131 replies to this topic

[kmoody]

Ichor Therapeutics has an ongoing study to repeat the Longecity sponsored pilot study we ran last year. Our sponsors have requested that we provide interim reporting to benefit the Longecity community, so current data will be provided in this post.

 

BACKGROUND

The extreme life extension in Winsar rats given c60oo reported by Baati et al is well known to the Life Extension community. A less well known aspect of the study is the effect of c60oo on age-related cancer incidence. Batti's control rats all showed evidence of tumors at necropsy, as is typical for rodents. Yet despite their advanced age, none of the treatment groups had tumors. Based on these findings, our group conducted a pilot study to directly assess the utility of c60oo as a treatment for human cancer using a xenograft leukemia model. We observed a dose-dependent increase in median lifespan, though these results were not conclusive because of limited sample size (n=5 per group).

 

 

In the present study, we propose to replicate and confirm our findings with a larger cohort. We also aim to assess c60oo biodistribution within C57BL/6 and NOD/SCID mouse strains, and compare these results with reports from Winsar rats to validate our model. Lastly, it is thought that c60oo may persist in the body for extended periods of time, so we will include time-points after c60oo administration has ceased to determine its long-term presence.

 

RESEARCH PLAN

The first objective of the present study is to assess the effect of c60oo administration on human leukemia proliferation in vivo using a modified version of the protocol described by Lehne et al. Briefly, immunocompromised mice will be inoculated with 1x10⁷ Kg1a cells (human leukemia) by tail vein injection. After one week, mice will be treated with 8mg/kg c60oo or placebo every two days for six days, then once per week thereafter. Analytical endpoints will include complete blood counts and body mass. Additionally, a small cohort kept under identical conditions will be sacrificed 30 days post-inoculation to determine tumor burden by flow cytometric analysis of bone marrow aspirates. All procedures are routine to our laboratory.

 

The second objective of the present study is to determine c60oo biodistribution within standard C57BL/6 mice and the immunocompromised NOD/SCID strain. Briefly, mice will be treated by IP administration of 4mg/kg c60oo daily for 7 days and housed in metabolic cages. At days 1, 8, 30, and 90 mice (n=3) will be sacrificed and c60oo biodistribution within whole blood, liver, spleen, and brain will be quantified by HPLC analysis. These results will be compared to previous reports in Winsar rats to validate mice as a comparable model for current and future c60oo studies.

 

METHODS

HPLC

A PerkinElmer LC Flexar HPLC system, equipped with a UV/VIS detector and run with Chromera software was used for HPLC analysis (PerkinElmer, Inc. Waltham, Massachusetts, U.S). A reverse phase C18 column (4.6*150mm, 5um, Atlantis dc18, Waters Corp, US) was used for the entire HPLC analysis. A mixed solvent system of toluene and acetonitrile was used for the mobile phase, run isocratically with a composition of 70% toluene/30% acetonitrile with a flow rate of 0.5ml/min for a total run time of 15 minutes. Injection volumes were 20ul and the column was held at a constant temperature of 26 C.

 

Tissue extractions

10ul of internal standard (1mg/ml C70 in toluene) was placed into marked 15 ml falcon tubes and dried under a constant stream of air for 15 minutes. 100mg of liver and brain and 10mg of spleen were excised and placed into 1.7ml microcentrifuge tubes. 0.5ml 0.1M SDS solution was added to the pulverized tissue, and tissue samples were pulverized with glass rods until homogenous. The mixture was then transferred to the 15ml falcon tubes containing the dried internal standard. the original microcentrifuge tubes were rinsed with 0.5ml acetic acid and the rinsate was added to the same 15ml falcon tube as the tissue homogenate. samples were vortex mixed for 5 minutes. 3 ml of toluene were added and the mixture was agitated for 12 hours overnight in the dark at room temperature. Tubes were centrifuged for 10 minutes at 5000g and the top toluene layer was removed to a new 15ml falcon tube. Another 3 ml of toluene was added to the samples, which were agitated for 12 hours at room temperature in the dark once more. The centrifugation step was repeated and the toluene containing upper layer was removed and added to the same tube as the previous extraction. Supernatant was then dried down under a stream of warm air for 2 hours and the residue was dissolved in 500ul of mobile phase solution. 20ul of the sample was then injected into the HPLC.

