Background & Aims

Painful diabetic neuropathy (PDN) is a long-term complication of diabetes affecting up to 30% of patients1. It is challenging to treat with clinically available drugs due to suboptimal efficacy and dose-limiting side effects1. Hence, there is a need for new analgesics with novel mechanisms of action. In our previous research on the pathophysiology of PDN, a long-term complication of diabetes, impaired nitric oxide (NO) availability appeared to be a contributing factor2,3. The aim herein was to assess the extent of in vitro NO release by three novel NO donor compounds (DH-MO-1, DH-MO-2 and DH-MO-6) as well as their in vivo pain-relief efficacy during the morphine-sensitive (8-11 weeks post-STZ) and morphine-hyposensitive phases (16-18 weeks post-STZ) in a widely used rat model of PDN.

Methods

Mu-opioid (MOP) receptor expressing human embryonic kidney cells were incubated with 3 novel NO donor compounds for a 2-h period relative to the positive control NO donor, SIN-1. The extent of NO (as total nitrite) released was measured using a Griess reagent assay. Blood glucose levels (BGLs), von Frey paw withdrawal thresholds (PWTs) and body weights of male Wistar rats were recorded before diabetes induction with intravenous streptozotocin (STZ) at 70 mg/kg. On day 9 post-STZ, rats with BGLs ?15 mM were confirmed as diabetic. Rats with PWTs ?8g at 8 weeks post-STZ had fully developed allodynia. These rats received single subcutaneous doses of an NO donor at 3-30 µmol/kg, gabapentin (100 mg/kg) or vehicle. A ‘washout’ protocol was used with ?2 days of washout between each dose. PWTs were measured by a tester blinded to treatment. Mean (±SEM) PWT versus time curves and the ED50 values were generated using GraphPadTM Prism (v9.3.1).

Results

The in vitro release of NO from DH-MO-2 was high at 71% of that of the positive control, SIN-1, whereas NO release from DH-MO-1 and DH-MO-6 was much lower at 6% and 5.5% respectively relative to SIN-1. In the STZ-diabetic rat model of PDN, during the morphine-sensitive phase, the ED50 (95% CI) values for DH-MO-1, DH-MO-2 and DH-MO-6 for the relief of mechanical allodynia in the bilateral hindpaws were 21.6 (11.0-47.8) µmol/kg, not attainable (NA), and 5.2 (1.7-13.5) µmol/kg respectively. During the morphine-hyposensitive phase, the corresponding ED50 (95% CI) values for DH-MO-1, DH-MO-2 and DH-MO-6 were NA, NA, and 3.5 (1.6-7.9) µmol/kg respectively. In both phases of the model, the positive control, gabapentin, evoked a similar extent of anti-allodynia and vehicle was inactive as expected.

Conclusions

The high-releasing novel NO donor, DH-MO-2, at the doses tested, did not alleviate mechanical hypersensitivity in the bilateral hindpaws in a widely used rat model of PDN during both the morphine-sensitive and morphine-hyposensitive phases, mimicking that reported previously by our laboratory for the high-releasing NO donor, SIN-1. By contrast, single bolus doses of the low-releasing novel NO donors, DH-MO-1 and DH-MO-6, evoked dose-dependent relief of mechanical hypersensitivity in the bilateral hindpaws in our rat model of PDN during the morphine-sensitive phase, similar to that reported previously by our laboratory for the low-releasing NO donor, PRG1502. During the morphine-hyposensitive phase of this rat model of PDN, DH-MO-1 was inactive similar to vehicle whereas the potency of DH-MO-6 was significantly increased, not decreased, in these animals with advanced STZ-diabetes. DH-MO-6 is worthy of further investigation as a potential novel therapeutic agent for the relief of PDN.

References

1.Sloan G, Selvarajah D, Tesfaye S. Nat Rev Endocrinol 2021;17:400-420.
2.Huang et al., Clin Exp Pharmacol Physiol 2015;42:921-929.
3.Lotfipour S, Smith MT. Clin Exp Pharmacol Physiol 2018;45:42-49.
4.Australian code for the care and use of animals for scientific purposes. NHMRC. 8th ed. 2013.

Presenting Author

Andy Kuo

Poster Authors

Andy Kuo

PhD

The University of Queensland

Lead Author

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Mohammad Zafar Imam

PhD

The University of Queensland

Lead Author

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Alexander Mueller

PhD

Lead Author

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Rui Li

PhD

Lead Author

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Dehui Kong

PhD

Lead Author

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Craig Williams

PhD

Lead Author

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Maree Smith

PhD

University of Queensland in Brisbane, Australia

Lead Author

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Topics

  • Models: Chronic Pain - Neuropathic