Peptides with therapeutic potential

Living With Hep B Treatment and Monitoring
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• HBV produces HBx, a protein that has many functions, one of which is floating around and making holes in the outer mitochondria membrane, which results in mitchodondrial dysfunction and the generation of ROS

• HBx specifically binds to cardiolipin: Hepatitis B virus X protein induces size-selective membrane permeabilization through interaction with cardiolipin - PubMed “In this study, we report that HBx specifically targets cardiolipin (CL) and induces membrane permeabilization depending on CL concentration in mitochondrial outer membrane-mimic artificial liposomes.”

• SS-31, a simple peptide that is only 4 amino acids long, selectively binds to cardiolipin and stabilizes it. It’s currently in use for several mitochondrial diseases, and has shown to be very successful in many areas. It appears to be completely safe, and appears to have a significantly higher affinity for cardiolipin than HBx: The mitochondria-targeted peptide SS-31 binds lipid bilayers and modulates surface electrostatics as a key component of its mechanism of action - PubMed

• Although cardiolipin comprises around 20% of the inner mitochondrial membrane, and only about 3% of the outer mitochondria membrane, given that both HBx and SS-31 bind to cardiolipin (with HBx primarily binding to the cardiolipin on the outer mitochondrial membrane and SS-31 primarily to the inner), they would be competing for the same binding sites

• Since SS-31 is safe, very small, and accumulates 1-5000 fold in mitchondria (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7247319/: “exogenously added SS peptides traverse the plasma membrane in an energy-independent and nonsaturable manner and accumulate strongly (1000–5000-fold) at the mitochondrial inner membrane”), it seems likely to disrupt HBx’s effects on mitochondria

I have my own speculations here which are very bullish on the therapeutic potential of this peptide, but I’ll spare the scientists here as I’m sure some of them will have an unpleasant enough experience reading what I’ve already said. I will add, however, that I have tried SS-31 and the results appear to be incredible. For the backstory, I’m 43 and have always had chronic HBV. I was completely fine and very fit and healthy up until 3 years ago, where I caught covid then I had a big HBV flare where my blood test results were shocking - sky high homocysteine, platelets, kidney dysfunction, high ALT, INR ratio was at 2.0 (this is extremely high), and a bunch of other things. And I felt like I was dying. After that flare I never really went back to normal, then I had another flare about 6 months later which was just as bad, my AFP went to 45 during that one among other things. I then went on antivirals but they didn’t help that much, even after 1.4 years. They made things more stable, but I still had extreme lethargy and just felt like hell in general

Anyway, I’m not saying all the above to complain, just to say that a few days ago I started taking 10-20 mg of SS-31 per day, and the effects have been absolutely stunning. After about 24 hours I was feeling so much better, and now several days in I can suddenly remember what it’s like to feel normal again. Time will tell if these effects last but in this case I have the strong feeling that they will. As I said I’ll resist speculating on the curative therapeutic potential of this peptide too much but I think it’s safe to say that a big part of HBV’s mechanism for establishing viral persistence is in its induction of ROS, which reduces the immune response and generally makes a more pathogen-friendly environment, and in particular disables the exact local immune reactions which may otherwise be able to remove the specific infected cells. Undoing this effect would be very significant, potentially allowing the proper innate and adaptive immune response to take place

Another peptide which I’m bullish on, and have been taking and which helped but not nearly as much as SS-31, is called mots-c. Mots-c increases mitochondrial biogenesis, and it up regulates mitochondria fusion, but most importantly it up regulates MAVS: Novel function of MOTS-c in mitochondrial remodelling contributes to its antiviral role during HBV infection - PubMed (I actually found a recent study that showed therapeutic potential of mots-c in the lab specifically in regard to it up regulating MAVS and that helping against HBV, but I’m unable to find that study now, so have just posted this correlation study in the interim)
In any case the thought here is that HBx down-regulates MAVS (by binding to it), and by up-regulating it with MOTS-C may allow it to be expressed enough for the infected cell to be removed. MAVS is a protein expressed on the mitchondria to say “I am infected” and then the immune system can remove it. Because it works, HBV produces stuff to disable that mechanism (specifically it’s that same HBx protein). Unlike in the case of SS-31 it being quite clear that SS-31 would be the winner in trying to bind to the same site as HBx, it’s unclear how strong the up-regulating effects of mots-c would be versus the down-regulating effects of HBx on MAVS, so this is a lot more speculative

I was torn whether I should write this up and post this. On one hand it could be inflammatory, perhaps seen as disrespectful, and perhaps irresponsible. On the other hand if I had read this 3 years ago it would have saved me 3 years of suffering and trial and error. So I’m doing it in case it helps anybody. It also seems like a therapeutic avenue worth exploring, but given the resources and time it takes to run trials, that seems a lot less likely in the short or medium term

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Hi @bob,

This is interesting. Yes, peptides can be used as drugs, but they work best for extracellular targets (ie, things in the blood or on the surface of cells) and almost always need to be injected. This is because peptide drugs do not enter cells except in very rare circumstances, and because peptides are just small chunks of proteins so they get digested like food when eaten. Good example of peptide drugs are insulin (that is a very small protein, but can be considered as a big peptide) for diabetes, and hepcludex/myrcludex for HBV (it is a preS1-derived peptide that blocks HBV and HDV entry to cells).

