Re: [DIYbio] Re: Intein control in a targeted cancer therapy

It's great to have ideas and write them up for others to see and comment on. I assume you want people to comment on your manuscript as that is why you posted it so I will attempt to give you a critique.

On first writing a scientific paper I would suggest doing a thorough literature search to uncover what people have done before. I also would suggest reading current literature. 32 Citations is scant for a theory paper and only ~5 of them are have been published in the last 3 years(all in 2010). This has no general bearing on the quality of a paper but is definitely a red flag because alot happens in Science in 3 years.

What you are proposing is not worded clearly but let me try. You want to create a T-cell that has an extracellular protein that will bind a cancer cell and then cause the uptake of a active caspase to kill the cell? All of that is daunting. One cannot just do this so easy.

Similar attempts have been tried by Adam Arkins group:

Anderson, J.C., Clarke, E.J., Arkin, A.P. and Voigt, C.A. (2006) Environmentally Controlled Invasion of Cancer Cells by Engineered Bacteria. J Mol Biol, 355, 619-627.

I just saw a talk by him recently. The bacteria kill the host even when having layers of specificity to target cancer cells. So this is no easy task.

Induction of apoptosis of cancer cells my using a monoclonal antibody to EGF to activate caspase, been done:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2363248/

Synthetic antibody work by Koide, Sidhu, Kossiakoff

Specifically targeting cancer cells, so many papers on that:
http://scholar.google.com/scholar?hl=en&as_sdt=0,14&q=specific+targeting+cancer+cells

Including some new cool work by Kobi Benenson
http://www.sciencemag.org/content/333/6047/1307.short

Protein design by David Baker and many others

Sometimes the problem why we don't see an accomplishment in Science is not that people don't have good ideas. The problem is that everything people have tried has not worked. Sure, people can have a treatment target a cancer cell in culture and have a protein taken up and activated. That's not real life. It is far from it.

Some statements you make in the work are completely wrong "However, with regard to oncogenes, the research suggests it isn't the function of the protein product that is necessarily altered in an oncogenic mutation, but rather a change in transcription regulation". Even the papers you cite disagree with this! Go here for a nice summary and tons of citations: http://en.wikipedia.org/wiki/Oncogene

"The interaction of forces at the atomic and quantum level that are the fabric in which proteins fold are not well understood. It is for this reason that, just as we confirm the very existence of proteins, an indirect path must be followed to engineer them."

Protein folding is very well understood. Read works by Firsht, Sosnick, Baker, DE Shaw.
If we don't understand the forces how can Scientists run simulations that recapitulate folding experiments to a high degree:
http://www.sciencemag.org/content/334/6055/517.short

Sure we don't understand everything about protein folding and have lots of trouble folding multi-domain proteins but "not well understood" is completely false. More like "not an easy way to calculate". Calculating changes in nuclei polarization or QM/MM requires so much computational power. Our understanding is great, it is the computation that is having a hard time keeping up.

"Marc Ostermeier wrote in 2009 given that allosteric effects depend on the "protein's sequence,structure, and energetics" one might reasonably conclude that computational modeling would be required to engineer the same. However, it has been rational design strategies that have proven most often successful. This is due, he continues, to the "modularity of protein function""

Rational design strategies have not proven the most successful. I work on designing new protein function. Computational modelling is so far beyond rational design. That is like saying a human is better at maths than a computer. Most every major modern protein design strategy that has worked has been done with a computer.
http://www.sciencemag.org/content/329/5989/309.short
http://www.sciencemag.org/content/302/5649/1364.short
http://www.sciencemag.org/content/319/5868/1387.short

"Upon encountering its tumor-specific ligand the resultant binding will be a conformational change that induces the intein self-excision and extein ligation. "
I feel like I need a meme here so:
http://cdn0.meme.li/instances/300x300/39360153.jpg

Inducing conformational change is one of the most complicated things in protein engineering. It is a huge goal of most protein engineers. The interaction networks that induce conformational change are so complicated and multi-noded. This in itself would be a huge feat much less all the other aims in the writing.

Many many other things I will not touch on for the sake of brevity as I am sure you are understanding my point.

Lots and lots of writing is uncited and aims are proposed as if they can just be done easily. Your goal in a writing such as this is that unless a statement made is an undeniable truth have a citation. READ the citations also. It is very apparent that you perhaps only read the title or abstract of many of these papers or chose to ignore much of what is written in the papers.

Good, to try your hand at some scientific theory work but I would recommend staying away from experimental theory unless you can test the experiments. One is so far removed from the experimental process that they have a hard time coming to terms with what it really takes to perform an experiment.

With your motivation I would suggest investing more in computational theory, thermodynamics theory or physical chemistry. You seem interested in it from the writing and it is an area that in many cases one can directly compare their work to experiment to validate it.

Hopefully this doesn't discourage you to stop learning and instead encourages you to to learn more and think deeply and Scientifically minded about things you propose.

Remember 90% of experimental planning is a good literature search and reading.


On Monday, July 1, 2013 11:07:36 PM UTC-5, Nathan McCorkle wrote:

aka http://academia.edu/1079278/intracellular_Caspase-Modulating_Chimeric_Antigen_Receptor

On Mon, Jul 1, 2013 at 7:44 PM, InteinHereAndThere
<sta...@webmeetssight.com> wrote:
>
> Thank you...
>
> My paper can be read here:  http://tinyurl.com/9kdbbqb
>
>
>
>
>
> On Monday, July 1, 2013 4:47:47 PM UTC-4, Mackenzie Cowell wrote:
>>
>> Passing your message on to the main list, Scott.
>>
>> ---------- Forwarded message ----------
>> From: Scott Tarone <sta...@synapticsynthase.com>
>> Date: Sun, Jun 30, 2013 at 7:16 PM
>> Subject: Intein control in a targeted cancer therapy
>> To: con...@diybio.org
>>
>>
>> Hello,
>>
>>
>>
>> I'm not sure if your group is the right place for me to look for help, but
>> I thought it worth a shot.
>>
>>
>>
>> I am self-taught in the biosciences… Neuroscience, Genetics, Immunology,
>> and Targeted Cancer Therapies.  After watching a presentation on Chimeric
>> Antigen Receptors on the NIH site, I had an idea on another approach that
>> would remove the limitations inherent in the technique presented (though it
>> was a fantastic step forward, and very successful).  My approach is based on
>> a constitutively active Caspase 3 mutant controlled by a molecular switch.
>>
>>
>>
>> Long story short… I just wrote up a few paragraphs on my idea and sent it
>> off to a few people in the field.  One of them (a senior scientist from
>> Pfizer) got back to me a few months later and said I may be on to something
>> and that I needed to write a paper.  I did  that and then wrote an article
>> on that paper with him and that was published in a small time online journal
>> last year.  Since then my paper and the article have been download several
>> hundred times.
>>
>>
>>
>> I didn't provide the links or an attachment as my first attempt to send
>> this email was blocked by spam filters.. I will provide that information if
>> you have interest.  In the interim, you can search for "intracellular
>> Caspase modulating Chimeric Antigen Receptor".
>>
>>
>>
>> Thank you very much for your time…
>>
>>
>>
>> Take care,
>>
>> Scott
>>
>>
>>
>>
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--
-Nathan

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