Honokiol and Sirt3

A brief nod to three targets of honokiol

• A 2022 publication suggested that honokiol is effective in ameliorating cognitive impairment and mitochondrial dysfunction in a rodent and cell culture models of Alzheimer’s Disease. [1] Activation of sirtuin 3, Sirt3, was the proposed mechanism.
• Honokiol and magnolol are positive allosteric modualators of the GABAA receptor meaning that it takes less gamma amino butyric acid (GABA) to cause this chloride channel to open and hyperpolarize the neuron in question. This inter-relatedness of GABA, the agonist, the GABA receptors A and B, and sirt1 was demonstrated in pancreatic beta cells. Both GABA_A and GABA_B agonists increased Sirt1 and NAD⁺ levels in a manner that was blocked by antagonists of both receptors. [3]
• Back in 2010 a methanol extract of magnolia bark was screened for compounds that could activate the retinoid X alpha transcription factor. [4] These authors were interested in compounds that could activate the transcription factor hetero dimer RXR/LXR (liver X rector).

Instead of getting lost exploring multiple targets of honokiol, let’s focus on just one.

What is Sirt3?

Dr Wenzhen Duan of John Hopkins University wrote an interesting review on sirtuins, primarily Sirt1 and Sirt3. [5]

partial list of enzymes regulated by acetylation/Sirt3 activity [5]

• long-chain acyl coenzyme A dehydrogenase (LCAD) a key enzyme that breaks down fatty acids and generates acetyl-CoA
• other β-oxidation enzymes, including the short-chain L-3-hydroxyacyl-CoA dehydrogenase and the very-long-chain acyl coenzyme A dehydrogase facilitating mitochondrial adaptation to fuel changes.
• l cyclophilin D, which leads to the dissociation of hexokinase II and mitochondria from the outer membrane of the mitochondria, decreases glucose metabolism, and stimulates oxidative phosphorylation
• isocitrate dehydrogenase 2 (IDH2) is one of the first enzyms after acetylCoA enters the TCA cycle pictured below.
• NADH dehydrogenase 1 alpha sub complex subunit 9 (NDUFA9), This is where NADH +H⁺ enters the OXPHOS electron transport chain below.
• complex I. Complex II, of the electron transport chain
• mitochondrial MnSOD a scavenger of reactive oxygen species superoxide.
• 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2), a mitochondrial enzyme that converts acetyl-CoA into ketone bodies (acetoacetate, β-hydroxybutyrate, and acetone)

Where is Sirt3 expressed?

Noteworthy is how ubiquitous the expression of Sirt3 really is.

Retracing of steps, does reveratrol really activate Sirt3? [6]

Some background on the rcsb.org structure of Sirt3 with bromo-resveratrol was speculated to be an inhibitor of both Sirt1 and Sirt3. This abstract made it uncertain if red wine resveratrol is in fact an activator of both Sirt1 and Sirt3. The 2020 Wang publication spells it out in uncertain terms that not only does resveratrol activate Sirt3, it also promotes angiotensin II induced cardiac cell hypertrophy as a cell culture model of whole organ angiotensin II induced cardiac hypertrophy. [6] The investigators found that Angiotensin II increased the expression of “bad” genes and deceased the expression of “good” genes. [6]. Resveratrol can not only activate the expression of its target Sirt3 but also activate the transcription of autophagy related genes thus promoting the removal of damaged mitgochondria via autophagy. [6] This protective effect was blocked by silencing mRNA that prevented the translation of SIRT3 mRNA into the enzyme. [6] So now that we’ve double checked on honokiol’s polyphenol cousin resveratrol, let’s take a look at this not so well known polyphenol.

Honokiol in cardiac myocytes and mice

A followup study came out of Emory University, North Western fUniversity, and University of Chicago. This group used wildtype (normal) and Sirt3 knockout mice as well as their cultured cardiac myocytes. [6] The following image is from summary Figure 10 [7] Data from the paper deemed particularly interesting are pasted into the summary figure 10 Structurally somewhat similar honokiol was used as the putative Sirt3 agonist. [7]

• Panel 9B This is a Western blot for total mitochondrial Manganese Superoxide Dismutase (MnSOD) showing total (bottom) and just the acetylated (top). The more hnokiol that was added to the myocytes, the less intense the acetylated MnSOD bands while the intensity of the total remains the same. These data point to a direct effect on deleterious signally via reactive oxygen species.
• Panel 9 The authors used a technique called fluorescence anisotropy. What is exciting about these data is that they suggest a direct interaction.
• Panel 4D is an interesting representative visual.
• Panel 9F illustrates the ability of honokiol to activate genes whose transcription is controlled by the PGC1α transcription factor. Transcription factors bind to regulatory elements upstream of the protein coding parts of the protein coding parts of genes. Transcription factors bring in RNA polymerase that transcribes the code into messenger RNA. mRNA is translated into protein by ribosomes.

Not shown are data showing improvements in mitochondrial bioenergetics and much more. The next quetion is , “Is this compound safe?”

Toxicology Studies

The parts of the magnolia, the species, and the region it was grown have an influence on the relative proportions of magnoliol and honokiol. [8] The genetic toxicology studies seemed limited according to this review. [8]

This review also covers honokiol and magnoliol modulation of the GABA A receptor and cannabinoid receptors CB1, CB2, and GPR55. The GABA A receptor binds gamma amino butyric acid.

