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Novel innovative approaches in the management of pain

Group leader: Mahmoud Al-Khrasani

Our research group continues to carry on pain and analgesics research with a long tradition background. Although pain relief seems to be solved following a superficial notice however, currently available drugs in some pain conditions, such as neuropathic pain are not efficient enough, and all available painkillers have serious adverse effects.

Beside to in vivo pain models most often during our research in vitro, biological (isolated organs), biochemical (binding experiments) and immunohistochemistry assays are applied to strengthen the message of the obtained results. These studies are carried out in our laboratories and national or international cooperative research partners’ laboratories.

In vivo pain models:
In vitro modeling of pain hitherto is not solved, therefore in vivo animal experiments are still indispensable for the analysis of analgesic agents.

Models to measure acute pain:
• Tail-flick test (in rats and mice)
• Hot-plate test (conventional and rising temperature)
Models to measure inflammatory pain:
• Acute inflammatory pain: formalin test, acetic acid writhing test (duration: 1 hour)
• Subacute inflammatory pain: Carrageenan-induced inflammatory hyperalgesia (duration: 3-5 hours)
• Sub chronic inflammatory pain: Freund’s Complete Adjuvant (FCA or CFA) -induced hyperalgesia (3-7 days duration)
Models to measure neuropathic  pain:
• Seltzer’s model:  partial sciatic nerve ligation modelling mononeuropathic pain (duration: 1-4 weeks)
• Diabetic polyneuropathy animal model (advanced diabetes 3-12 weeks duration)

In the inflammatory and neuropathic pain models the developed hyperalgesia and allodynia are assessed by Randall-Selitto, DPA methods or Hargreaves (detecting change in Cold-Hot Sensitivity). To determine the analgesic effect of test compounds, the pain threshold is assessed prior to and after treatment with test compounds. In case of formalin or mouse writhing test the test compounds are preemptively administered (prior to pain stimuli). The analgesic effect is determined from base threshold and the threshold measured following drug administration. To assess development of hyperalgesia and allodynia, the pain thresholds are measured before and after pain inductions by irritant chemical, nerve damage (nerve ligation) or diabetes. Then the analgesic action of test compounds is assessed. In case of pain evoked by injection of formalin or acetic acid the pain behaviors are rapidly developed. The number of these pain behaviors are reduced by painkillers. In these models the analgesic effect is quantifiable.

Isolated organs

Isolated organs are widely used to assess the pharmocology profiles of opioid analgesics. In our labs three different isolated organs namely mouse vas deferens (MVD), rat vas deferens (RVD) and guinea pig ileum (GPI) are used for screening the novel opioid compounds. MVD hosts d>µ>k opioid receptors. RVD hosts µ-opioid receptor of low reserve, and only opioid of high efficacy could show effect on it.  GPI hosts µ and k opioid receptors. Following isolation of organs, the organs are suspended between upper and lower electrodes in organ baths of 2-10ml values. Electrical field stimulation is used to evoke the smooth contractions of these organs. Opioid agonists do inhibit the contractions of isolated organs. Applying different agonist concentrations allow us to construct concentration response curves, from which we can calculate the receptor constants of agonists. In addition, we can also use these isolated organs to assess the affinity of opioid antagonists.

Ongoing research projects:

Peripheral opioid analgesia

Project leader:  Dr. Mihály Balogh

Opioid analgesics are mostly known to produce analgesia by a central mechanism of action, however growing data support the existence of functional opioid receptors at the periphery on peripheral nociceptive fibers. This finding provides a new target for opioid analgesics. The drawback is that peripheral opioid receptors located on pain traffic points are low in reserve and only compounds with high efficacy could activate them. Most opioids used in the clinic are considered to be full agonists however, their intrinsic activity is typically far behind the clinical need. Limiting the central effects of opioids and activating the peripheral opioid receptors is beneficial to eliminate several important central side effects such as sedation, euphoria, respiratory depression as well as abuse liability, though the intestinal transit inhibition is remained. Despite of the availability of synthetic approach to produce an opioid agonist or antagonist of limited CNS penetration, the low efficacy and or affinity was the matter of debate. Cooperating with the Institute of Pharmaceutical Chemistry (S. Hosztafi), we could overcome this issue and did develop more promising opioid molecules. In addition, in this field we have national and international cooperative research framework, with the Biological Research Center of Szeged (S. Benyhe and F. Zádor; Hungarian Academy of Sciences), and the University of Charité in Berlin (M. Schäfer és S. Mousa).

Compounds that appear to be promising in in vitro isolated organ- and binding assays are further analyzed in in vivo in pain models above mentioned.

