Dottorato in FARMACOLOGIA E TOSSICOLOGIA

Titolo della tesi: Pharmacological characterization of new compounds and herbal derivatives for pain and inflammation treatment

Introduction: Despite the availability of several analgesic and anti-inflammatory options, the effective treatment of pain and inflammation is still a challenge for clinicians, and the balance between efficacy and safety aspects remains both crucial and difficult. Thus, the identification of new potential targets which may affect pain and inflammatory processes is becoming an urgent clinical and therapeutic need. This scientific project derives from a close collaboration with various national and foreign research groups, in order to establish an efficient and highly multidisciplinary approach to investigate the pharmacological profile of new compounds and herbal derivatives with significant antinociceptive and antinflammatory effects to be used for the treatment of pain and inflammation. Materials and Methods: The research approach involves the in vivo studies of natural (ammonium glycyrrhizinate; rubiscolin-6 and soymorphin-6) and non-natural compounds (C-terminal amides derivatives of rubiscolin-6 and soymorphin-6; six analogues of endomorphin-2 and DAPEA; two compounds obtained in silico studies; a series of lonidamine with high CB1 receptor affinity and new pH-sensitive formulation of niosomes containing Polysorbate 20 derivatized by Glycine and loaded with ibuprofen) using murine models of pain and inflammation. All experiments were carried out using male CD-1 mice weighing 25 g. Results: The results suggest the potential use of ammonium glycyrrhizinate for clinical treatment of pain and/or inflammatory-related diseases, included diabetic peripheral neuropathy. The results obtained by in vivo studies testing two natural compounds, rubiscolin-6 and soymorphin-6, and their C-terminal amides derivatives, show that rubiscolin-6 was able to increase the nociceptive threshold to thermal stimuli after supraspinal administration, but was ineffective after subcutaneous administration, in an experimental paradigm of chemical-induced nociception as the formalin test. On the other hand, instead, its derivatives (rubiscolin-6 C-amide) centrally administered demonstrates a strong antinociceptive effect higher than rubiscolin-6, and it is effective after subcutaneous administration. Both soymorphin-6 and its derivative soymorphin-6 C-amide induced a robust antinociceptive effect after central administration in tail flick test but both peptides have not been able to change the behavioral response to chemical-induced nociception in the formalin test. The in vivo antinociceptive profile of new six tetrapeptide models containing α-amino-γ-lactam of Freidinger in position 2 and 3, as analogues of endomorphin-2 (EM-2) and DAPEA, revealed that one of them (peptide A2D) exhibited a strong antinociceptive effect in vivo after intracerebroventricular administration, performing better than the parent compounds EM-2 and DAPEA. Peptide A3D is also able to produce antinociceptive effect both in the acute and in the inflammatory phase of the formalin test. The in vivo results obtained from two compounds (tripeptides 6 and 11) derived from previous in silico studies revealed their ability to induce an antinociceptive effect after intracerebroventricular and subcutaneous administrations in the tail flick and formalin tests, respectively. Subsequently, it was analyzed the in vivo orexant/anorexant and antinociceptive profile of a series of lonidamine joined Leu-, tert/Leu- and Val-amino acids with different C-terminal functional groups (LONI 1-4,11) as novel compounds with high CB1 receptor affinity and selectivity with different biological activity depending on the C-terminal substitution and amino acid residues and endowed with activities. The results demonstrated that LONI2 and LONI4 were able to inhibit food intake (anorexant effect) consistent with an inverse agonism at CB1 receptors. On the other hand, LONI11, an agonist towards cannabinoid receptors CB1 and CB2, proved to carry out a significant orexant an antinociceptive effect at the central and periphery levels. Finally, a new pH-sensitive formulation of niosomes containing Polysorbate 20 derivatized by Glycine and loaded with ibuprofen (NioIbu) was in vivo tested, using models of pain and inflammation. The results demonstrated that NioIbu, administered 2h before testing, reduced nociception, whereas the free form of ibuprofen was ineffective. Conclusion: In conclusion, these findings create prospects for conducting clinical studies with the compounds presented, which show in fact their effectiveness for chronic pain and inflammation control.