Very preliminary first impression and questions.
SCIENCE: Its a completely daft idea - but that is great, in keeping with the Imminst 'niche'. People don't really seem to understand LLLT either. Personally, I can't imagine that you'll get any effective deep tissue penetration. Laser ablation is successful in the eye and on the skin where you have easy access and acess issues are why people want to use nanoparticles as (laser)inducible agents. Regrettably, this project will skirt around the issue by lasering cells in the dish, but I fully accept that, as a proof of principle, this may be a useful first step. Maybe you could check if you are able to penetrate (porcine) skin explants and make a difference to cells placed just underneath?
This is getting a little ahead of myself, but the penetration depth problem has already been solved. Deeper tissues are routinely treated with lasers percutaneously (needles with fiber-optic tips are injected to the site to be treated), which works very well in deep brain tumors, liver, lung, etc. All tissues at all depths can, and are already being reached with lasers in a clinical setting. Here is a selection of references demonstrating this technique:
Thermal therapy of canine cerebral tumors using a 980 nm diode laser with MR temperature-sensitive imaging feedback. Kangasniemi M, McNichols RJ, Bankson JA, Gowda A, Price RE, Hazle JD. Lasers Surg Med. 2004;35(1):41-50. PMID: 15278927
Real-time magnetic resonance-guided laser thermal therapy for focal metastatic brain tumors. Carpentier A, McNichols RJ, Stafford RJ, Itzcovitz J, Guichard JP, Reizine D, Delaloge S, Vicaut E, Payen D, Gowda A, George B. Neurosurgery. 2008 Jul;63(1 Suppl 1):ONS21-8; discussion ONS28-9. PMID: 18728600
Feasibility study of particle-assisted laser ablation of brain tumors in orthotopic canine model. Schwartz JA, Shetty AM, Price RE, Stafford RJ, Wang JC, Uthamanthil RK, Pham K, McNichols RJ, Coleman CL, Payne JD. Cancer Res. 2009 Feb 15;69(4):1659-67. Epub 2009 Feb 10. PMID: 19208847
Laser-induced thermotherapy for lung tissue--evaluation of two different internally cooled application systems for clinical use. Ritz JP, Lehmann KS, Mols A, Frericks B, Knappe V, Buhr HJ, Holmer C. Lasers Med Sci. 2008 Apr;23(2):195-202. Epub 2007 Jun 29. PMID: 17599236
Furthermore, since the energy required to heat non-pigmented tissue is many orders of magnitude higher than that needed to destroy pigment granules, we will be able to greatly increase the laser power for deeper tissues, once overlying tissues have been safely cleared of pigment granules. The experiment I really want to do is treating two-inch thick steaks from old cows or pigs (which will contain lipofuscin granules throughout the cells) to find the maximum practical treatment depths without using needles. I had that in the proposal initially, but John thought it would be best to just deal with cell culture first.
(Btw the underlying assumption that a broken-up plaque = easily digestible plaque is also doubtful, but that is not a useful discussion to have at this stage imo).
Laser removal of tattoos, pigmented lesions (including age spots which are all lipofuscin), and lipofuscin-loaded RPE cells in macular degeneration all have shown that the granules, once disintegrated with the laser, are safely and efficiently cleared from the system. The body is set up to do this kind of thing already, and does it quite efficiently. I can provide more refs here if needed, but some in the proposal demonstrate this effect.
I'd like to see some details on the assessment. What software is used to evaluate lipofuscin destruction? What is the source of the "fibroblast cells"?
The quantization of granule number and size is easily done with a number of software packages. My favorite is the new quantization and analysis feature in Adobe Photoshop, which works quite nicely. Basically you take a number of sample images of treated and sham-treated cells, load them into Photoshop, and do your quantization analysis using the exact same settings for all images, to get an accurate comparison.
Right now we have WS1 (cat. # CRL-1502) and HFL1 (cat. # CCL-153) fibroblasts (originally from ATCC - a kind gift from my former lab). They are from human foreskin and lung, respectively.
COST: The prices quoted (those I can put into perspective, I don't know anything about lasers) are certainly very low. Can you really get a working cabinet at that price? What happened to the microsope in the second funding variant?
The scope is in the second variant too - it's just buried just above the 4-pulse setup equipment. Sorry for the poor formatting!
Yes, the prices are as low as we can get them we're scraping bottom here just to get this done with the bare minimum. The laser price is insanely cheap - thanks to the Chinese! Lasers made elsewhere are in the tens of thousands of dollars, but this one will do nicely for initial (and even later) experiments. We'll have to build the cabinet. John already built one using an upside-down rubbermaid tub and a HEPA filter, which works pretty well.
However, I think it is very difficult for the Institute to fund project-specific equipments grants, especially if they are of such basic nature. Maybe the laser needs to be bought but why can't one just use a friendly scientists lab? Imo, if you want to start a basic laboratory from scratch, the application should focus on that, rather than conflate it with a project application. The full costing would likely mean a significant contraction in Imminst finances. For the Institute to take such a big hit, a referendum and further discussion may be in order. Also, Imminst is not really in the business of funding 'big players' - we don't have enough money to make any difference in that area for starters. It may come as a surprise to the community that the SENS foundation is not able to fund such a -in the sceme of things- small project itself.
As far as I understand it, SENS funding is stretched to the limit as it is. We're in a fairly bad situation as John had to axe the funding for the entire cell culture facility in order to get the IBG budget down to what the foundation could fund, with the understanding that we would leech off of existing labs. For this project, if we get another lab involved then they have to be included in IP rights, so John wants to keep it all in-house to make sure it's patent-able and potential future investors will have a reason to fund clinical trials. I'm not sure about all the motivating factors, as I'm not involved in administrative decisions for IBG, but that is my understanding of the facts at this point.
Personnel: Costs are ok from our end, but Mr Schooler does not appear to have any cell culture experience? It would be good to see a CV. Will there be training and supervision? What does that mean: "The project will nevertheless begin 5/1/2009, lasting a total of 8 months"? If Imminst funding should come online in July, what happening in the interval? Note that if you want general IP on this idea (rather than on specific wavelegths, pluse combinations) you better file now.
I spent the last 2.5 years doing cell culture for my recent MS degree in Pharmacology and Toxicology from University of Louisville. I have extensive experience with the Fibroblasts we will be using, as well as a number of other cell types, as well as various viability assays, cell counting, colony formation, etc. Furthermore, I have 9 months worth of experience in Laser use, setup and alignment of optics, laser treatment of biological specimens, etc. Also, I spent the last 6 months selecting and purchasing equipment for, and setting up a Biology lab in Minnesota, so I have experience in this area as well. SENS funding can take up the slack for May and June, so I can start working full time right away. John tells me we need some good results before we apply for a patent, to convince investors to fund the patent application process - I'm not very knowledgeable about this end of things so I'm taking his word for it... He says it's too late for general IP since I've already given a presentation on the technique at UABBA.
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