Efficacy and preventive potential of apitoxin and melittin in pharmacoacupuncture to control neuropathic pain: Safety, determination of action mechanisms and search for active fragments

• Grant number: 24/04901-7
• Support Opportunities: Scholarships in Brazil - Doctorate
• Start date: December 01, 2024
• End date: July 31, 2028
• Field of knowledge: Health Sciences - Collective Health - Public Health
• Principal Investigator: Gisele Picolo
• Grantee: Ana Maria Boaventura de Oliveira
• Host Institution: Instituto Butantan. Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Associated research grant: 13/07467-1 - CeTICS - Center of Toxins, Immune-Response and Cell Signaling, AP.CEPID

Abstract

Chronic pain is a multifactorial disease affecting approximately 20% of the global population, being a major cause of suffering and disability, since it interferes with daily activities. Despite the availability of various pharmaceuticals in medical practice, treatment remains challenging since patients rarely achieve complete reversal of pain, and often suffer from the diverse adverse effects of medication use. Due to these challenges, the adoption of Integrative and Complementary Practices (ICP) alongside traditional pharmacological therapy has proven beneficial in improving quality of life and reducing pain in chronic patients, especially since these practices, including acupuncture, have been included in public health systems such as the Brazilian Unified Health System (SUS).Among these techniques, pharmacopuncture has been highlighted. In pharmacopuncture, which involves the application of drugs into acupuncture points-specific points in acupuncture-analgesia is achieved with smaller doses than those required in systemic treatments, thereby potentially enhancing the expected effects and reducing medication-related adverse effects. Pharmacopuncture combined with apitoxin, the venom produced by the honeybee Apis mellifera, has gained attention due to its potential antinociceptive properties, especially after apitherapy was included in the National Policy on Integrative and Complementary Practices (PNPIC).Previous trials conducted by the group have demonstrated the efficacy not only of apitoxin but also of melittin, the most abundant peptide in apitoxin. Both have proven attractive when used in conjunction with pharmacopuncture for promoting prolonged analgesia and improving body distribution in neuropathic pain models. However, considering the toxicity associated with bee venom and some of its components, ensuring the therapy's safety and specifically searching for smaller peptide portions for evaluation of analgesic activity and subsequent formulation of a new pharmaceutical formula are still necessary.Therefore, the objective of this study is to verify the safety of performing pharmacopuncture with melittin, investigate the mechanisms associated with analgesia through this route, and identify a peptide portion derived from melittin that exhibits analgesic activity for kinetic and behavioral assays. For safety trials, apitoxin or melittin will be applied in multiple treatments, and at the end, organs will be collected for histopathological evaluation, and blood will be analyzed for blood cell counts and immunogenicity assessment. Concurrently, to investigate the mechanisms underlying analgesia, two strategies will be employed: the first involves the use of specific antagonists of analgesic pathways (serotonergic, purinergic, opioidergic, and endocannabinoid), administered prior to toxin treatments; for the second strategy, collected material will undergo immunoassays to determine levels of endogenous opioids involved in pharmacopuncture analgesia, as well as the role of the somatostatin pathway.Subsequently, to identify the peptide portion, the Expasy Peptide Mass platform will be utilized for predicting fragments generated from the parent molecule, or degradation assays will be conducted to generate fragments, followed by in vivo and in silico assays aimed at confirming the analgesic activity (local and systemic) and determining the pharmacokinetic and physicochemical characteristics of the peptides.By the end of the project, we aim to establish a pharmacopuncture protocol capable of preventing the onset of chronic pain, determine the mechanisms involved in this effect, and identify melittin-derived peptide portions capable of promoting analgesia and generating new analgesic compounds.