 

Blood extractions

10ul of internal standard (1mg/ml C70 in toluene) was placed into marked 15 ml falcon tubes and dried under a constant stream of air for 15 minutes. 100ul of blood was placed into a 1.7ml microcentrifuge tube. 0.1ml 0.1M SDS solution was added and the mixture was then transferred to the 15ml falcon tubes containing the dried internal standard. the original microcentrifuge tubes were rinsed with 1ml acetic acid and the rinsate was added to the same 15ml falcon tube as the tissue homogenate. samples were vortex mixed for 5 minutes. 3 ml of toluene were added and the mixture was agitated for 12 hours overnight in the dark at room temperature. Tubes were centrifuged for 10 minutes at 5000g and the top toluene layer was removed to a new 15ml falcon tube. Another 3 ml of toluene was added to the samples, which were agitated for 12 hours at room temperature in the dark once more. The centrifugation step was repeated and the toluene containing upper layer was removed and added to the same tube as the previous extraction. Supernatant was then dried down under a stream of warm air for 2 hours and the residue was dissolved in 500ul of mobile phase solution. 20ul of the sample was then injected into the HPLC.

 

RESULTS

C60 was identified by HPLC and readily resolved from C70, the internal standard.

 

Concentrations of C60 in the blood, brain, liver, and spleen were quantified at D1 (group 1), D8 (group 2), and D30 (group 3). D90 data has not yet been collected.

 

 

 

 

During the pilot study, we failed to reliably quantify tumor burden by flow cytometry when analyzing Kg1a in peripheral mouse blood. In the present study, we modified our method to look in bone marrow instead. Kg1a populations clearly resolved at levels less than 1%.

 

A linearity test was performed comparing expected vs. actual Kg1a when mixed with bone marrow cells. The actual Kg1a measured was at lower levels than expected. This has been attributed to non-specificity in the FSC vs SSC gating. A viability dye was not included in the present assay, so a reliable all-cells gate could not be established. However, this assays appears to be valid as a relative measure of tumor burden.

 

 

Interestingly, tumor burden in the bone marrow of NOD/SCID mice (D30) was higher than that of olive oil or saline controls. This data cannot be fully evaluated in the absence of all-cause mortality data because multiple variables were changed since the pilot study. For example, the pilot study relied on in-house manufactured C60oo and CIEA NOG mice whereas the current study relied upon SES research C60oo and NOD/SCID mice.

 

 

Ichor is currently developing GMP grade analytical chemistry methods to fully evaluate C60oo quality and remove this variable from consideration. Death curves among the mice are expected to begin within 1-2 weeks. Data will be posted as it becomes available.

[bixbyte]

Briefly, immunocompromised mice will be inoculated with 1x10⁷ Kg1a cells (human leukemia) by tail vein injection. After one week, mice will be treated with 8mg/kg c60oo or placebo every two days for six days, then once per week thereafter. 

 

 

I would have first started the mice on C60 for a couple weeks and then inoculated them with Human Leukemia.

 

[aribadabar]

 

Briefly, immunocompromised mice will be inoculated with 1x10⁷ Kg1a cells (human leukemia) by tail vein injection. After one week, mice will be treated with 8mg/kg c60oo or placebo every two days for six days, then once per week thereafter. 

 

 

I would have first started the mice on C60 for a couple weeks and then inoculated them with Human Leukemia.

 

 

That's a good idea but in real world you get diagnosed first with a disease and THEN start treatment - which this study tries to simulate.

[sensei]

 

 

Briefly, immunocompromised mice will be inoculated with 1x10⁷ Kg1a cells (human leukemia) by tail vein injection. After one week, mice will be treated with 8mg/kg c60oo or placebo every two days for six days, then once per week thereafter. 

 

 

I would have first started the mice on C60 for a couple weeks and then inoculated them with Human Leukemia.

 

 

That's a good idea but in real world you get diagnosed first with a disease and THEN start treatment - which this study tries to simulate.

 

 

So pre-exposure prophylaxis is not a real-world phenomenon?

 

Preventative trials are NEVER executed?

 

Really?

 

In the Baati study they pre-treated with C60OO -- before CCl4 (Carbon tetrachloride) challenge.

 

Pre-treatment would more closely mimic the effects of C60  on emergent cancer cells caused by environmental and epigenetic oncological mutations

Edited by sensei, 21 December 2015 - 05:52 AM.

[aribadabar]

 

 

 

Briefly, immunocompromised mice will be inoculated with 1x10⁷ Kg1a cells (human leukemia) by tail vein injection. After one week, mice will be treated with 8mg/kg c60oo or placebo every two days for six days, then once per week thereafter. 

 

 

I would have first started the mice on C60 for a couple weeks and then inoculated them with Human Leukemia.

 

 

That's a good idea but in real world you get diagnosed first with a disease and THEN start treatment - which this study tries to simulate.

 

 

So pre-exposure prophylaxis is not a real-world phenomenon?

 

Preventative trials are NEVER executed?

 

Really?

 

In the Baati study they pre-treated with C60OO -- before CCl4 (Carbon tetrachloride) challenge.

