One question about time of egf inhibition

Stem cell; cell based hair therapy
1

i was re-reading the first patent of Cotsarelis (2006) to make some comparison about the methods .

While in new patent egf inhibition could start also before wounding , in 2006
we can read only after , 3-12 days.

Pag 253 of pdf version :
there is a picture with a timeline about a mouse experiment .
The egfr inhbition (with AG1487 compound) seems ( if i well understand)
to start at day 11.

even if this could not prove nothing (obviusly) i think cal has well said that results obtained do not confirm anyway an inhibition before wounding,
so ,with the hope to begin as soon as my experiment , i’ll go to inhibit egf
after wound and more of 3-4 days.

Any idea / suggest ?

you can find old patent here :
http://www.wipo.int/pctdb/en/fetch.jsp?LANG=ENG&DBSELECT=PCT&SERVER_TYPE=19-10&SORT=11252868-KEY&TYPE_FIELD=256&IDB=0&IDOC=1353744&C=10&ELEMENT_SET=B&RESULT=3&TOTAL=5&START=1&DISP=25&FORM=SEP-0/HITNUM,B-ENG,DP,MC,AN,PA,ABSUM-ENG&SEARCH_IA=US2006011319&QUERY=(IN%2Fcotsarelis)+ )

2

EXAMPLE 12

ENHANCEMENT OF EDIHN BY INHIBITION OF EGF RECEPTOR

[000229] To determine the effect of administration of EGF receptor inhibitors on DIHN, the inhibitor

P-7628-PC

AG1478 (150 μM in 10 μL volume) was administered as a single injection. 11 days after incisional wounding (1 cm2) to the middle of the wound near the skin surface. EGF receptor inhibitor administration led to generation of more and larger hair follicles compared with control mice that were wounded only (Figure 26A). As shown in Figure 26B, large hair follicles developed in the wounded area in the AG1478-injected mice. Left panel: epidermis stained for Kl 7, with three large hair follicles next to each other. Right panel: dermis stained for AP with large coalescing dermal papilla areas.

[000230] The findings of this Example confirm the results of the previous Example, and show that more and larger HF can be generated when EDIHN comprises, or is followed by, administration of EGFR inhibitors, or with compounds with a similar mechanism of action; e.g. Hedgehog protein and androgen antagonists.

The mice re-epilithialization period is longer than a humans is. This is why there is at least a three day wait. They noted this in two experiments (also experiment 11, they waited 11 days until the administered EGF to STOP hair from growing at the wound site…

Now weve got this cleared and out of the way. You need to wait at least three days before the EGF-antagonist.

3

I’m surprised to hear you referring to this issue as “cleared” benji.

I thought we were all still pretty unsettled on it. The Gentifib cancer regrowth case wasn’t using any waiting at all. And we were amassing a bunch of trains of thought that it made more sense not to wait at all either.

4

» I’m surprised to hear you referring to this issue as “cleared” benji.
»
» I thought we were all still pretty unsettled on it. The Gentifib cancer
» regrowth case wasn’t using any waiting at all. And we were amassing a
» bunch of trains of thought that it made more sense not to wait at all
» either.

For once I have never understood why the cancer patient have anything to do with the follica technology. Everyone seems surtain that he got a sunburn and that was what caused the hair growth. Sorry to say but that is just pure speculation, he could just be a freak accident who happend to regrow hair on medicine that is in the follica patent.

And second I dont understand why it makes more sense not to wait at all when the patent clearly states 3-14 days post wounding. A patent is to protect the technology from competition so why would they want to throw off some guys at a internet forum? I am for once convinced that cotsarelis and company knows more than us, even if it makes more sense not to wait.

I would also like to point out that no one seems to talk about dosage, most people seem to think that we just have to find an egfr inhibitor and everything will work. The person who failed (rev? cal?) with oral egfr inhibitor may not have had enough of it in his body/scalp. I believe a topical would be more efficient if we just could get a good formulation, the patent states how big concentration of egfr we need.

Just my two cents

5

» I’m surprised to hear you referring to this issue as “cleared” benji.
»
» I thought we were all still pretty unsettled on it. The Gentifib cancer
» regrowth case wasn’t using any waiting at all. And we were amassing a
» bunch of trains of thought that it made more sense not to wait at all
» either.

