Updated Research and Knowledge - Cutting Edge

[jjo]

ketoconazole cream 2% twice a day + finasteride + minox will cover the Androgen side if you've got good density or less aggressive AGA.

The problem with 5ar inhibitors, especially oral ones, is that they have a tendency to increase LH production and subsequently raise blood T levels and a local reduction in T's conversion to DHT leading to even more more T within cells. Suraphysical levels of T can activate the AR just as potently as DHT so it just offsets the potential gain from DHT inhibition. It's a tricky decision, yes Dut will kill more DHT, but you'll have even more T in comparison to Fin. I would stick to Fin.

Hi Chemical, thanks so much for all your time and effort. Can you go more into what you are saying about oral 5ar inhibitors please?

When fin stopped working for me after years I switched to dut and got a couple years more. Now i'm loosing ground on 1 mg / day dut and have even added 8% ru daily.

This has been really hard on me and I don't know what to do anymore... these drugs worked super good for me and now they don't.. what can I do Chemical? Please any advice would be so much appreciated.

thanks

[Chemical]

i got the epilium ones 500mg tablets. so can i just crush these up and put them into the solution, or do i need to filter the coating after.

I couldn't find any images of the EC500 and was hoping to get them myself. If theres a coloured coating you might be able to dissolve it until you see the actual powder idk. You could try scraping it off too. I get mine from 4nrx (dirt cheap) and the orange coating just chips off with a little effort. If you cant get it off then it shouldn't cause too much of an issue, probably just sugar and colouring.

30 mg/mL (a.k.a. 3%) VPA or SV probably isn't going to be strong enough, considering that even at it's best, VPA/SV isn't a super robust hair grower. There was a mouse study that I think the human study used as a preliminary research. In mice, they tried several different concentrations of SV. 3% was not very effective as compared to around 7%.

You're right I dont know why I keep thinking 1mg/ml = 1%. The study used 80mg/ml to get 8%. Also can you post the mouse study, I prefer not to go assumptions and random numbers. As for the dose, I've come to realise it's a bit useless working with the solubility if you can control how much of the actual drug you apply. If you went with ethanol and saturated completely to get 30mg/ml, you'd want 2ml to get 60mg and so forth. The human study advised subjects to use around 0.8ml/64mg:

The subjects were instructed to use either the VPA (sodium valproate, 8.3%) or placebo spray twice daily, with seven or eight pumps (~0.8 mL) for each dose.

I've been making different formulations, and one batch I made had too much water & glycerin, and not enough ethanol & butylene glycol. The VPA completely separated. Then I remade this formula, but tweaked it to have less water/glycerin and more ethanol/BG, and the VPA was totally soluble. I took notes on all the amounts used, but I'm going from memory at the moment. If you look at the molecule though, it's obvious it can't dissolve in pure water.

Aha, here it is: a pubchem (good site to look up molecules) reference for VPA (it didn't have much to say about SV solubility). VPA in pure water will only go to about 1 mg/mL, or 0.1%. Although, you can't always trust these solubility numbers, sometimes it's less or more. But that sounds about right to me, because it's a mostly fat-like molecule.

https://pubchem.ncbi.nlm.nih.gov/compound/3121#section=Solubility

You might have missed this comment I made along with the source for the solubility numbers

Since Sodium Valproate is the more readily available form and has excellent water solubility at 50mg/ml, in comparison to 1.3mg/ml for Valproic Acid (source) it looks like Sodium Valproate is a strong candidate to be used in our protocols.

Also pubchem didnt have anything on sodium valproate and the human study used sodium valproate. I dont think its fair to compare VPA and SV hence I looked deeper

http://www.scbt.com/datasheet-202378.html

I trust scbt if pubchem doesnt list the exact molecule and both the pdf and scbt show the Sodium Valproate has 50mg/ml solubility in water. Whereas Valproic Acid specifically has very low water solubility corroborated by multiple sources and the trustworthy pubchem. You've also come to realise this with your own experiments that Valproic Acid isn't very poorly soluble in water. But the real question is did you try Sodium Valproate? I'm sure you acknowledge that the two are chemically different.

Ethanol isn't hard to get. You don't even need 95% Everclear. Most states that make it tough to get the 95% will offer 75.5% ethanol at the liquor store. And from 75.5%, you can easily add water to get down to ~30% ethanol.

Good point but water is still an option. Not everyone is in the states remember!

I wouldn't recommend PG as a solvent, but rather BG instead. Butylene glycol has similar solvent properties to PG, but BG is typically a lot less irritating. For those buying all kinds of ingredients to use, it would be a shame to go cheap and have skin irritation with PG. I have to avoid PG or my scalp will break out, and a surprising number of people have PG sensitivity. I got BG from lotioncrafters online.

Another good solvent that is under-utilized by the hair loss community is glycerin. Glycerin also looks chemically a lot like PG, but that one extra -OH group makes a big difference. Rather than being an irritant, glycerin is a nutrient that moisturizes the skin, adds some viscosity to the formula (think of hand sanitizers), and the glycerin is actually a 3-carbon sugar that can be used as fuel by the skin cells, or other aspects of cell metabolism. Glycerin is good for skin at up to 20% in a formula.