 

Pre-treatment would more closely mimic the effects of C60  on emergent cancer cells caused by environmental and epigenetic oncological mutations

 

 I didn't dismiss the idea - in fact I said it is a good one...it just doesn't happen.

I didn't mean trials, I meant real-life treatments.

 

How many undergo preventative chemotherapy prior to cancer diagnosis?

 

Again, I don't disagree that pre-treatment would be beneficial.

[sensei]

 

 

How many undergo preventative chemotherapy prior to cancer diagnosis?

 

 

 

Everyone that believes Pauling and Cathcart and takes multi-gram doses of ascorbate daily.

 

I'd put the number of people that take large doses of vitamin C (1000mg/day or more) specifically as a cancer preventative in the millions.

 

 

That NONE of the Baati C60 rats had tumors is astounding -- pointing more to a preventative role.  The C60 was administered years before death, and months to years before most Wistar rats even get cancer. 

Edited by sensei, 21 December 2015 - 02:04 PM.

[kmoody]

 

 

 

 

Briefly, immunocompromised mice will be inoculated with 1x10⁷ Kg1a cells (human leukemia) by tail vein injection. After one week, mice will be treated with 8mg/kg c60oo or placebo every two days for six days, then once per week thereafter. 

 

 

I would have first started the mice on C60 for a couple weeks and then inoculated them with Human Leukemia.

 

 

That's a good idea but in real world you get diagnosed first with a disease and THEN start treatment - which this study tries to simulate.

 

 

So pre-exposure prophylaxis is not a real-world phenomenon?

 

Preventative trials are NEVER executed?

 

Really?

 

In the Baati study they pre-treated with C60OO -- before CCl4 (Carbon tetrachloride) challenge.

 

Pre-treatment would more closely mimic the effects of C60  on emergent cancer cells caused by environmental and epigenetic oncological mutations

 

 I didn't dismiss the idea - in fact I said it is a good one...it just doesn't happen.

I didn't mean trials, I meant real-life treatments.

 

How many undergo preventative chemotherapy prior to cancer diagnosis?

 

Again, I don't disagree that pre-treatment would be beneficial.

 

 

I think we can agree that the prophylactic properties and direct therapeutic properties of C60oo may be distinct, and may need to be answered in different ways. The xenograft model we have elected to use here is not suitable for prophylactic studies. The Kg1a cells are grown ex vivo and administered by tail vein to the mice. This is not a suitable representation of de novo cancer formation. Any "prophylactic" efficacy we might have observed could just as easily been attributable to confounding variables in this model, such as limiting engraftment efficiency. We have an ongoing lifespan study using C60oo as an intervention arm so that group should be telling about any prophylactic properties C60oo may have.

 

 

 

 

 

How many undergo preventative chemotherapy prior to cancer diagnosis?

 

 

 

Everyone that believes Pauling and Cathcart and takes multi-gram doses of ascorbate daily.

 

I'd put the number of people that take large doses of vitamin C (1000mg/day or more) specifically as a cancer preventative in the millions.

 

 

That NONE of the Baati C60 rats had tumors is astounding -- pointing more to a preventative role.  The C60 was administered years before death, and months to years before most Wistar rats even get cancer. 

 

 

 

Assuming the results are valid. I have yet to see any group confirm Baati's study (which is why we are attempting to do so). It would be very interesting if this claim holds true.

[Wilberforce]

I believe I saw a study where c60 fed rats were then injected with carcinogens or cancer cells and they remained immune. I agree with the OP, wouldn't this be a valid experiment to extend?

[kmoody]

I believe I saw a study where c60 fed rats were then injected with carcinogens or cancer cells and they remained immune. I agree with the OP, wouldn't this be a valid experiment to extend?

I would be happy to review the paper and see if this is something we could do in house if there were interest. Could you post a link or the PMID?

 

The reality is that there are tons of valid experiments we could do to characterize the effects of C60oo, but funding is always a limiting factor. Part of why we have chosen the experiments we are currently working on is that positive results could be leveraged more easily to finance further studies. It is very hard to raise capital for prophylactic interventions with nebulously defined mechanisms of action, and the studies take a lot longer (i.e. more expensive) than acute models. Additionally, there are important differences between aging and cancer in rodents vs. humans. The pharmaceutical industry is littered with drug candidates that cured every disease known to man in mouse models, but failed to do anything substantial in people.

Edited by kmoody, 22 December 2015 - 07:07 PM.

[Wilberforce]

It's valid to defend the aim of an experiment , I think what's being said here is that there could be a different aim.

[sensei]

It's valid to defend the aim of an experiment , I think what's being said here is that there could be a different aim.