Publications on this topic:

  • Al-Khrasani, E. Lacko, P. Riba, K. Kiraly, M. Sobor, J. Timar, S. Mousa, M. Schafer, S. Furst, The central versus peripheral antinociceptive effects of mu-opioid receptor agonists in the new model of rat visceral pain, Brain Res Bull, 87 (2012) 238-243.10.1016/j.brainresbull.2011.10.018
  • Al-Khrasani, M. Spetea, T. Friedmann, P. Riba, K. Kiraly, H. Schmidhammer, S. Furst, DAMGO and 6beta-glycine substituted 14-O-methyloxymorphone but not morphine show peripheral, preemptive antinociception after systemic administration in a mouse visceral pain model and high intrinsic efficacy in the isolated rat vas deferens, Brain Res Bull, 74 (2007) 369-375.10.1016/j.brainresbull.2007.07.008
  • Balogh, Z.S. Zadori, B. Lazar, D. Karadi, S. Laszlo, S.A. Mousa, S. Hosztafi, F. Zador, P. Riba, M. Schafer, S. Furst, M. Al-Khrasani, The Peripheral Versus Central Antinociception of a Novel Opioid Agonist: Acute Inflammatory Pain in Rats, Neurochem Res, 43 (2018) 1250-1257.10.1007/s11064-018-2542-7
  • Furst, P. Riba, T. Friedmann, J. Timar, M. Al-Khrasani, I. Obara, W. Makuch, M. Spetea, J. Schutz, R. Przewlocki, B. Przewlocka, H. Schmidhammer, Peripheral versus central antinociceptive actions of 6-amino acid-substituted derivatives of 14-O-methyloxymorphone in acute and inflammatory pain in the rat, J Pharmacol Exp Ther, 312 (2005) 609-618.10.1124/jpet.104.075176
  • B.I. Khalefa, S.A. Mousa, M. Shaqura, E. Lacko, S. Hosztafi, P. Riba, M. Schafer, P. Ferdinandy, S. Furst, M. Al-Khrasani, Peripheral antinociceptive efficacy and potency of a novel opioid compound 14-O-MeM6SU in comparison to known peptide and non-peptide opioid agonists in a rat model of inflammatory pain, Eur J Pharmacol, 713 (2013) 54-57.10.1016/j.ejphar.2013.04.043
  • B.I. Khalefa, M. Shaqura, M. Al-Khrasani, S. Furst, S.A. Mousa, M. Schafer, Relative contributions of peripheral versus supraspinal or spinal opioid receptors to the antinociception of systemic opioids, Eur J Pain, 16 (2012) 690-705.10.1002/j.1532-2149.2011.00070.x
  • Kiraly, F.F. Caputi, A. Hanuska, E. Kato, M. Balogh, L. Koles, M. Palmisano, P. Riba, S. Hosztafi, P. Romualdi, S. Candeletti, P. Ferdinandy, S. Furst, M. Al-Khrasani, A new potent analgesic agent with reduced liability to produce morphine tolerance, Brain Res Bull, 117 (2015) 32-38.10.1016/j.brainresbull.2015.07.005
  • Lacko, P. Riba, Z. Giricz, A. Varadi, L. Cornic, M. Balogh, K. Kiraly, K. Cseko, S.A. Mousa, S. Hosztafi, M. Schafer, Z.S. Zadori, Z. Helyes, P. Ferdinandy, S. Furst, M. Al-Khrasani, New Morphine Analogs Produce Peripheral Antinociception within a Certain Dose Range of Their Systemic Administration, J Pharmacol Exp Ther, 359 (2016) 171-181.10.1124/jpet.116.233551
  • S.A. Mousa, M. Shaqura, U. Brendl, M. Al-Khrasani, S. Furst, M. Schafer, Involvement of the peripheral sensory and sympathetic nervous system in the vascular endothelial expression of ICAM-1 and the recruitment of opioid-containing immune cells to inhibit inflammatory pain, Brain Behav Immun, 24 (2010) 1310-1323.10.1016/j.bbi.2010.06.008
  • Riba, T. Friedmann, K.P. Kiraly, M. Al-Khrasani, M. Sobor, M.F. Asim, M. Spetea, H. Schmidhammer, S. Furst, Novel approach to demonstrate high efficacy of mu opioids in the rat vas deferens: a simple model of predictive value, Brain Res Bull, 81 (2010) 178-184.10.1016/j.brainresbull.2009.09.011
  • Schmidhammer, M. Spetea, P. Windisch, J. Schutz, P. Riba, M. Al-Khrasani, S. Furst, Functionalization of the carbonyl group in position 6 of morphinan-6-ones. Development of novel 6-amino and 6-guanidino substituted 14-alkoxymorphinans, Curr Pharm Des, 19 (2013) 7391-7399
  • Shaqura, B.I. Khalefa, M. Shakibaei, J. Winkler, M. Al-Khrasani, S. Furst, S.A. Mousa, M. Schafer, Reduced number, G protein coupling, and antinociceptive efficacy of spinal mu-opioid receptors in diabetic rats are reversed by nerve growth factor, J Pain, 14 (2013) 720-730.10.1016/j.jpain.2013.01.776