The PG is mainly to prevent the ethanol from drying up too quick and give the molecules enough time to soak in after the barrier is weakened. As you can tell I dont know jack about chemistry so couldn't recommend anything other than PG even though many people are sensitive to it. Looks you are right about BG and glycerols being equally effective substitutes to PG - according to these guys. Glad we've finally got someone with a chemistry background to correct my mistakes!

[bej]

Correct, both VPA and SV are soluble in most solvents and I highly doubt you've want to go beyond 30mg/ml anyway. The only reason I suggested water is because its easier for people to make a solution without too much effort. I have a strong suspicion the study used ethanol to enhance the skin penetration seeing as ethanol strips the lipid barrier and weakens the stratum corneum. I personally encourage the use of ethanol + PG as a primary vehicle for nearly everything. Ideally a you'd want a water based solution with any amount of ethanol (maybe PG too) to enhance skin permeation but I dont see why a water based solution wouldn't work in this case for people who cant get hold of 190 proof alcohol.

Regarding separation? - I've never seen a dissolved molecule separate from a solution before its reached saturation. But I'm not a molecular biologist and only have experience mixing water and sugar so correct me if I'm wrong lol.

30 mg/mL (a.k.a. 3%) VPA or SV probably isn't going to be strong enough, considering that even at it's best, VPA/SV isn't a super robust hair grower. There was a mouse study that I think the human study used as a preliminary research. In mice, they tried several different concentrations of SV. 3% was not very effective as compared to around 7%.

I believe the SV studies had to use ethanol simply for solubility reasons. Take a look at the molecule, it is, for the most part, a very fatty-like molecule. I've been making different formulations, and one batch I made had too much water & glycerin, and not enough ethanol & butylene glycol. The VPA completely separated. Then I remade this formula, but tweaked it to have less water/glycerin and more ethanol/BG, and the VPA was totally soluble. I took notes on all the amounts used, but I'm going from memory at the moment. If you look at the molecule though, it's obvious it can't dissolve in pure water.

Aha, here it is: a pubchem (good site to look up molecules) reference for VPA (it didn't have much to say about SV solubility). VPA in pure water will only go to about 1 mg/mL, or 0.1%. Although, you can't always trust these solubility numbers, sometimes it's less or more. But that sounds about right to me, because it's a mostly fat-like molecule.

https://pubchem.ncbi.nlm.nih.gov/compound/3121#section=Solubility

Ethanol isn't hard to get. You don't even need 95% Everclear. Most states that make it tough to get the 95% will offer 75.5% ethanol at the liquor store. And from 75.5%, you can easily add water to get down to ~30% ethanol.

I wouldn't recommend PG as a solvent, but rather BG instead. Butylene glycol has similar solvent properties to PG, but BG is typically a lot less irritating. For those buying all kinds of ingredients to use, it would be a shame to go cheap and have skin irritation with PG. I have to avoid PG or my scalp will break out, and a surprising number of people have PG sensitivity. I got BG from lotioncrafters online.

Another good solvent that is under-utilized by the hair loss community is glycerin. Glycerin also looks chemically a lot like PG, but that one extra -OH group makes a big difference. Rather than being an irritant, glycerin is a nutrient that moisturizes the skin, adds some viscosity to the formula (think of hand sanitizers), and the glycerin is actually a 3-carbon sugar that can be used as fuel by the skin cells, or other aspects of cell metabolism. Glycerin is good for skin at up to 20% in a formula.

[joshuk]

i got the epilium ones 500mg tablets. so can i just crush these up and put them into the solution, or do i need to filter the coating after.

[Chemical]

@Chemical

Putting aside the negative effects and looking purely at the effects on hair, would Propecia and Nizoral 1% plus Minoxidil be enough to cover the Androgen angle to this or should I just use dut?

ketoconazole cream 2% twice a day + finasteride + minox will cover the Androgen side if you've got good density or less aggressive AGA.

The problem with 5ar inhibitors, especially oral ones, is that they have a tendency to increase LH production and subsequently raise blood T levels and a local reduction in T's conversion to DHT leading to even more more T within cells. Suraphysical levels of T can activate the AR just as potently as DHT so it just offsets the potential gain from DHT inhibition. It's a tricky decision, yes Dut will kill more DHT, but you'll have even more T in comparison to Fin. I would stick to Fin.

I have just ordered VPA from inhouse, if i crush a 500mg tablet into 60ml of solution this gives me 8.3mg per ML. can i use a 70/30 eth/pg mixture or is it better to use 60ml of distilled water instead??? want to try this as i cannot use minoxdil.

Did you get the depakote ones or the epilium? It looks like the tablets have some coating, I want tablets without a coating, preferably capsules.

I think I'll scrap the whole mg/ml crap, so long as we know what doses work and there is okayish solubility we can just control the dose with the application. Go with 50/30/20 Ethanol/Water/PG and use as much mg as you want. You dont want the ethanol to dry up quickly. Given that the study used 8mg per application and some individuals probably used less or more, with some cases of slight systemic absorption I think it's pretty safe to bump the dose higher than 8mg. The plasma therapeutic range is 50-125 µg/mL according to google and the topical absorption didnt get anywhere near those numbers. The cases of hairloss associated with oral VA require chronic high doses and even then it only affects a small minority:

http://www.bioline.org.br/pdf?pt05030

Among 211 patients who received sodium valproate (enteric coated tablet), 51% were male and 49% were female (no significant difference). Patients’ mean age of epilepsy onset was 11 years old and mean age starting to receive sodium valproate was 18 years old. More than 78% of patients had experienced tonic–clonic seizures (primary or secondary). All patients received monotherapy (single drug treatment). Overall, there were 6 cases (3.5%) of hair loss and curling of hair. Among them 3 cases were female and 3 were male. This side effect could be observed 3 months after first dose of therapy It took 1-2 years before appearance of hair loss and curly hair in other cases.