 

Not taking anything away from Ichor, however:

 

there is little monetization available for a preventative/prophylactic anything, and I don't believe that the development of prophylactic drugs is part of Ichor's business model -- except as perhaps part of the contract research.

 

With the emerging data that shows environmental and lifestyle choices are overwhelmingly the cause of cancer, a cancer preventative would face a huge epidemiological and evidence battle, as would any "life-extension" drug.

 

Marie Calment ate 2 lbs of dark chocolate a week and reportedly drowned her food (most meals) in olive oil -- combination of anti-oxidant and olive oil  -- correlated? causal? how long a study would suffice.

 

Tweak a C60OO molecule enough to get a patent, and show efficacy against existing disease and you have a cheap to manufacture almost priceless drug.

 

Truvada was initially a  HAART treatment for AIDS and HIV patients, that just so happens to be an extremely potent and efficacious pre-exposure prophylactic anti-viral with respect to HIV.

Edited by sensei, 23 December 2015 - 03:47 AM.

[password]

http://www.jimmunol....gAbstracts/55.2

 

kmoody, this is exiting work, thanks for keeping us informed on your progress.

 

I was drawn to this forum because of a thoughtful post by niner on fullerenes and asthma.  In regards to the study design, the study linked above (for some reason I can't link inline) had two treatment groups, one treated during the development of asthma and another treated after.  So they were able to test both the preventative and "after infection" treatment value of C70 for asthma.

 

On a different topic, could someone help put these initial results in layman terms?  Is this indicating that C60oo treatment is causing more tumors to form in bone marrow?

 

 

 

[ambivalent]

In determining the structure of the study a critical question might be what are or should be the overarching objectives of SENS’s investment? Is it to demonstrate c60oo as a potential effective cancer therapy? Or is a significant part of the aspiration to gain wide-reaching attention and subsequent investment in what the community here certainly believe to be something truly spectacular within and outside the domain of cancer treatment? The inclusion of a pre-AML C60oo group at the expense of an intermediate-dosing group would improve the chances of this occurring (if demonstrating high efficacy of course). Personally I would favour three groups: control; high dose and pre-AML C60oo. Although intermediate dosing would be useful, I feel, the investment required to fund those groups could be spent demonstrating the efficacy of c60oo elsewhere (or, still, albeit less usefully, creating larger groups within the study)

I am concerned that given the anecdotes here over the years of the potential for c60oo to be an effective treatment against a number of wide ranging conditions we seem only to be pursuing a line of deductive research based on the Baati & Moody studies. We have considerable albeit uncatalogued and uncollated human usage here and yet we have no lines of research based on any of it. It is hard to see for example, given the current line of c60oo related cancer research, how investment in c60oo for its efficacy on say burn injuries or respiratory disorders (or even one hopes neurological disorders*) will begin. Anecdotes are discounted in value against hard evidence when allocating research investment but they should not be rendered irrelevant : there is a very significant non-zero probability that the effects claimed by members of the forum are real and there is a very large impact on society if they are demonstrated to be true - this reasoning renders the justification of the relatively small investment of tens of thousands of dollars on what could be a life-changing therapy for millions a trivial matter.  

Isn’t it time as a community we start to raise funds for studies in what would would seem to be orthogonal conditions to those currently pursued the research?

 

 

          

*I’m not sure we have forum accounts for efficacy here.

Edited by ambivalent, 23 December 2015 - 09:56 PM.

[kmoody]

http://www.jimmunol....gAbstracts/55.2

 

kmoody, this is exiting work, thanks for keeping us informed on your progress.

 

I was drawn to this forum because of a thoughtful post by niner on fullerenes and asthma.  In regards to the study design, the study linked above (for some reason I can't link inline) had two treatment groups, one treated during the development of asthma and another treated after.  So they were able to test both the preventative and "after infection" treatment value of C70 for asthma.

 

On a different topic, could someone help put these initial results in layman terms?  Is this indicating that C60oo treatment is causing more tumors to form in bone marrow?

The C60oo treated mice have double the tumor burden within the bone marrow as compared to controls. Most of our C60oo treated mice also have palpable tumors whereas most controls do not. We have a few working hypotheses to explain this data.

 

It could be that C60oo exacerbates cancer. However, I do not believe this to be the case. Our pilot study showed a very compelling, dose-dependent decrease in all-cause mortality.

 

I am aware of three major changes to the study design between the pilot and our current study. One was a switch from CIEA NOG to NOD/SCID mice. These are very similar immunocompromised mouse models that I don't think are sufficiently distinct to cause this discrepancy, but an essential non-obvious difference could exist. Another possibility is the lag time with treatment. In the pilot study we began treating with C60oo when the mice were inoculated with the leukemia cells. In the repeat, we wanted to clearly differentiate the effects of prophylactic treatment vs. reduced engraftment of cancer vs. cancer treatment. We chose to study the cancer treatment, and waited until 1 week after the cancer was injected before beginning treatment. It could be that properties of C60 reduce tumor engraftment but exacerbate cancer. Again, I do not believe this to be the case.