EXAMPLE 11

INHIBITION OF EDIHN BY EPIDERMAL GROWTH FACTOR INJECTION

[000226] 21 day-old mice were wounded as described in previous Examples. Starting from day 11 after wounding, a time point corresponding to the point at which the wound had recently re- epithelialized, 10 μL of 1 μg/ml EGF was injected into the wound bed. EGF was injected once per day after this point for a total of 5 days. Three days later, the skin was collected, and whole-mount

EDIHN assays were performed. EGF prevented HF formation as assessed by gross morphology. In addition, whole mounts of control and treated skin were analyzed with anti-K17 antibody immimostaining. All mice injected with EGF (n=4) exhibited no new HF formation (Figures 25 A-

B), while control mice (n=2) had many new HF5 as expected. (Figures 25 C-D).

[000227] In an additional experiment, recombinant EGF (1 microgram (mcg)/microliter (mcl)) was injected at days 11, 13 and 15 after wounding. Skin was collected at 18 days after wounding and stained for K,17 and alkaline phosphatase. Once again, administration of EGF inhibited EDIHN.

[000228] The findings of this Example show that EGF inhibits HF formation. Thus, inhibiting EGF, EGFR, or one of the pathways in which they participate increases EDIHN-induced HF formation.

Me again…I cant believe I overlooked this. These mice WILL form some hair when abraded alone, but by administering EGF when their skin had re-epilithialized, they were able to stop this. If they block EGF at day 11 (EXPERIMENT 12), they could enhance hair follicle formation. Obviously they mean what they say about skin needing to re-epilithialize. Whether or not EGF could be inhibited before re-epilthialization and hair placodes still form is anybody’s guess, but we KNOW they got growth when the agonist was used on day 11 post wounding in the mice. This would correspond to day 3-5 in us depending on the depth of the wound.

I think the immune system is the “biggie”. The immune system in a human being, MUCH MORE EVOLVED than what it is in a mouse, is going to respond to a disruption in its protective layer of the whole organism (the dermis) without fail. If anti-microbials and anti-inflammatories and anti-histamines can calm the wounded area and keep it infection free…then perhaps the immune system might not respond so mightily, but thats just conjecture. We know if the immune system is suppressed, hairs will grow (EXPERIMENT 7).
Haroldo (Harold Hallman) had noted in other experiments in mice that if there is inflammation the mice wont form hair when they are baby mice. It somehow will interdict the whole process.

6

» »»
» For once I have never understood why the cancer patient have anything to
» do with the follica technology. Everyone seems surtain that he got a
» sunburn and that was what caused the hair growth. Sorry to say but that is
» just pure speculation, he could just be a freak accident who happend to
» regrow hair on medicine that is in the follica patent.
Actually Z79, there are TWO people who got very unusual hair growth using Getfitinib. One of them got whisker-like growth on their nose. Thick growth of big dark hairs all over their nose. EGF is necessary for signalling to initiate the anagen phase in cycling hair follicles. If you blocked EGF all the time, it would suppress hair----not enhance it. The one guy who got the head growth had it right in the middle of long bald scalp-----where finasteride and minox and ketoconazole all together would not have “regrown it”. He was going grey also-----and the hair was dark brown. This is splendid evidence that the hair in question was indeed “new” hair formed via skin stem cells----and NOT old rejuvinated hair. It wouldn’t have been dark as the melanocytes in that old hair would have no doubt been aged like they were on the rest of his head. Over a thousand people have taken getfitinib. There would be more hypertrichotic effects if it grew hair out of its own accord. Its in the patent for a reason, which is EGF inhibtion at a unique time post-epidermal disrutption. »

» And second I dont understand why it makes more sense not to wait at all
» when the patent clearly states 3-14 days post wounding. A patent is to
» protect the technology from competition so why would they want to throw off
» some guys at a internet forum? I am for once convinced that cotsarelis and
» company knows more than us, even if it makes more sense not to wait.
There is also verbiage in the patent that claims that a topical with all the necessary embodiments can be applied at the same time as the wounding and even before the wounding takes place------albeit with the usage of “timed release” compounds therein. I thought it sounded rather flimsy and was wondering much as Cal is whether this was to throw people off so they would wait several days and get nothing. Make no mistake Z79, people can obtain (docs and nurses certainly can) just about everything in this patent. It can certainly be performed at home with the exception of some of the exotic “possible” adjuvants like beta catenin and ectodysplasin and some DNA fragments mentioned in the patent literature that very very likely wont be used. »

» I would also like to point out that no one seems to talk about dosage,
» most people seem to think that we just have to find an egfr inhibitor and
» everything will work. The person who failed (rev? cal?) with oral egfr
» inhibitor may not have had enough of it in his body/scalp. I believe a
» topical would be more efficient if we just could get a good formulation,
» the patent states how big concentration of egfr we need.
It does mention the concentration in the “kit” patent, but they “protect” all sorts of concentrations from down to .05% up to much much higher. They obviously plan on this being topical…but the therapuetic dosage internally of getfitinib is obviously “enough” to get hair. It has done it on two people. »