I believe the AR inhibitory properties are due to it's ER potentiating effects and I'm seeing more studies showing various ER agonists have the ability to downregulate AR in a dose dependent manner. The hairloss effects of VA could be more noticeable in women than men since female follicles require certain levels of ER to grow and maintain anagen. The study I posted before showed the AR suppressive effects were maximal at 2mM+ so I'm going to try and figure how that translates to topical use. I'm tempted to try alfatradiol (17-alpha Estradiol - available from Amazon.de as Ell Cranell) which supposedly increases the levels of Aromatase - more E2 should help VA work at a lower concentration. 17alpha Estradiol doesnt have the feminizing effects of 17beta Estradiol so you dont need to worry about gyno as long as the dose is low.

Chemical, why did you stop using oleuropein?

The powder dried up in the capsules and I need to buy more. I'm experimenting without it for now to see if I can get regrowth using other stuff.

[FeelsBad]

Chemical, why did you stop using oleuropein?

[joshuk]

I have just ordered VPA from inhouse, if i crush a 500mg tablet into 60ml of solution this gives me 8.3mg per ML. can i use a 70/30 eth/pg mixture or is it better to use 60ml of distilled water instead??? want to try this as i cannot use minoxdil.

[BaldingEagle]

@Chemical

Putting aside the negative effects and looking purely at the effects on hair, would Propecia and Nizoral 1% plus Minoxidil be enough to cover the Androgen angle to this or should I just use dut?

[Chemical]

Hmm, so you think using bicalutamide could be good ?

If you're okay with castration then yes. It will go systemic even if you use it as topical. You want a special AR blocker that doesnt function in the blood stream. Like CB or RU, or a combination of the experimental ones in this thread.

Chemical,

I've followed this thread with interest and appreciate all your hard work and research. I'd like to replicate your treatment on myself, but I don't have a chemistry background and want to make sure I'm using the right products and dosages. Is it possible to go into the details of your current regimen so that the followers of this thread can easily replicate what you're doing? If you can provide exact dosages and even brand names you've been using to buy some of these products, that would be really helpful. Thanks again!

Here's a good place to start:

https://www.baldtruthtalk.com/threads/22991-Treatments-and-Science-Read-Only

My protocol is always changing so you're better off sticking to the main components:

Ketoconazole cream 2% - Daktarin - 2x day
Kirkland minox 5% - 2x day
Borage oil - Solgar (capsules poured into dispenser) - night
Evening primrose oil - Natures Aid capsules (same dispenser) - night
Oleuropein - Swanson (1mg/ml in minox) - night (stopped for now)
Sodium Valproate tablets crushed - 10mg/ml in distilled water and equal amount of minox solution (stopped for now)
Teavigo EGCG - Swanson - 8-10/mg/ml in separate minox solution - night (stopped)

chemical , I have no knowledge of alopecia. I suffer aga for 16 years . I was deceived by a surgeon in Spain twice and had to go to Dr. Hasson to solve the problem .
I took finasteride for 7 years with very good results but I gave it up because of side effects . 8 years ago I left finasteride.
currently I do not take drugs for hair loss and I 'm sentenced because I have several capillaries operations.
I read your posts but I do not understand anything . I'm depressed , bitter. please chemical , you seem to have much knowledge. I think we are many users who believe in you. Do you think you come to any conclusion on your research and get a possible future treatment ?

Hey machi, I want to tell you that I know what it feels like but I havent had it for that long so I can only imagine how hard it is for guys like you. Growing hair is a difficult process and it takes alot time, for some people nothing seems to work and it can be frustrating so I'd encourage you try and stay hopeful. We're in the 21st century and there are alot of promising treatments on the horizon. I'm not going to give you false hope by saying the research we're doing here is going to lead to a cure but we've made some huge leaps in understanding AGA better which will increase the chances of coming across something that helps. You can try the treatments listed in this thread but it will take time and perseverance to see results especially given that you've had AGA for a while. Stay strong.

Chemical, have you seen:

http://www.dannyroddy.com/weblog/antihistaminebaldnessstress

which if correct indicates that Estrogen plays a negative role (causes terminal hair - vellus, is involved in fibrosis of scalp around follicles, etc) rather than a positive role?

I like the authors scientific approach but he seems to use alot of associative reasoning. It pains me to see scientific minds become a victim of confirmational bias, they end up looking for literature that reaffirms their hypothesis and form conclusions based on loose inductive logic.

The heavy focus on androgens and “the genes” as causes of baldness have led people to believe that pattern hair loss is a compartmentalized problem rooted in vanity that has nothing to do with their metabolism or lifestyle. Confusion about the role of androgens probably relates to testosterone’s conversion into estrogen during metabolic stress and that dihydrotestosterone (DHT), like DHEA, can increase to buffer the effects of metabolic stress, for example, as an anti-estrogen.