 

Our current lead hypothesis is sourcing. For our pilot study, we made C60oo fresh in-house. At the time of the current study, we wanted to add quality assurance to the study design so we chose to go with a vendor that claimed robust product specifications. We have 3 ongoing studies that are using C60oo sourced from that vendor, including the leukemia one described here. In all three studies, the C60oo treated mice are trending heavily in the direction of accelerated death from all-cause mortality. In our repeat lifespan study, we have confirmed by blinded 3rd party histopathology that at least some C60oo treated mice are dying of cancer. The product from this vendor has failed every one of our quality control tests as we have brought them online in-house. We have repeatedly asked for CoA and product specification documents but these forms have not been provided.

 

So in layman terms, something bad appears to be happening to these mice. A plausible common denominator is that C60oo sourcing is incredibly important. Some vendors may have a product that could promote all the health benefits we hear about with C60oo. Some vendors may have a product that can dramatically increase all cause mortality. We are aggressively studying the differences between our formulation and that of the vendor to understand what the difference may be.

 

 

It's valid to defend the aim of an experiment , I think what's being said here is that there could be a different aim.

 

Not taking anything away from Ichor, however:

 

there is little monetization available for a preventative/prophylactic anything, and I don't believe that the development of prophylactic drugs is part of Ichor's business model -- except as perhaps part of the contract research.

 

With the emerging data that shows environmental and lifestyle choices are overwhelmingly the cause of cancer, a cancer preventative would face a huge epidemiological and evidence battle, as would any "life-extension" drug.

 

Marie Calment ate 2 lbs of dark chocolate a week and reportedly drowned her food (most meals) in olive oil -- combination of anti-oxidant and olive oil  -- correlated? causal? how long a study would suffice.

 

Tweak a C60OO molecule enough to get a patent, and show efficacy against existing disease and you have a cheap to manufacture almost priceless drug.

 

Truvada was initially a  HAART treatment for AIDS and HIV patients, that just so happens to be an extremely potent and efficacious pre-exposure prophylactic anti-viral with respect to HIV.

 

I agree with sensei on this one. Happy to run contract research near cost for the benefit of the community, but the value to us as a business is in drugs. That said, there is obvious overlap in the safety profile of C60oo as a drug candidate or a prophylactic supplement, so robust characterization of manufacturing, formulation, stability, and safety issues is where everyone's interests are likely aligned. Note that the initial study was not really meant to show C60oo can treat cancer. Rather, it was to see if it would exacerbate a human cancer model in mice.

[password]

...
 
Our current lead hypothesis is sourcing. For our pilot study, we made C60oo fresh in-house. At the time of the current study, we wanted to add quality assurance to the study design so we chose to go with a vendor that claimed robust product specifications. We have 3 ongoing studies that are using C60oo sourced from that vendor, including the leukemia one described here. In all three studies, the C60oo treated mice are trending heavily in the direction of accelerated death from all-cause mortality. In our repeat lifespan study, we have confirmed by blinded 3rd party histopathology that at least some C60oo treated mice are dying of cancer. The product from this vendor has failed every one of our quality control tests as we have brought them online in-house. We have repeatedly asked for CoA and product specification documents but these forms have not been provided.
...

 
That's scary.  Many of us in the US making home-made C60oo are sourcing from SES; which, if I understand correctly, is the same source you used for your recent studies.
 
That's probably a discussion for this thread, but maybe you can help clarify the HPLC graph you provided.  What is this graph measuring?  The premade C60oo?  Or is it demonstrating calibration?  My apologies if the answer to these questions are self-evident (I'm not a trained scientist).

[Turnbuckle]

 

 

 

 

 
That's scary.  Many of us in the US making home-made C60oo are sourcing from SES; which, if I understand correctly, is the same source you used for your recent studies.
 

 

 

People are using the SES C60, but not necessarily the dissolved product. I would steer clear of it as they are apparently using ultrasonic energy that might create a different spectrum of adducts. They also give it a 3-year shelf life, which suggests they may be selling rather old stuff.

[sensei]

 

 

 

 

 

 
That's scary.  Many of us in the US making home-made C60oo are sourcing from SES; which, if I understand correctly, is the same source you used for your recent studies.
 

 

 

People are using the SES C60, but not necessarily the dissolved product. I would steer clear of it as they are apparently using ultrasonic energy that might create a different spectrum of adducts. They also give it a 3-year shelf life, which suggests they may be selling rather old stuff.