Metabolism of honokiol

treatment
Single or repeated doses of [14C] magnolol were orally or i. p. administered to male Wistar rats	Radioactivity mainly distributed in the gastrointestinal tract and liver, but also in kidney, pancreas and lung. A similar excretion dynamic was observed after a single oral and i. p. administration. Within 12 – 24 h more than 72% of magnolol was excreted in feces and 24% in urine. Repeated oral doses resulted in the accumulation of magnolol sulfates/glucuronides but not free magnolol
Magnolol was orally administered at the dose of 20 mg/kg b. w. to male Sprague-Dawley rats	Thirty minutes after the administration the concentration of glucoronidated magnolol and free magnolol were 1.79 µg/mL and 0.16 µg/mL, respectively
Magnolol was orally administered at 5 – 100 mg/kg/b. w. to male Sprague-Dawley rats	The absorption half-life was 0.63 h, the elimination half-life 2.33 h, the time of maximum concentration 1.12 h, and the maximum concentration is 0.16 µg/mL. Oral bioavailability was 4 – 9%. The locomotor activity, measured as indicative of the pharmacodynamic profile, was affected starting from 20 mg/kg/b. w.
Magnolol was administered i. v. at 2 – 10 mg/kg b. w. to male Sprague-Dawley rats	Increasing dosages have same half-life but increasing AUC. Magnolol distributes evenly in different brain regions with concentration higher than plasma
Magnolol was administered to male Sprague-Dawley rats as a single i. v. dose 20 mg/kg b. w. or as single or multiple oral doses (50 mg/kg/b. w.)	Comparable levels of magnolol and magnolol glucuronides were found in the blood after i. v. administration whereas in orally treated rats the levels of magnolol glucuronides and sulfates were higher than that of free magnolol. The highest concentrations were found in the liver. Magnolol was found also in kidney, brain, lung, and heart
Honokiol was administered i. v. to male Sprague-Dawley rats at the dose of 5 – 10 mg/kg b. w.	A biphasic process consisting of a rapid distribution phase followed by a slower elimination phase was observed from the plasma concentration-time curves
Honokiol was orally administered to male Wistar rats at 40 mg/kg/b. w.	Honokiol was rapidly absorbed reaching its maximal plasma concentration within 20 min. It was rapidly metabolized to mono-glucuronidated honokiol and slowly eliminated (T1/2 = 290.4 min). Honokiol rapidly distributed in liver, kidney, and brain. The concentrations of honokiol and its metabolites were highest in liver, followed by kidney and brain. At central level only honokiol was detected indicating that its metabolites cannot cross the blood-brain barrier
M. officinalis cortex extract (corresponding to 12.78 mg/kg b. w. of magnolol) was administered intragastrically to male Sprague-Dawley rats	Within the first 35 min of administration, magnolol and honokiol crossed the blood brain barrier and accumulated in different brain regions
Healthy subjects and asthmatic patients were treated with 5 g/d of Saiboku-To (corresponding to 2.1 mg/d of magnolol)	Both asthmatic patients and the healthy subjects excreted the 10% of administered magnolol in the urine within 9 h. About the 95% of urinary magnolol were glucuronidated

Future outlook?

It is hard to look past the multiple targets of natural products as well as Big Pharma drugs.

References

1. Hou M, Bao W, Gao Y, Chen J, Song G. Honokiol improves cognitive impairment in APP/PS1 mice through activating mitophagy and mitochondrial unfolded protein response. Chem Biol Interact. 2022 Jan 5;351:109741. Epub 2021 Nov 6. PubMed
2. Alexeev M, Grosenbaugh DK, Mott DD, Fisher JL. The natural products magnolol and honokiol are positive allosteric modulators of both synaptic and extra-synaptic GABA(A) receptors. Neuropharmacology. 2012 Jun;62(8):2507-14. PMC free article
3. Prud’homme GJ, Glinka Y, Udovyk O, Hasilo C, Paraskevas S, Wang Q. GABA protects pancreatic beta cells against apoptosis by increasing SIRT1 expression and activity. Biochem Biophys Res Commun. 2014 Sep 26;452(3):649-54. Sci-Hub free
4. Kotani H, Tanabe H, Mizukami H, Makishima M, Inoue M. Identification of a naturally occurring rexinoid, honokiol, that activates the retinoid X receptor. J Nat Prod. 2010 Aug 27;73(8):1332-6 Sci-Hub free article
5. Duan W. Sirtuins: from metabolic regulation to brain aging. Front Aging Neurosci. 2013 Jul 23;5:36. PMC free article
6. Wang HN, Li JL, Xu T, Yao HQ, Chen GH, Hu J. Effects of Sirt3‑autophagy and resveratrol activation on myocardial hypertrophy and energy metabolism. Mol Med Rep. 2020 Aug;22(2):1342-1350 PMC free article
7. Pillai VB, Samant S, Sundaresan NR, Raghuraman H, Kim G, Bonner MY, Arbiser JL, Walker DI, Jones DP, Gius D, Gupta MP. Honokiol blocks and reverses cardiac hypertrophy in mice by activating mitochondrial Sirt3. Nat Commun. 2015 Apr PMC free article
8. Sarrica A, Kirika N, Romeo M, Salmona M, Diomede L. Safety and Toxicology of Magnolol and Honokiol. Planta Med. 2018 Nov;84(16):1151-1164. PMC free article