He believes AGA is more a lifestyle/endocrinology related pathogenesis. Oxidative stress, Immune system, prostaglandins, glucose metabolism, diet, exercise - yeah they all play a part just like breathing provides oxygen to all mitochondria in all the cells. Side effects and symptoms of AGA do not mean that they are causative factors. Alot of people are in the PGD2 bandwagon and with good reason, but InBeforeTheCure has demonstrated that this whole PGD2 thing is caused by Androgens. How are you going to deny that Androgens arent the main factor driving the progression of baldness?

Estrogen, free fatty acids, prostaglandins, mast cells, and histamine are involved in the development of fibrosis, or the abnormal progression of the normal formation of fibrous material between cells due to inflammation. In 1992, Jaworsky et al. found that a prominent feature of baldness was mast cell degranulation and the activation of fibroblasts resulting in fibrotic thickening of the hair follicle.[28] In another experiment, 412 people with pattern baldness (193 men and 219 women) confirmed the presence of a significant degree of perifollicular fibrosis in at least 37% of cases. Moreover, balding men with higher levels of inflammation and fibrosis led to worse outcomes using the traditional hair loss remedy minoxidil compared to those with lower levels.[29]

There is fibrosis in AGA. Estrogens, prostaglandins and histamine are involved in the development of fibrosis. Therefore, Estrogen is bad. Also AGA has three letters. A triangle has three sides. The illuminati created AGA.

I've had this debate about fibrosis and AGA in the past, feel free to read the exchange with youngin from this post onwards. In short, the Reactive oxygen species generated by the DPC as a result of Androgen overexpression leads to excessive TGF-beta production. TGF-beta has been shown to kick off the chain that causes fibroblasts to deposit fibrotic tissue.

Estrogens are actually beneficial to frontal hair follicles, being able to stimulate the elongation of hair shafts - I made an in-depth post here here.

Both sodium valproate and VPA are very soluble in vehicles with 30% (or more) ethanol. The human AGA study I think used 27% ethanol with sodium valproate. To have the concentration you want (around 7% or 8%) you probably need the ethanol there or it might separate. I've made a lot of mixtures with VPA and various other components, and the ethanol seems necessary to have useful concentrations.

Correct, both VPA and SV are soluble in most solvents and I highly doubt you've want to go beyond 30mg/ml anyway. The only reason I suggested water is because its easier for people to make a solution without too much effort. I have a strong suspicion the study used ethanol to enhance the skin penetration seeing as ethanol strips the lipid barrier and weakens the stratum corneum. I personally encourage the use of ethanol + PG as a primary vehicle for nearly everything. Ideally a you'd want a water based solution with any amount of ethanol (maybe PG too) to enhance skin permeation but I dont see why a water based solution wouldn't work in this case for people who cant get hold of 190 proof alcohol.

Regarding separation? - I've never seen a dissolved molecule separate from a solution before its reached saturation. But I'm not a molecular biologist and only have experience mixing water and sugar so correct me if I'm wrong lol.

[Seuxin]

Bej,

I have a very very importante question ( It's Seuxin...form HairLossHelp too ).

About VPA, what is important....the amount of VPA per day, or the % ?

By example :

If i use VPA at 7% in stemoxydine, and if i use 6ml per day of stemox...it represent more VPA ( as mg) than if i use 1-2ml of solution, right ?

For RU, CB, or Minox for example the important is the amount of solution...but i don't know if for acidic solution it's the same.

Do you understand my question ? What do you think ?

Thanks

[bej]

Both sodium valproate and VPA are very soluble in vehicles with 30% (or more) ethanol. The human AGA study I think used 27% ethanol with sodium valproate. To have the concentration you want (around 7% or 8%) you probably need the ethanol there or it might separate. I've made a lot of mixtures with VPA and various other components, and the ethanol seems necessary to have useful concentrations.

[Rog]

Chemical, have you seen:

http://www.dannyroddy.com/weblog/antihistaminebaldnessstress

which if correct indicates that Estrogen plays a negative role (causes terminal hair - vellus, is involved in fibrosis of scalp around follicles, etc) rather than a positive role?

[Chemical]

For those of you that havent been following the science, our current understanding is that for as long as Testosterone and DHT is around, any treatment that tries to increase beta-catenin will be significantly blunted. The AR will just suck up the beta-catenin and prevent it from working. So we need to reduce AR and 5ar using suppressors and increase aromatase within hair follicles to further divert Testosterone to convert to Estrogen which can enhance the growth of frontal hair follicles. Also since there will be some DHT/T floating around you will need to boost beta-catenin so that even after AR has bound beta-catenin until saturation, there will still be enough left to activate the good TCF/LEF genes that induce and maintain anagen.

Existing Androgen Receptor inhibitors

Ketoconazole

Ketoconazole binds to the human androgen receptor.

Ketoconazole competition with [3H]methyltrienolone (R1881) for androgen binding sites in dispersed, intact cultured human skin fibroblasts was determined at 22 degrees C. Fifty percent displacement of [3H]R1881 binding to AR was achieved by 6.4 +/- 1.8 (SE) x 10(-5) M ketoconazole. Additional binding studies performed with ketoconazole in the presence of increasing amounts of [3H]R1881 showed that the interaction of ketoconazole with AR was competitive when the data were analyzed by the Scatchard method. It should be noted, however, that the dose of ketoconazole required for 50% occupancy of the androgen receptor is not likely to be achieved in vivo, at least in plasma. Finally, androgen binding studies performed with other imidazoles, such as clotrimazole, miconazole, and fluconozole, revealed that in this class of compounds only ketoconazole appears to interact with the androgen receptor. Ketoconazole appears to be the first example of a non-steroidal compound which binds competitively to both SSBG and multiple steroid hormone receptors, suggesting that the ligand binding sites of these proteins share some features in common.