 

 

Ultrasonic cavitation has recently been shown to induce extremely [unexpectedly] high energies within the micro-bubble,  and also contribute energy to chemical processes.

 

Here is an old paper from MIT http://web.mit.edu/d...nochemistry.pdf

 

there is newer literature as well.

 

I completely concur with Turnbuckle that SES is likely creating FrankenC60OO by using ultrasound.   The issue may not even be with the C60 adducts -- it could easily be from non C60 related sonochemical interaction of constituents of olive oil.

Edited by sensei, 24 December 2015 - 01:56 AM.

[Major Legend]

 

 

 

 

 

 

 
That's scary.  Many of us in the US making home-made C60oo are sourcing from SES; which, if I understand correctly, is the same source you used for your recent studies.
 

 

 

People are using the SES C60, but not necessarily the dissolved product. I would steer clear of it as they are apparently using ultrasonic energy that might create a different spectrum of adducts. They also give it a 3-year shelf life, which suggests they may be selling rather old stuff.

 

 

Ultrasonic cavitation has recently been shown to induce extremely [unexpectedly] high energies within the micro-bubble,  and also contribute energy to chemical processes.

 

Here is an old paper from MIT http://web.mit.edu/d...nochemistry.pdf

 

there is newer literature as well.

 

I completely concur with Turnbuckle that SES is likely creating FrankenC60OO by using ultrasound.   The issue may not even be with the C60 adducts -- it could easily be from non C60 related sonochemical interaction of constituents of olive oil.

 

 

Thats worrying because I have been using SES fortunately, I've only taken a very small amount of c60. This issue may explain why different vendors have different colours compared to each other (light red, ruby red, or violet) . There has also been documented change in colour over time, which may indicate ratios of c60 derivatives are changing via reaction with air which consists of a bunch of stuff more than just Oxygen.

 

Would be good to get a clarification of the vendors used in the ongoing tests and the pilot study

Edited by Major Legend, 24 December 2015 - 03:49 AM.

[Kalliste]

Looking forward to the entire quality control. This is a bet.

[kmoody]

 

...

 
That's probably a discussion for this thread, but maybe you can help clarify the HPLC graph you provided.  What is this graph measuring?  The premade C60oo?  Or is it demonstrating calibration?  My apologies if the answer to these questions are self-evident (I'm not a trained scientist).

 

The HPLC graph just shows we can easily resolve C60 by HPLC, and we show decent linearity down to pretty low concentrations. We use C70 as an internal standard when harvesting C60 from tissues and organs to assess biodistribution (extraction isn't 100%, so adding a known concentration of C70 allows us to know what our percent yield is and we can adjust the C60 measurements to account for loss during the extraction process).

 

 

People are using the SES C60, but not necessarily the dissolved product. I would steer clear of it as they are apparently using ultrasonic energy that might create a different spectrum of adducts. They also give it a 3-year shelf life, which suggests they may be selling rather old stuff.

Agreed. I should emphasize this point. Our pilot study used SES research C60 mixed in house with off-the-shelf olive oil. Our repeat studies used SES research C60oo formulation. It may be as simple an issue as freshness of the product.

 

 

 

Ultrasonic cavitation has recently been shown to induce extremely [unexpectedly] high energies within the micro-bubble,  and also contribute energy to chemical processes.

 

Here is an old paper from MIT http://web.mit.edu/d...nochemistry.pdf

 

there is newer literature as well.

 

I completely concur with Turnbuckle that SES is likely creating FrankenC60OO by using ultrasound.   The issue may not even be with the C60 adducts -- it could easily be from non C60 related sonochemical interaction of constituents of olive oil.

Do we know SES research makes their C60oo by ultrasound or are we speculating here?

 

 

 

Thats worrying because I have been using SES fortunately, I've only taken a very small amount of c60. This issue may explain why different vendors have different colours compared to each other (light red, ruby red, or violet) . There has also been documented change in colour over time, which may indicate ratios of c60 derivatives are changing via reaction with air which consists of a bunch of stuff more than just Oxygen.

 

Would be good to get a clarification of the vendors used in the ongoing tests and the pilot study

 

Agreed. I have requested this information repeatedly over the last month or so (approximately weekly). They shipped me pure olive oil which is helpful for our method development, but they have yet to reply with any quality assurance data or discussion thereof. It is the holidays though so it could be that their QC person is simply out of the office.

Edited by kmoody, 24 December 2015 - 02:35 PM.

[ambivalent]

Once again thanks very much for your dilligence to the c60 community; is there are anyway of helping fund the cost of your research?

 

 

I wonder if it is useful to study a group of mice on a course of your in-house c60 after being previously being dosed with the apparent dodgy batch. It might be useful information for those concerned about the product they've taken.