» Just my two cents
Any other observations that you have, please share it with us. We need all the perspectives we can get. There has got to be a way to do this…and its right there in the patent, but we are just not “seeing” it yet. My present position is that either the immune system will need to be antagonized or the skin will have to have an anti-inflammatory like a COX2 inhibitor put upon it so the immune system thinks it has nothing to react to. An anti-microbial might also be necessary…

7

if density and thickness is directly correlated with egf-inhibition topical dosage is important ( should be so… i’m wrong?)

We should try to eliminate all useless variables like immune system and skin too low dosage (in patent we can read "contacting skin with an amount sufficient).

I mean , is not any logically reason to believe immune system (or other) don’t became necessary in particulary moment of the process .Maybe dosage too high put in trouble the entire procedure.

But my feeling is :
-use medium high dosage for topical to overcame the possibility of lack of inhibition
(this if the assumption for dosage/hair density is true …in fact
goata007 asked for supplementary tables of the Nature article in which should be more data about )

-drop or reduce immune system

all this is quite possible in a topical way with tacrolimus (protopic/elidel cream) and gefitinib.

edit:
I’d also like to know if what i’ve read about the solubility of gefitinib is true
( i mean… gefitinib is soluble in DMSO but not in alcol).
if somebody can help, let we know :slight_smile:

I’m agree with benji, we’ve the need to more visual prospective ,
so every observation is appreciated.

I’m agree also with z79 when asserts that patent sure don’t want to throw off
internet guys .
Sorry for my english.

8

As for the question of whether I got enough oral EGF-R inhibition to adequately get the growth or not:

I took the normally recommended dosage of the drug. The gentifib cancer patients would most likely not have been taking a whole lot more than this either.

The drugs may not have been invented for hair purposes. But they were still invented to inhibit EGF-R regardless of what you’re trying to accomplish by doing that. And if Folica wants us to take a lot more than what’s ever been approved in the past, then this would be out of bounds of their desire to skip the full FDA trials.

Given that Folica’s patents explicitly say that oral systemic drugs WILL work, the normal dosage seemed like the obvious choice for me to try. I’m not gonna try taking a lot more than that unless Folica itself gives me specific reason to. (Hey, this stuff isn’t aspirin. Risks.)

9

»
» Now weve got this cleared and out of the way. You need to wait at least
» three days before the EGF-antagonist.

I’m still inclined to try inhibition DURING wounding. EGF has a definite role in epithelization. I can’t see any harm in extending the window to prior to the wounding or at least at the same time.

10

cal,
Is the EGFR inhibitor that you took, a small molecule drug? The patent & Zohar specifically mentioned small molecule drug(s), I’d say there probably is a good reason for that as well.

11

» As for the question of whether I got enough oral EGF-R inhibition to
» adequately get the growth or not:
»
»
» I took the normally recommended dosage of the drug. The gentifib cancer
» patients would most likely not have been taking a whole lot more than this
» either.
»
» The drugs may not have been invented for hair purposes. But they were
» still invented to inhibit EGF-R regardless of what you’re trying to
» accomplish by doing that. And if Folica wants us to take a lot more than
» what’s ever been approved in the past, then this would be out of bounds of
» their desire to skip the full FDA trials.
»
» Given that Folica’s patents explicitly say that oral systemic drugs WILL
» work, the normal dosage seemed like the obvious choice for me to try. I’m
» not gonna try taking a lot more than that unless Folica itself gives me
» specific reason to. (Hey, this stuff isn’t aspirin. Risks.)

But the patent focuses on topical application wouldn´t you agree? The oral mentioning is just a line or two and it it feels more like a “just in case someone tries to find a loop hole that eventually turns out to work”. And in your case it did not work which makes me believe even stronger that the wound needs to be in direct contact with the egfr inhibitor.

So the question I would like an answer to is how do we make a topical that can carry the small molecular egfr inhibitor through the skin without it loosing its function? Disolving leflunomide in alcohol did not work at least, I know that from my own experiments.

12

» »
» » Now weve got this cleared and out of the way. You need to wait at least
» » three days before the EGF-antagonist.
»
» I’m still inclined to try inhibition DURING wounding. EGF has a definite
» role in epithelization. I can’t see any harm in extending the window to
» prior to the wounding or at least at the same time.