Ketoconazole as an adjunct to finasteride in the treatment of androgenetic alopecia in men.

Dihydrotestosterone (DHT) binding to androgen receptors (AR) in hair follicles is commonly accepted as the first step leading to the miniaturizing of follicles associated with androgenetic alopecia (AGA). Testosterone is converted to DHT by the enzyme 5alpha-reductase. Finasateride a 5alpha-reducase inhibitor blocks the production of DHT and is currently used to treat AGA. The inhibition is not complete but a reduction of DHT systemically and in the scalp is accomplished. Ketoconazole has been clinically shown to be effective in the treatment of AGA. In this paper, evidence is presented to support the hypothesis that ketoconazole 2% shampoo has a local disruption of the DHT pathway. It is proposed that using ketoconazole 2% shampoo as an adjunct to finasteride treatment could lead to a more complete inhibition of DHT and thus better treat AGA.

Ketoconazole has a plasma half life of 8 hours and I'm not too sure if that relates to skin retention too. The current recommended dosing is 3x a week using the shampoo but I personally encourage the use of the cream 2-3x a day. Its easier to apply, more convenient and you dont need to worry about the shampoo causing irritation from excessive use. Even though keto is a mild AR blocker people still see results using it so its not entirely useless - remember that this is using it 3x a week so imagine if you used it 2-3x a day.

Minoxidil

Minoxidil may suppress androgen receptor-related functions

Minoxidil concentrations were chosen based on most common reports in the literature: 0.1 to 10 μM for oral intake and about 1 mM for topical application of 5% minoxidil (100 mM) in skin tissue (Regaine 5% minoxidil topical solution; monograph, February 2013), assuming 1.7% absorption. In in vitro organ culture or animal studies, high concentrations, ranging from 1-100 mM, have been reported in skin tissue [31, 32]. After treating with different concentrations of minoxidil (1 to 100 μM), cells were harvested and cellular extracts were assayed for luciferase activity. As shown in Fig. Fig.1A,1A, minoxidil suppressed AR reporter activity at the concentrations tested.

To further test minoxidil suppression of AR-related function, we tested the effects of minoxidil on the growth of LNCaP cells, an androgen-sensitive prostate cancer cell line that, as noted above, endogenously expresses AR. As shown in Fig. Fig.1D,1D, minoxidil suppressed DHT-dependent LNCaP cell growth in concentration-dependent manner. Notably, at the highest concentration, cell growth was suppressed to a level comparable to that observed in the absence of DHT. These data provide the first demonstration that minoxidil can suppress AR-related functions, including AR transcription and AR-related cell growth.

To assess possible nonspecific effects of minoxidil at high concentrations, we performed control studies, testing different concentrations of minoxidil (1-100 μM) on glucocorticoid receptor (GR) transcriptional activity. As shown in Fig. Fig.1E,1E, the highest concentration minoxidil (100 μM) did affect GR transcriptional activity in reporter assays in PC-3 cells, whereas lower concentrations (1-10 μM) did not. These data suggest that minoxidil, a small hydrophobic molecule, may have multiple targets in the cell when used at high concentrations.

You can see that minoxidil can suppress AR activity in a dose dependent and time dependent manner providing more evidence that minoxidil works on two levels necessary to treat AGA. This explains why alot of people can achieve regrowth solely using minox seen both in studies and anecdotal reports.

Its important to note that minoxidil does not work if the follicles do not contain adequate sulfotransferase enzymes necessary to convert minoxidil to minoxidil sulphate which is reported to be 14 times more potent.

Minoxidil sulfate is the active metabolite that stimulates hair follicles.

An important step in understanding minoxidil's mechanism of action on hair follicles was to determine the drug's active form. We used organ-cultured vibrissa follicles to test whether it is minoxidil or its sulfated metabolite, minoxidil sulfate, that stimulates hair growth. Follicles from neonatal mice were cultured with or without drugs and effects were assessed by measuring incorporation of radiolabeled cysteine in hair shafts of the treated follicles. Assays of minoxidil sulfotransferase activity indicated that vibrissae follicles metabolize minoxidil to minoxidil sulfate. Dose-response studies showed that minoxidil sulfate is 14 times more potent than minoxidil in stimulating cysteine incorporation in cultured follicles. Three drugs that block production of intrafollicular minoxidil sulfate were tested for their effects on drug-induced hair growth. Diethylcarbamazine proved to be a noncompetitive inhibitor of sulfotransferase and prevented hair growth stimulation by minoxidil but not by minoxidil sulfate. Inhibiting the formation of intracellular PAPS with chlorate also blocked the action of minoxidil but not of minoxidil sulfate. Acetaminophen, a potent sulfate scavenger blocked cysteine incorporation by minoxidil. It also blocked follicular stimulation by minoxidil sulfate apparently by directly removing the sulfate from the drug. Experiments with U-51,607, a potent minoxidil analog that also forms a sulfated metabolite, showed that its activity was inhibited by both chlorate and diethylcarbamazine. These studies show that sulfation is a critical step for hair-growth effects of minoxidil and that it is the sulfated metabolite that directly affects hair follicles.

