Site selective C-H functionalization of Mitragyna alkaloids reveals a molecular switch for tuning opioid receptor signaling efficacy.

Mitragynine (MG) is the most abundant alkaloid component of the psychoactive plant material "kratom", which according to numerous anecdotal reports shows efficacy in self-medication for pain syndromes, depression, anxiety, and substance use disorders. We have developed a synthetic method for selective functionalization of the unexplored C11 position of the MG scaffold (C6 position in indole numbering) via the use of an indole-ethylene glycol adduct and subsequent iridium-catalyzed borylation. Through this work we discover that C11 represents a key locant for fine-tuning opioid receptor signaling efficacy. 7-Hydroxymitragynine (7OH), the parent compound with low efficacy on par with buprenorphine, is transformed to an even lower efficacy agonist by introducing a fluorine substituent in this position (11-F-7OH), as demonstrated in vitro at both mouse and human mu opioid receptors (mMOR/hMOR) and in vivo in mouse analgesia tests. Low efficacy opioid agonists are of high interest as candidates for generating safer opioid medications with mitigated adverse effects.

Electroacupuncture analgesia is enhanced in transgenic nociceptin/orphanin FQ knock-out mice.

OBJECTIVE: The modulatory effects of nociceptin/orphanin FQ (OFQ) on acupuncture (EA, a modern version of acupuncture)-induced analgesia are still controversial. Transgenic OFQ knock-out mice provide us a useful tool to investigate the role of endogenous OFQ in EA analgesia. The present study aims to investigate the role of OFQ in the EA-induced analgesia with OFQ knock-out mice. METHODS: Acupoints were selected as "Zusanli" (ST36) and "Sanyinjiao" (SP6), EA parameters were as follows: constant current output, rectangular (square) wave pulses, increased intensities of 0.5, 0.7, 0.9 mA with stepwise fashion, 10 min for each intensity, frequency of 100 Hz (with 0.2 ms as pulse width) or 2 Hz (with 0.6 ms as pulse width). Tail flick latency (TFL) evoked by radiant heat was used to evaluate the change of pain threshold before, during and after EA application. The knock-out mice were divided randomly into 3 groups: needling control, EA at 100 Hz and 2 Hz. Wild-type mice of littermates of were used as Control. RESULTS: It was found that OFQ knock -out mice had a longer basal thermal threshold; EA had enhanced analgesic effect in the knock-out mice than in wild-type Control mice. CONCLUSION: Endogenous OFQ might be algesic in basal condition and antagonize EA analgesia.

The role of prohormone convertase-2 in hypothalamic neuropeptide processing: a quantitative neuropeptidomic study.

Prohormone convertase (PC) 1/3 and 2 are involved in the generation of neuropeptides from their precursors. A quantitative peptidomic approach was used to explore the role PC2 plays in the processing of hypothalamic peptides. In this approach, extracts from mice lacking PC2 activity and from wild-type littermates were labeled with isotopic tags, combined, fractionated on a reverse phase HPLC column, and analyzed by electrospray ionization mass spectrometry. Altogether, 53 neuropeptides or other peptides derived from secretory pathway proteins were identified and sequenced using tandem mass spectrometry. These peptides arise from 21 distinct proteins: proenkephalin, proopiomelanocortin, prodynorphin, protachykinin A and B, procholecystokinin, promelanin-concentrating hormone, proneurotensin, proneuropeptide Y, provasopressin, pronociceptin/orphanin, prothyrotropin-releasing hormone, cocaine- and amphetamine-regulated transcript, chromogranin A and B, secretogranin II, prohormone convertase 1 and 2, propeptidyl-amidating monooxygenase, and proteins designated proSAAS and VGF. Approximately one third of the peptides found in wild-type mice were not detectable in PC2 knock-out mice, and another third were present at levels ranging from 25 to 75% of wild-type levels. Comparison of the cleavage sites suggests that sequences with a Trp, Tyr and/or Pro in the P1' or P2' position, or a basic residue in the P3 position, are preferentially cleaved by PC2 and not by other enzymes present in the secretory pathway.