P3 - Modification of Flupirtine for the Treatment of Chemotherapy-Induced Neuropathic Pain

Introduction: : Chemotherapy induced neuropathic pain (CINP) is the most common neurological syndrome in patients undergoing antineoplastic therapy. CINP is predominantly a sensory neuropathy with various symptoms, most prominent and classical among them include tingling, hypersensitivity to extreme temperatures, numbness and pain. When CINP occurs, it leads to treatment interruption or dose modification which in turn leads to a negative impact on the success of the treatment. Current treatment strategies are mainly based on modification of the chemotherapy dose, treatment cycle, timing, and duration. However, in most patients, CINP persists or even intensifies after termination of chemotherapy, and thus can only be treated through additional therapies. At present, there are no clinically established treatments capable of reversing or preventing the neurological damage associated with CINP. Therefore, a critical and unmet medical need exists for the development of potent therapeutic agents for CINP. Flupirtine is a non-opioid and non-steroidal anti-inflammatory drug analgesic used in various disease conditions where neuronal hyperexcitability is involved. Flupirtine has demonstrated efficacy in a range of pain conditions, including musculoskeletal pain, neuropathic pain, and postoperative pain in clinical studies. Despite its initial success, flupirtine faced safety concerns related to rare but severe hepatotoxicity. This led to restrictions on its use and its withdrawal from the European market. Currently, flupirtine is still sold under various brand names in some countries and regions. The current study focuses on the design and synthesis of flupirtine analogs to maintain the pain relief capacity while significantly reducing liver toxicity adverse effects.

Materials and

Methods: : One postulated mechanism attributed to the hepatotoxicity for flupirtine is metabolism to the reactive azaquinone dimine intermediate. During oxidative metabolism of flupirtine, it produces azaquinone diimine (and its ortho-azaquinone diimine isomer), a known hepatotoxic metabolite and major cause of its hepatotoxicity, similar to the N-acetyl-p-benzoquinone imine produced during the metabolism of acetaminophen. We have developed a library of multifunctional aromatic carbamates based around the flupirtine scaffold and the related phenyl bioisostere retigabine. Among the many compounds synthesized, we identified compound 5 as a promising candidate. We further evaluated compound 5 in vitro cytotoxicity by using CCK-8 assay and in vivo hepatotoxicity in mice after intraperitoneal (i.p.) injection. To enable preclinical study, CINP rodent models have been well established by repetitive administration of a certain chemotherapy agent, such as paclitaxel (PTX), a widely used chemotherapeutical agent for the treatment of a wide range of cancers. We developed a mouse CINP model by i.p. injection of PTX on four alternate days (day 1, 3, 5, and 7) at a dose of 8 mg/kg, so as to reach a cumulative dose of 32 mg/kg. On the same days as PTX injections, compound 5 was administered i.p. at a dose of 10 mg/kg. To evaluate CINP development and the potential therapeutic effects of compound 5, we conducted a series of behavioral tests, including the von Frey test for mechanical allodynia, the hotplate test for thermal nociception, and the acetone test for cold sensitivity.

Results, Conclusions, and Discussions:: We demonstrated that the cytocompatibility of compound 5 was concentration-dependent. Following i.p. injection in mice at a dose of 10 mg/kg for 24 hours, there were no significant differences in the levels of blood urea nitrogen, aspartate aminotransferase, or creatinine between the groups treated with compound 5 and those receiving the vehicle (PBS). These findings suggest that chemical modification of the flupirtine scaffold can reduce hepatotoxicity while maintaining biocompatibility. We found that compound 5 was able to ameliorate CINP as evidenced by significantly increasing the paw withdrawal threshold and latency in response to mechanical and heat stimulation and decreased the response duration to the cold stimulation. Indeed, in the heat and cold stimulation experiments compound 5 restored response to near baseline control levels. Taken together, these results demonstrated compound 5 as a promising candidate for the treatment of CINP, offering both improved safety and therapeutic efficacy over the parent compound flupirtine. Future studies will focus on elucidating the molecular mechanisms underlying compound 5's protective effects against CINP. Additionally, pharmacokinetic and biodistribution analyses will be conducted to optimize dosing regimens and understand tissue-specific exposure. Long-term safety assessments and efficacy studies will also be evaluated.

Acknowledgements and/or References (Optional):: UN Collaboration Initiative Grant, LB606 Nebraska Stem Cell Grant, UNMC Center for Drug Design and Innovation (CDDI) Seed Grant, NIH R21CA298918