Sulfotransferase levels in scalp vary alot between individuals and that can account for the lack of response in some individuals

Biochemical evidence for minoxidil sulphation by two phenol sulphotransferases has been found in human scalp skin[22] and Dooley[21] reported finding mRNA expression for four sulphotransferases in human epidermal keratinocytes. There are interindividual variations in scalp sulphotransferase activity and this correlates with the level in platelets.[22] In a clinical setting, scalp sulphotransferase activity was higher in men who responded to minoxidil compared with those who did not respond.[23]

http://www.medscape.com/viewarticle/470297_3

I'm currently looking into ways of boosting scalp sulfotransferase levels to improve the efficacy of minoxidil.

Valproic Acid - Sodium Valproate

I was quite surprised when I found out VA was a serious Androgen Receptor suppressor. Before we look at that I'd like to show you the study they did on men with AGA with a 8.3% Sodium Valproate spray in a 27% EtOH solution over 6 months, twice a day applications of 1ml.

Topical valproic acid increases the hair count in male patients with androgenetic alopecia: a randomized, comparative, clinical feasibility study using phototrichogram analysis.

Full Study

The representative clinical photographs and macrophotographs are presented at baseline and after 24 weeks of treatment (Fig. 2). The total hair count increased after 24 weeks of treatment in the VPA group; the median hair counts were 181/cm2 (range, 125–241) at baseline and 192/cm2 (range, 153–271) at 24 weeks. However, the total hair count did not change in the placebo group; the median hair counts were 194/cm2 (range, 155–244) at baseline and 197/cm2 (range, 132–253) at 24 weeks (Fig. 3a). The median change in total hair count from the baseline was 23/cm2 (range, 17 to 39) in the VPA group and 1/cm2 (range, 68 to 70) in the placebo group, and the difference between groups was statistically significant (P = 0.047; Fig. 3b).

In the full study you can see a photo of the results, and I can definitely see an increase in hair count both in the closeup and wide angle. You have to understand that there is a very high probability the subjects might not have been too diligent with the treatment and the ages ranged from 27-45. The AGA has probably become alot worse and its very unlikely that 6 months of monotherapy will completely regrow hair. For those of you that expect complete regrowth from one treatment please reevalute your expectations, we do not live in the world of movies and fiction where scientists come up single drugs that cure diseases completely. A small but consistent positive effect of a treatment on a reasonable sample size is a good predictor that a treatment has potential to have additive or even synergistic effect when combined with other treatments. Most of the placebo individuals saw a decline in hair count but a vast majority of VA subjects saw no reduction in hair count or an increase - which means it definitely did something.

The serum levels achieved by topical VA dont seem too bad but its something to keep in mind

The serum VPA concentration was measured in 20 subjects in the VPA group and 19 subjects in the placebo group. The serum VPA level was detectable in only seven subjects in the VPA group (range, 0.4–2.3 lg/mL), whereas it was too low (<0.4 lg/mL) to be detected in the other 13 subjects in the VPA group and all of the subjects in the placebo group.

Valproic acid and its derivatives enhanced estrogenic activity but not androgenic activity in a structure dependent manner.

Full Study

Steroid hormones affect metabolic pathways and cellular functions. Valproic acid (VPA), used as antiepileptic drug, inhibits histone deacetylases and interacts with intracellular receptors. We analyzed the impact of VPA and VPA derivatives on activation of estrogen and androgen receptors (ER and AR) using reporter gene assays. VPA and its long-chain derivatives (long name 1), (long name 2) and (long name 3) enhanced 17β-estradiol-induced ERα and ERβ activation partly synergistically with a structure-activity correlation. The extent of this effect regarding to ERα activation increased with prolongation of the aliphatic side chain. Regarding AR activation, VPA, S-pentyl-4-yn- and heptyl-4-yn-VPA slightly induced AR activity when tested alone. In combination with the AR agonist 5α-dihydrotestosterone, VPA, S-pentyl-4-yn- and heptyl-4-yn-VPA showed anti-androgenic effects without an apparent structural relation. Our results indicate that VPA and its derivatives affect estrogen signaling with a structural specificity, while the (anti-)androgenic effects of these compounds are not structurally correlated.

So here we have our first indication that VPA can indeed suppress AR. No mention of how much so lets delve into the study

For VPA, concentrations between 0.001 and 3 mM and for its derivatives between 0.01 and 1 mM were investigated. The therapeutic range of VPA comprises 0.35–0.7 mM in serum [42], whereas concentrations of 3 mM can be attained in the serum of patients after an acute VPA ingestion [43].

In combination with 10−9 M E2, VPA enhanced the E2-induced ER activation synergistically in a concentration-dependent manner with a first significant effect at 0.001 mM. For instance, 2 mM VPA caused a 3.5-fold increase of ER activity compared to 10−9 M E2 alone (E2: 100% vs VPA plus E2: 348% of maximum E2- activation; Fig. 4A). Similarly for ER activation, the co-incubation of VPA (0.001–3 mM) with 10−9 M E2 caused synergistic effects (e.g. E2: 100% vs. 1 mM VPA plus E2: 408% of maximum E2-activation; Fig. 4B). Thus, VPA enhanced the E2-induced activation of both ER types in a dose-dependent manner with a slightly greater effect on ER activation.