 

 

 

 

[sthira]

Once again thanks very much for your dilligence to the c60 community; is there are anyway of helping fund the cost of your research?
.

Yes I agree, and would happily donate to this, and I'm sure others here would, too. We want human studies, though, and since there already exist human self-experimenters with this stuff, why not study people and leave the poor mice alone? But I think I already know your answer :-(

[sensei]

 

Do we know SES research makes their C60oo by ultrasound or are we speculating here?

 

 

 

 

According to smithx, he contacted SES Research regarding their C60OO and they told him they used ultrasound to dissolve it faster  - quote and thread below

 

"SES:

SES has their own C60OO, which is made using their 99.95% C60. (https://sesres.com/C60-olive-oil.asp)

Normally one would think this would be a great product, because it's made by the manufacturer of the C60 using their purest product, but I spoke with them and I have doubts. First of all, they use ultrasound sonication to get the C60 to dissolve faster. I've previously read here that this may not form the same adducts we are looking for. Secondly, they filter using a 5 micron filter, and do see a lot of undissolved solids on the filter paper. This implies that a lot of clumps are getting through their process."

 

http://www.longecity...e-oil-supplier/

[ambivalent]

 

Once again thanks very much for your dilligence to the c60 community; is there are anyway of helping fund the cost of your research?
.

Yes I agree, and would happily donate to this, and I'm sure others here would, too. We want human studies, though, and since there already exist human self-experimenters with this stuff, why not study people and leave the poor mice alone? But I think I already know your answer :-(

 

 

I certainly agree the community is well overdue funding tests on long-term and high-dose users.   

[kmoody]

I certainly agree the community is well overdue funding tests on long-term and high-dose users.   

Agreed. My group may be able to do some inexpensive/pro bono work (helping to coordinate, track C60 plasma half-life in peripheral blood, etc.) if there is serious interest here.

[sensei]

 

I certainly agree the community is well overdue funding tests on long-term and high-dose users.   

Agreed. My group may be able to do some inexpensive/pro bono work (helping to coordinate, track C60 plasma half-life in peripheral blood, etc.) if there is serious interest here.

 

 

That is very interesting.

 

The Baati study showed that all C60 was gone from blood by 97h (based on half-life evaluation).

 

What protocols would be necessary to capture multiple blood draws over such a short course of elimination?

 

As a high dose user, not afraid to be a guinea pig, I would be willing to volunteer for dosing of 1.7 mg/kg daily for a week and may even consider individual doses as high as 4 mg/kg (to see the effects of such an acute dose on a human -- as the effects on rats (accumulation in spleen and liver) are documented in Baati)  -- although gulping down 400ml

(3600 kcal- talk about sweats)

of C60OO may be challenging     no ip administration only oral -- and I'm not really that partial to gavage 

 

If the community/Ichor is interested in funding travel and associated costs -- I am willing to support as my schedule would permit

Edited by sensei, 24 December 2015 - 10:26 PM.

[ambivalent]

It had to be you Sensei :-) Can we set up a fund raiser here shortly and figure out what we need later (let's harness festive community spirit!)? I don't think we should just cost out the work for Sensei's potential study and fund on that basis alone but rather provide ongoing resource for Kelsey: I'm pretty sure we'd not have a funding problem. Would there be any other useful metrics?

Edited by ambivalent, 24 December 2015 - 11:12 PM.

[sensei]

It had to be you Sensei :-) Can we set up a fund raiser here shortly and figure out what we need later (let's harness festive community spirit!)? I don't think we should just cost out the work for Sensei's potential study and fund on that basis alone but rather provide ongoing resource for Kelsey: I'm pretty sure we'd not have a funding problem. Would there be any other useful metrics?

 

 

I think some of the metrics we should look at as changed from baseline (and that raises issues of washout for long-term and/or high dose users like me and others) include but are not limited to

 

GSH-Px

SOD

MDA

 

As noted earlier months ago in another thread, I had blood tests prior to starting and twice during dosing and there were no statistically significant changes in blood panels.