 

Analgesic effects of glycine transporter inhibitors

Project leader: Dr. Mohammadzadeh Amir

Glycine has a dual role in the central nervous system: acts as an inhibitory transmitter and as a positive allosteric modulator of NMDA receptors. The glycine transporters regulate the functions of glycine via uptake mechanism. There are growing data about the role of glycine and its transporter in different pain conditions. Glycine transporter inhibition might be a novel therapeutic approach to alleviate pain.

 

Assessment of the analgesic effect of the dipeptidyl peptidase 4 (DPP4) enzyme inhibitorss

Project leader: Dr. Kornél Király

DPP4 has been subjected to intensive research over the past decades. It takes part in the cleavage of many biologically active peptides, including hormones (eg. GLP-1, GIP), neuropeptides and chemokines (molecules responsible for chemotaxis). In addition, more publications have been published on its immunological role, which is independent of its enzymatic function – DPP4 on the cell surface may dimerize with adenosine deaminase, affecting its function. DPP4 enzyme inhibitors (so-called gliptins) are currently mainly known as antidiabetics, are prolonging the duration of GLP-1 and GIP, leading to an increased insulin release. The role of DPP4 enzyme in nociceptive transmission is poorly understood, however DPP4 is an important metabolizing enzyme of several endogenous peptide transmitters which are primarily involved in nociceptive transmission at the spinal cord level (e.g., SP, NPY, endomorphin 1 and 2, PACAP).  It has been reported that spinally injected DPP4 inhibitors alleviate inflammatory hyperalgesia, though they are ineffective in an acute nociceptive test. This analgesic effect is mediated by opioid receptors however, NPY also plays a role in some pain conditions.

The role of the DPP4 enzyme in transmission is carried out in permanent cooperation with the Institute of Anatomy, Histology and Embriology (Z. Puskár, M.Kozsurek).

Publications on this topic:

  • Kiraly, M. Kozsurek, E. Lukacsi, B. Barta, A. Alpar, T. Balazsa, C. Fekete, J. Szabon, Z. Helyes, K. Bolcskei, V. Tekus, Z.E. Toth, K. Pap, G. Gerber, Z. Puskar, Glial cell type-specific changes in spinal dipeptidyl peptidase 4 expression and effects of its inhibitors in inflammatory and neuropatic pain, Sci Rep, 8 (2018) 3490.10.1038/s41598-018-21799-8
  • Kiraly, A.M. Lambeir, J. Szalai, A. Szentirmay, W. Luyten, I. Barna, Z. Puskar, M. Kozsurek, A.Z. Ronai, The dipeptidyl peptidase IV (CD26, EC 3.4.14.5) inhibitor vildagliptin is a potent antihyperalgesic in rats by promoting endomorphin-2 generation in the spinal cord, Eur J Pharmacol, 650 (2011) 195-199.10.1016/j.ejphar.2010.09.060
  • Kiraly, B. Szalay, J. Szalai, I. Barna, K. Gyires, M. Verbeken, A.Z. Ronai, Intrathecally injected Ile-Pro-Ile, an inhibitor of membrane ectoenzyme dipeptidyl peptidase IV, is antihyperalgesic in rats by switching the enzyme from hydrolase to synthase functional mode to generate endomorphin 2, Eur J Pharmacol, 620 (2009) 21-26.10.1016/j.ejphar.2009.08.018
  • A.Z. Ronai, K. Kiraly, A. Szebeni, E. Szemenyei, Z. Prohaszka, Z. Darula, G. Toth, I. Till, B. Szalay, E. Kato, I. Barna, Immunoreactive endomorphin 2 is generated extracellularly in rat isolated L4,5 dorsal root ganglia by DPP-IV, Regul Pept, 157 (2009) 1-2.10.1016/j.regpep.2009.06.006

Novel approaches to alleviate neuropathic pain – Consortial grant with Meditop Ltd.

Project leader: Mahmoud Al-Khrasani

The alleviation of neuropathic pain (NP) is still a clinical challenge. The first line treatment includes gabapentinoids and antidepressants, which are commonly lack of proper analgesic effects. In this project a novel promising compound is investigated in NP pain model.