VPA is quite powerful at potentiating the ER pathways which is exactly what we want for frontal DPC.

Co-administration of VPA (0.01–4 mM) with 10−9 M DHT reduced AR activity in a dose-dependent manner with a first significant effect at 1 mM (Fig. 7). Butyl-4-yn-VPA did not alter DHT-induced AR activation. S-pentyl-4-yn- and heptyl-4-yn-VPA (≥0.2 mM) also decreased DHT-mediated AR activation. At the highest concentration, VPA and heptyl-4-yn-VPA inhibited the AR activity induced by DHT to control values of untreated cells (Fig. 7).

Just look at what 1mM could do to DHT's ability to bind to AR. Just look at it! And look at what 2mM could do, its identical to frickin flutamide.

The anti-androgenic effects of VPA are also characterized through a reduced AR expression and cell growth inhibition revealed in cell cultures of prostate cancer cells [64,65]. For example, Iacopino et al. [66] have shown that 0.45 mM VPA suppressed DHT-stimulated proliferation of human androgen-sensitive LNCaP prostate cancer cells. Epigenetic regulations byVPA might be involved in the modulation ofAR activity. HDAC inhibition leads to AR acetylation [67] causing reduced cell growth, apoptosis induction and differentiation [64,68]. However in our study, VPA seems to have both anti-androgenic and marginal androgenic effects dependent on the VPA concentration and the presence or absence of DHT. A structural dependency of the VPA derivatives could not be attributed to the AR activity in the presence of DHT.

That was Valproic Acid, what about Sodium Valproate?

Valproate is an anti-androgen and anti-progestin.

We examined the widely used anti-convulsants valproate (VPA) and carbamazepine (CBZ) for steroidal bioactivity using a yeast-based steroid receptor-beta-galactosidase reporter assay for the androgen receptor (AR), progesterone receptor (PR) or estrogen receptor (ER). Bioassays were performed (a) to detect agonist activity by exposing yeast to 100 microM CBZ or VPA or (b) to detect antagonist activity by exposing yeast stimulated with testosterone (5 x 10(-9) M, AR), progesterone (1.6 x 10(-9) M, PR) or estradiol (2.6 x 10(-11) M, ER) together with either VPA or CBZ for 4 (PR) or 16 (AR, ER) hours. VPA showed dose-dependent (1-800 microM) inhibition of progesterone-induced PR- and testosterone-induced AR activity but had no ER antagonist bioactivity and no significant PR, AR or ER agonist bioactivity. VPA also showed a dose-dependent (1-200 microM) blockade of DHT's suppression of AR-mediated NF-kappaB activation in human mammalian cells. By contrast, CBZ had no significant PR, AR or ER agonist or AR and ER antagonist bioactivity but at the highest concentration tested (800 microM) it did antagonize PR activity. We conclude that VPA is a non-steroidal antagonist for human AR and PR but not ER. VPA's androgen and progesterone antagonism at concentrations within therapeutic blood levels (350-700 microM) seems likely to contribute to the frequency of reproductive endocrine disturbances among patients treated with VPA.

Since Sodium Valproate is the more readily available form and has excellent water solubility at 50mg/ml, in comparison to 1.3mg/ml for Valproic Acid (source) it looks like Sodium Valproate is a strong candidate to be used in our protocols. I use distilled water to dissolve the tablet after crushing them and removing the orange coating but I'm looking for capsules which should make my life easier. I would suggest starting at 5mg/ml then increasing until you get irritation or notice side effects. You can just dilute to bring the concentration down again.

This post is quite lengthy so I'll continue with the other treatments in the next post.

[Seuxin]

Hmm, so you think using bicalutamide could be good ?

[Chemical]

Did some more reading on prostate cancer and AR beta-catenin interaction.

Functional localization and competition between the androgen receptor and T-cell factor for nuclear beta-catenin: a means for inhibition of the Tcf signaling axis

Reports have shown a role for the beta-catenin/Tcf pathway in prostate cancer including nuclear localization and beta-catenin mutations in primary prostate tumor samples (Chesire and Isaacs, 2002; Chesire et al., 2002). beta-Catenin has also been described as a ligand-dependent coactivator of the AR (Truica et al., 2000; Mulholland et al., 2002; Pawlowski et al., 2002; Yang et al., 2002). Previously, we have shown that translocating AR can provide a means of nuclear entry and accumulation of beta-catenin (Mulholland et al., 2002). This mode of beta-catenin trafficking has also been shown to hold true in neuronal cells (Pawlowski et al., 2002). Cotrafficking of AR and beta-catenin to the nucleus likely has important implications both for AR and Wnt signaling. While nuclear accumulation of beta-catenin has been shown to correlate with increased AR transcriptional activity, the effects on Tcf signaling have only begun to be explored. Recently, it has been shown that the AR has the ability to inhibit the beta-catenin/Tcf signaling pathway, ligand-dependently (Chesire and Isaacs, 2002). By way of transcriptional reporter assay, this report showed reduced luciferase activity for the Tcf reporter in a ligand-dependent manner in several prostate and colon cancer cell lines. This study also observed reduced transcriptional activity with the use of an AR deletion mutant (for beta-catenin binding), suggesting the possibility of a reciprocal balance of nuclear beta-catenin between the AR and Tcf. In the present study, we corroborate these results but provide mechanistic data to support the hypothesis that repression of the beta-catenin/Tcf signaling is mediated by ligand-occupied AR that is in competition with Tcf for nuclear beta-catenin. Specifically, using transcriptional reporter assays, we show that overexpression of WT Tcf reduced the activity of an AR (ARR3-Luc)-responsive reporter, while overexpression of a DeltaNt Tcf mutant did not have this effect.