I still think that the best approach would be not to wound the whole scalp at once but rather do one smaller area every day for a week (or every other day) and on the last day(s) you can apply your egfr inhibitor. By doing so you have covered a big span of healing time and even been on egfr inhibitor when doing the wounding. Do you follow me? Dont you think this would be the best way of doing, it would save a lot of time instead of doing many separate experiments.
Best of luck!

13

» » »
» » » Now weve got this cleared and out of the way. You need to wait at
» least
» » » three days before the EGF-antagonist.
» »
» » I’m still inclined to try inhibition DURING wounding. EGF has a
» definite
» » role in epithelization. I can’t see any harm in extending the window to
» » prior to the wounding or at least at the same time.
»
» I still think that the best approach would be not to wound the whole scalp
» at once but rather do one smaller area every day for a week (or every other
» day)
Thats a fair suggestion, but the thing is that we do not understand really how this approach works. Baccy did his entire scalp and yet the growth he had was ONLY in the crown area. WHY is that ? we dont know ! of course he talked about that wig …but what about the frontline ? so reducing the area involved with the experimentation can be counter-productive.

and on the last day(s) you can apply your egfr inhibitor. By doing so
» you have covered a big span of healing time and even been on egfr inhibitor
» when doing the wounding. Do you follow me? Dont you think this would be the
» best way of doing, it would save a lot of time instead of doing many
» separate experiments.
» Best of luck!

Baccy , you said that you didt realise that there was much activity until you saw, yourself, the picture. My question for you is the following : ARE you sure that you were not missing some hair left alive on your crown as you were always under the wig or shaving it to 0 . Men do tend to radically give up on hair after a certain level of hair loss is reached and think it’s all gone although they still have some of it but who cares as it does not give the desired appearance anymore, so they just forget about the few follicules left and consider they lost it all.

One more thing, was your crown the last castle to surrender ( to hairloss) ?What about frontline (was it the first) ?

14

My oral inhibitor drug was Leflunomide. Mentioned in the patents in the same breath as Gentifib. A small-molecule non-natural EGF-R inhibition drug.

Leflunomide and Gentifib . . . . they are alike in some ways but different in others. We need to keep this issue straight in our minds in order to correctly interpret what we are (and are not) discovering with all these experiments.

When comparing both drugs orally, Leflunomide should function like Gentifib for our EGF-R purposes. (Which is why I used it this first time in my oral EGF-R inhibition attempt.)

But when comparing them in terms of skin absorption, these two drugs ARE NOT the same stuff. Gentifib has a chemical makeup that leads one to think it will be skin-absorbed well. But Leflunomide is an unknown this way. It may work fine or it not get matabolized correctly enough to do anything.

Next time I will be switching my experimentation over to Gentifib. I think I’m gonna stay with Gentifib in the future whether I’m trying something orally or topically.

Further testing might eventually reveal that Leflunomide still works equally fine for our purposes. But in retrospect it’s also one more variable that’s not really necessary right now. Gentifib isn’t THAT much more money.

15

» My oral inhibitor drug was Leflunomide. Mentioned in the patents in the
» same breath as Gentifib. A small-molecule non-natural EGF-R inhibition
» drug.
»
»
»
» Leflunomide and Gentifib . . . . they are alike in some ways but different
» in others. We need to keep this issue straight in our minds in order to
» correctly interpret what we are (and are not) discovering with all these
» experiments.
»
» When comparing both drugs orally, Leflunomide should
» function like Gentifib for our EGF-R purposes. (Which is why I
» used it this first time in my oral EGF-R inhibition attempt.)
»
» But when comparing them in terms of skin absorption, these
» two drugs ARE NOT the same stuff. Gentifib has a chemical
» makeup that leads one to think it will be skin-absorbed well. But
» Leflunomide is an unknown this way. It may work fine or it not get
» matabolized correctly enough to do anything.
»
»
»
»
»
» Next time I will be switching my experimentation over to Gentifib. I
» think I’m gonna stay with Gentifib in the future whether I’m trying
» something orally or topically.
»
» Further testing might eventually reveal that Leflunomide still works
» equally fine for our purposes. But in retrospect it’s also one more
» variable that’s not really necessary right now. Gentifib isn’t THAT much
» more money.

Isn´t gentifib one of those expensive cancer drugs? It is good if we try different drugs and different methods (oral/topical), that is the only way we ever will crack this.

And regarding Baccys regrowth I suspect that the regrowth is not new follicles but rejuvenated since it is growing in areas which look like a late norwood patern (last to go, first to grow back). Otherwise I think he should have had more spread result all over the scalp. If I am right he should have little or no result in further experiments. I hope to be wrong of course.

