research. CGRP signaling has been discussed in relation to neuroimmune mechanisms in psoriasis, but a practical development of α-CGRP 8-37 requires analytical methods and formulation strategies that can handle peptide instability, adsorption, and matrix effects. The aim of this thesis was to develop and evaluate analytical and formulation approaches for α-CGRP 8-37, with emphasis on subcutaneous formulation relevance and pilot blood-matrix method development.
An HPLC method with UV detection at 214 nm was used for formulation and stability samples. LC-MS was used for pilot blood-matrix experiments. Formulation studies evaluated α-CGRP 8-37 in aqueous and excipient-containing vehicles at different peptide concentrations during room-temperature storage. An adsorption experiment compared apparent peptide recovery in BSA-coated and non-coated vials. Blood-matrix experiments tested dilution and protein-removal ap-proaches, including direct dilution into aqueous formic acid, inhibitor-containing formic acid, and organic solvent, with and without SPE cleanup.
The formulation data showed concentration-dependent differences in apparent stability. At day 41, 10 and 20 mg/mL samples retained mean signals of 55.0% and 58.2% across tested condi-tions, while 2 mg/mL samples retained a mean signal of 15.3%, with several conditions becom-ing undetectable. BSA-coated vials increased apparent recovery in several low-concentration samples, supporting adsorption as a likely contributor to peptide loss. In the first whole-blood LC-MS pilot, CGRP and CGRP 8-37 signals for 30 second old samples were approximately 19.1% and 24.5% of blank-spiked controls, and signals decreased by 84.6% and 79.5%, respec-tively, from 30 seconds to 10 min. In the SPE-based experiment, enzyme inhibitor containing unspiked samples gave a high CGRP 8-37 signal while CGRP remained low. Likely indicating that sample preparation chemistry strongly affected the measured peptide signal.
The HPLC method was suitable for formulation screening, while the current blood-matrix LC-MS workflow was not ready for quantitative volunteer or patient analysis. The blood results should be interpreted as method development findings rather than biological concentration data. Future work should prioritize internal standards, controlled quenching immediately after blood draw, recovery studies, matrix-factor assessment, pH and osmolality testing, and longer formu-lation stability studies.

Popular Abstract

Making a fragile peptide measurable and usable as an injectable drug
Psoriasis is described as a skin disease, the immune system and sensory nerves help shape the disease process. Calcitonin gene-related peptide, shortened to CGRP, is one of the nerve-derived signaling molecules that is linked to skin inflammation. A shortened fragment of the peptide, called CGRP 8-37, can block some signaling caused by the whole version of CGRP. That makes it interesting as a research molecule and as a possible starting point for a future treatment strategy.
Before a peptide like CGRP 8-37 can become a realistic medicine, two practical questions need to be answered. First, can the peptide be measured, detected and observed in a reliable manner? Second, can it be formulated in a liquid that stays stable and works for injection under the skin? This thesis focused on those two questions.
The development part tested how CGRP 8-37 behaved after being dissolved in different liquid vehicles and observed what happened when they were stored over time, regarding stability. The peptide did not behave like a table salt or sugar dissolved in water. Its apparent stability depend-ed on concentration, the vehicle used to dissolve the peptide and contact with container surfaces that the samples came into contact with, such as storage vials. Low-concentration samples often lost measurable signal more strongly than higher-concentration samples. This led to an experi-ment where a protein called BSA was used to coat the inside of the storage vials, acting as a barrier between the peptide and the glass vial. The goal was to see if the peptide was getting stuck to the vials and therefore not getting detected when the samples were getting measured. In theory this matters more with lower concentration samples since only a set amount of peptide can get stuck to the containers before there is no more space for the peptide to stick to, for high-er concentration samples this amount could be neglectable, but for smaller concentration samples most of the peptide could get stuck to the container vials. The results were that samples stored in BSA-coated vials gave higher signal in several conditions, which points toward that the peptide is sticking to glass or vial surfaces as one source of peptide loss.
The analytical part used HPLC, a machine that can be used to separate and quantify chemicals. For formulation samples and LC-MS, a machine that can separate and quantify like the HPLC, but can also identify chemicals, for blood-matrix experiments. HPLC worked as a practical tool for formulation and stability screening. In pilot experiments, CGRP and CGRP 8-37 signals changed rapidly after blood contact. In one experiment, signal decreased strongly between 30 seconds and 10 minutes. In another, an inhibitor-containing preparation gave unexpectedly high CGRP 8-37 signal while CGRP stayed low. These results do not yet give reliable concentra-tions in blood. They however show why blood sampling needs tighter control before volunteer or patient samples are analyzed.
The main conclusion is that CGRP 8-37 development is dependent on analytical control. A sta-ble formulation cannot be judged if the method loses peptide to surfaces or sample preparation. Blood levels cannot be interpreted if the peptide changes during handling. The next research step should combine rapid blood quenching, internal standards for LC-MS, pH and osmolality test-ing, and long-term formulation stability studies.

• author: Magnason, Máni Þór
• supervisor: Lars Nilsson
• organization: Food Technology and Nutrition (M.Sc.)
• course: KLGM06 20261
• year: 2026
• type: H2 - Master's Degree (Two Years)
• subject: Technology and Engineering
• keywords: psoriasis, formulation, peptide, analytical methods, stability, pharmaceutical formulation
• language: English
• id: 9231394
• date added to LUP: 2026-06-10 14:17:40
• date last changed: 2026-06-10 14:17:40