Results

TOPFLASH activity = beta-catenin TCF/LEF activity

To evaluate the repressive effect that AR and its physiological ligand had on Tcf signaling, several titrations were performed in PC3 cells. Specifically, we observed decreased TOPFLASH activity with increased amounts of transfected AR (mug/three wells) with no detectable changes observed in either beta-catenin or Tcf4 protein levels (Figure 1a). Basal levels of BCT (TOPFLASH) in untreated PC3 cells were low, while in SW480 cells luciferase counts were 6–8-fold higher. In either cell line, control FOPFLASH values were about 10% of basal TOPFLASH levels. To confirm the ligand dependency of Tcf inhibition, we used AR deletion mutants (Figure 1b), including those coding the amino- and DNA-binding domain regions (Nt/DBD), as well as the DNA-binding region plus ligand-binding region (DBD/LBD). While the Nt/DBD mutant showed little ability to repress TOPFLASH, the DBD/LBD mutant was capable of a 3.5–4-fold repression in the presence of DHT. This suggests that in an AR overexpressed state, the ARNt is dispensable for Tcf repression. Further verification that the LBD is vital for repression is shown by increased relief of AR-mediated repression in cells treated with the pure AR antagonist Casodex (1 and 10 mum), which was able to efficiently relieve AR (DBD/LBD)-mediated repression of TOPFLASH. Having shown that repression is both AR and ligand dependent, we next evaluated the effect of increasing the concentration of DHT.

So bicalutamide reduced beta-catenin recruitment! Woohoo!

As a prerequisite, we verified that we could, in fact, detect an AR/beta-catenin complex upon transfection with AR (Figure 6b, arrow). Consistent with our previous studies (Mulholland et al., 2002), we detected more AR/beta-catenin complex from cells treated with DHT, suggesting a ligand-sensitive interaction. We also detected an association with beta-catenin and Tcf-HIS, but did not detect this association in the DeltaNt Tcf-myc deletion mutant. Importantly, using this assay, interactions between AR and Tcf were very weak as compared to our detected AR/beta-catenin and beta-catenin/Tcf complexes both in prostate and colon cancer cells. Nonimmune, control precipitations were only slightly less than those of the AR/Tcf complex, suggesting that only a small fraction of AR associates with Tcf. We wanted to know whether Casodex, which abrogated both HcRed-Tcf focal accumulation (data not shown) and TOPFLASH activity, could alter the binding of AR and beta-catenin. By treating cells with 5 mum Casodex (dissolved in EtOH) with and without 5 nm DHT, we observed reduced physical interaction between AR and beta-catenin (arrowhead), while also relieving AR-mediated repression of the beta-catenin/Tcf-HIS complex (Figure 6b, star and open arrow heads). To evaluate whether a decreased beta-catenin/TcfHIS complex could be directly affected by the AR TcfHIS and ARS35, in vitro translated products were used in series of binding assays with HIS-tagged recombinant beta-catenin (Figure 6c). Using Ni-NTA beads to precipitate in vitro binding reactions, we observed that upon increased levels of ARS35 (a–d) there was a corresponding decrease in TcfS35 (a–e) precipitated with the HIS-tagged recombinant beta-catenin. Although ARS35 levels up to 10 mul were added per binding reaction, we observed no further increase in detectable AR/beta-catenin-HIS binding greater than 6 mul (point d) or 60% of input by volume.

There is a saturation limit which we can exploit to get beta-catenin to activate TCF/LEF.

Some closing analysis:

The reduction of AR transcription (in the presence of DHT) upon overexpression of Tcf provides evidence of competition. These data are consistent with the notion that modulation of the beta-catenin-Tcf/Lef assembly may be the mechanism by which AR exerts its repression on Tcf/Lef signaling. To test this further, we evaluated the amount of beta-catenin associated with the Tcf/Lef complex with and without AR+androgen, both in vivo and in vitro. Data achieved with these data corroborate morphological data and our overall hypothesis.

Could it be that AR requires beta-catenin to exert its effects?

Other nuclear receptors and Tcf/Lef signaling

It is interesting how two stimulators of cell proliferation (AR/DHT and Wnt) can interact in a repressive manner. One possible interpretation is that androgens may not increase proliferation by, rather, promoting growth and differentiation in prostate epithelia. Prostate cancer cells have cell cycle deregulation compounded by a cell survival response to androgens. The contributions of Wnt signaling in prostate cancer are likely complex, although it is clear that nuclear beta-catenin can serve as a potent AR coactivator. We suggest a scenario whereby beta-catenin could be shuttling between Tcf/Lef-binding sites and AR elements. In the absence of androgen, AR resides mainly in the cytosol, while nuclear beta-catenin associates with Tcf (Figure 8a). In the presence of androgen, beta-catenin could be shuttled by translocating nuclear receptors to both Tcf- and AR-associated response elements to promote coactivation (Figure 8b). Consequently, less beta-catenin would be associated with Tcf and more with AR. This would simultaneously lower Tcf activity and augment AR transactivation.

Very very interesting stuff.