I’m very glad you are feeling better, but I’m skeptical that the effect is operating through the mechanism you propose. However, if its working and is safe, then keep doing it!

John.

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Hi Bob,
I was curious, so I looked up SS-31 and Mots-c on the Internet. This is not a very scientific way to learn about these peptides. My understanding is that SS-31 is not being investigated as a HBV drug. Mots-c is forbidden by the USADA. Both are not FDA approved. I do believe drugs should be developed against HBx as it has been implicated in the development of HCC in many scientific studies. Of course, the existing anti-viral HBV drugs will greatly reduce production of hbvdna, hence production of HBx.

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Fair enough, I guess it’s all speculative

How relevant would you say it is that SS-31 and HBx both bind to cardiolipin?

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As @john.tavis has mentioned, even if it does bind cardiolipin, that doesn’t say anything about orally ingested SS-31 being able to arrive intact into the mitochondria of liver cells (and with a high enough dosage to be useful). Without any clear data, it is difficult to recommend these compounds in any way.

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It’s not orally ingested. It is injected

In any case I was just trying to ask if it could potentially be of any relevance that SS-31 and HBx both bind to cardiolipin. And since around 50% of HBx proteins are found in the mitochondria it seems to be an important site

Regarding arriving to the liver cells - in therapeutic doses often 50mg a day is used. Which should be enough to saturate most of the mitochondria

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Not necessarily - I am not sure about how it is transported around the body, but if it isn’t complexed and protected, then it has the potential to be broken down in the blood. Also, peptides generally are difficult to get into cells due to their polar nature.

In my understanding of HBx biology, most of the effects on HBV control by HBx takes place in the nucleus (via DDB1/SMC5/6 interactions), not the mitochondria. HBx can localise to the mitochondria when over-expressed in different models, but in a real infection it is rarely seen outside of the nucleus. So in my interpretation, the binding of cardiolipin is not very strong evidence that it would have an effect through this mechanism.

Thomas

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HBx appears to make holes in the cardiolipin:
“In this study, we report that HBx specifically targets cardiolipin (CL) and induces membrane permeabilization depending on CL concentration in mitochondrial outer membrane–mimic artificial liposomes. Interestingly, HBx-induced membrane permeabilization was enhanced by liposomes containing phosphatidylethanolamine, which plays a crucial role in forming a negative curvature on the membrane. We also show that the 68-117 region of HBx, which interacts with mitochondria, is necessary for membrane permeabilization. We examined the size of the pores formed by HBx and found that HBx permeates fluorescent dyes depending on the hydrodynamic diameter with a pore size of approximately 10 nm.“

Source: Hepatitis B virus X protein induces size-selective membrane permeabilization through interaction with cardiolipin - ScienceDirect

From everything I’ve read at least a decent percentage of HBx is found in the mitochondria. Though I guess it could be just in the models they used, as you said

“Yoo et al. found more than 50% HBx in the cytoplasm tends to be distributed into the mitochondria by overexpressing the HBx‐Flag at 48 h after transfection. As a part of the hepatitis B virus, HBx affects various intracellular metabolic activities by regulating the signaling pathways in mitochondria.”

Regarding the potential degradation of peptides in blood - in my original post I quoted this:

Since SS-31 is safe, very small, and accumulates 1000-5000 fold in mitchondria (The mitochondria-targeted peptide SS-31 binds lipid bilayers and modulates surface electrostatics as a key component of its mechanism of action - PMC “exogenously added SS peptides traverse the plasma membrane in an energy-independent and nonsaturable manner and accumulate strongly (1000–5000-fold) at the mitochondrial inner membrane”. So it does definitely get into mitchondria. It’s been tested pretty extensively

Also, it’s excreted by the kidneys. It is particularly stable. It has a half life of about 4 hours. Besides that there’s loads of other therapeutic peptides which are just straight unprotected peptides. The ozempic weight loss glp-1 agonist blockbuster for example, is just unadulterated semaglutide. I took a look at the pending therapies by the FDA, and over 50% of them were just regular peptides, many of them found in the 60’s in Russia. It seems like we’re going to be seeing a resurgence of them as they get approved more for various uses. I guess the difference is those peptides are naturally occurring so the getting them into cells issue is already solved