Has anyone tried any other small molecular drugs mentioned in the patent besides leflunomide?

16

» When comparing both drugs orally, Leflunomide should
» function like Gentifib for our EGF-R purposes. (Which is why I
» used it this first time in my oral EGF-R inhibition attempt.)
»
» But when comparing them in terms of skin absorption, these
» two drugs ARE NOT the same stuff. Gentifib has a chemical
» makeup that leads one to think it will be skin-absorbed well. But
» Leflunomide is an unknown this way. It may work fine or it not get
» matabolized correctly enough to do anything.
»
» Next time I will be switching my experimentation over to Gentifib. I
» think I’m gonna stay with Gentifib in the future whether I’m trying
» something orally or topically.
»
» Further testing might eventually reveal that Leflunomide still works
» equally fine for our purposes. But in retrospect it’s also one more
» variable that’s not really necessary right now. Gentifib isn’t THAT much
» more money.

you’re fine , try to eliminate extra variables.

Also i’m agree with z79 about skin concetration of compound so please to consider topical way.

I don’t want suggest anybody to kill himself :slight_smile: so in previous post
my idea was referred to topical way .

gefitinib is soluble in DMSO ( unfortunately we know dmso can take the substance in system … but we need only 10 days so a concetration of 5 % maybe possibile)
tacrolimus is out there in cream/gel (protopic).

17

I disagree that we’re most likely looking at rejuvenated follicles. Maybe both things are going on, but I think it’s at least partly new ones being created.

Look at the nose regrowth pics. No way that’s existing follicles.

1109_image1.jpg1000×500

18

»
» Baccy , you said that you didt realise that there was much activity until
» you saw, yourself, the picture. My question for you is the following : ARE
» you sure that you were not missing some hair left alive on your crown as
» you were always under the wig or shaving it to 0 . Men do tend to radically
» give up on hair after a certain level of hair loss is reached and think
» it’s all gone although they still have some of it but who cares as it does
» not give the desired appearance anymore, so they just forget about the few
» follicules left and consider they lost it all.
»
»
» One more thing, was your crown the last castle to surrender ( to hairloss)
» ?What about frontline (was it the first) ?

First thing I did when I removed the dead rat that was resting on my head was to grow the hair out to see what I had left. The answer was that I had quite a bit of vellus hair but nothing terminal.
I don’t know whether these follicles are new or rejuvenated; in practical terms, it’s irrelevant. But there are some vague follicles appearing at the NW0 hairline. If they’re rejuvenated, they have been dormant since about 1979.

As for the last to surrender, I don’t know because while all that was going on, it was covered by the rat. I do know that the actual crown is being more stubborn than the top of the head to grow hair at the moment.

19

» I still think that the best approach would be not to wound the whole scalp
» at once but rather do one smaller area every day for a week (or every other
» day) and on the last day(s) you can apply your egfr inhibitor. By doing so
» you have covered a big span of healing time and even been on egfr inhibitor
» when doing the wounding. Do you follow me? Dont you think this would be the
» best way of doing, it would save a lot of time instead of doing many
» separate experiments.
» Best of luck!

I’m still going with the whole area at once. I think that the size of wound is important. And I want to inhibit EGF definitely BEFORE via topical tannic acid and internally via milk thistle extract. I’ll keep you apprised.

Note: If anyone starts messing with tannic acid, don’t be lulled into thinking it’s safe because it’s natural. In high concentrations, it causes severe liver damage. I’d hate for someone to get hurt trying out these things without considering the safety issues.

20

» » I still think that the best approach would be not to wound the whole
» scalp
» » at once but rather do one smaller area every day for a week (or every
» other
» » day) and on the last day(s) you can apply your egfr inhibitor. By doing
» so
» » you have covered a big span of healing time and even been on egfr
» inhibitor
» » when doing the wounding. Do you follow me? Dont you think this would be
» the
» » best way of doing, it would save a lot of time instead of doing many
» » separate experiments.
» » Best of luck!
»
» I’m still going with the whole area at once. I think that the size of
» wound is important. And I want to inhibit EGF definitely BEFORE via topical
» tannic acid and internally via milk thistle extract. I’ll keep you
» apprised.
»
» Note: If anyone starts messing with tannic acid, don’t be lulled into
» thinking it’s safe because it’s natural. In high concentrations, it causes
» severe liver damage. I’d hate for someone to get hurt trying out these
» things without considering the safety issues.

Baccy, why not give your temples special attention this time around? Your project didn’t seem to go too well on the temples on the last attempt. We all saw that there was nothing there and if you get that to sprout…