 

The blood panels are those taken during yearly physicals in the US including A1C, electrolytes, platelets, etc

[Logic]

Niner:

"I shouldn't have left that hanging...  There are multiple effects that c60oo has in the body, with a number of different pharmacologic mechanisms.  There is more than one possible product formed when c60 reacts with olive oil, and the spectrum of effects that you see depends on the levels of the different products.  One of the effects that I like is an immunomodulatory one that controls my eczema better than anything I've ever found, and also improves my breathing.  I noticed that a batch that was poorly stored lost this effect over time, and speculated that it was being destroyed by oxidation, since the oil had been kept in a large partially filled bottle "sealed" with a cork.  I made a batch using high quality ingredients and excluding air as much as possible.  This batch, to my surprise, had no anti-eczema effect to speak of.  I made a new batch where I ground the c60 in air, and used the same high quality oil.  It had a little of the effect, but not much.  I made another batch using a lower quality oil that had been in use for a while, and would be expected to have a much higher level of peroxides than the good oil.  I used the same c60 as the previous batch.  The first thing that I noticed was that the old oil reacted far faster; all solids were gone in three days, while the good oil took two weeks of stirring and still had some small specks.  This new batch appears to have the desired anti-eczema effect, and also feels like the enhanced endurance effect is better.   Recently, Franco Cataldo published a paper on the interaction of c60 with vegetable oils, and said that it goes through a peroxide intermediate.  I think it might be the case that the peroxide reaction is faster, but that there are other possible reactions as well.  It is likely that these other reactions, assuming they exist, result in different products, explaining the different pharmacodynamic effects.  The upshot of all this is that you might be better off using a cheap oil for making c60oo, and using the good oil on your salad and vegetables."

http://www.longecity...ndpost&p=697908

 

 

 

[kmoody]

 

What protocols would be necessary to capture multiple blood draws over such a short course of elimination?

 

As a high dose user, not afraid to be a guinea pig, I would be willing to volunteer for dosing of 1.7 mg/kg daily for a week and may even consider individual doses as high as 4 mg/kg (to see the effects of such an acute dose on a human -- as the effects on rats (accumulation in spleen and liver) are documented in Baati)  -- although gulping down 400ml

(3600 kcal- talk about sweats)

of C60OO may be challenging     no ip administration only oral -- and I'm not really that partial to gavage 

 

If the community/Ichor is interested in funding travel and associated costs -- I am willing to support as my schedule would permit

 

A good pilot may be to go without C60oo for at least two weeks (perhaps 1 month) then do a single dose at 1mg/kg. It would be useful to see peripheral blood samples at T0 (before ingestion) then at various time points over D1, 2, and 3. Based on Baati et al hourly for 12 hours on D1 then at 24 and 48 hours should give decent resolution. The easiest way to do this would be if you knew a phlebotomist who could just pull the samples and store in the fridge, then you could send overnight on ice for analysis. I cannot fund or directly support this work without IRB approval, but if you want to just send me blood samples to test for C60oo or biomarkers I should be able to run them.

 

It had to be you Sensei :-) Can we set up a fund raiser here shortly and figure out what we need later (let's harness festive community spirit!)? I don't think we should just cost out the work for Sensei's potential study and fund on that basis alone but rather provide ongoing resource for Kelsey: I'm pretty sure we'd not have a funding problem. Would there be any other useful metrics?

GSSG/GSH ratio would be interesting. This work should be done in consult with a primary care physician. Standard labs can be done via physician or LEF (see http://www.lifeexten...bc-blood-test).

 

COI Disclaimer: LEF has provided my group with substantial funding for our leukemia work.

 

Niner:

"I shouldn't have left that hanging...  There are multiple effects that c60oo has in the body, with a number of different pharmacologic mechanisms.  There is more than one possible product formed when c60 reacts with olive oil, and the spectrum of effects that you see depends on the levels of the different products.  One of the effects that I like is an immunomodulatory one that controls my eczema better than anything I've ever found, and also improves my breathing.  I noticed that a batch that was poorly stored lost this effect over time, and speculated that it was being destroyed by oxidation, since the oil had been kept in a large partially filled bottle "sealed" with a cork.  I made a batch using high quality ingredients and excluding air as much as possible.  This batch, to my surprise, had no anti-eczema effect to speak of.  I made a new batch where I ground the c60 in air, and used the same high quality oil.  It had a little of the effect, but not much.  I made another batch using a lower quality oil that had been in use for a while, and would be expected to have a much higher level of peroxides than the good oil.  I used the same c60 as the previous batch.  The first thing that I noticed was that the old oil reacted far faster; all solids were gone in three days, while the good oil took two weeks of stirring and still had some small specks.  This new batch appears to have the desired anti-eczema effect, and also feels like the enhanced endurance effect is better.   Recently, Franco Cataldo published a paper on the interaction of c60 with vegetable oils, and said that it goes through a peroxide intermediate.  I think it might be the case that the peroxide reaction is faster, but that there are other possible reactions as well.  It is likely that these other reactions, assuming they exist, result in different products, explaining the different pharmacodynamic effects.  The upshot of all this is that you might be better off using a cheap oil for making c60oo, and using the good oil on your salad and vegetables."

http://www.longecity...ndpost&p=697908

This is an interesting observation. The C60oo we used for the pilot study used standard olive oil, not the fancy stuff.