Sciencemadness Discussion Board - Targeted Apoptosis of Senescent Cells

Targeted Apoptosis of Senescent Cells

Jekyll - 24-3-2017 at 21:11

I may have found my next synthesis target after decades away from the lab.

Unfortunately, my skills aren't up to it. Should I dust off my chemistry degree from last millennium and see if I can get into the local Masters program in biochemistry and molecular biology? Hire some starving grad student and build her a lab? Outsource to a custom biosynthesis company? Or try the Science Madness approach and start learning and building myself a lab? Any thoughts or advice would be greatly appreciated.

Targeted Apoptosis of Senescent Cells Restores Tissue Homeostasis in Response to Chemotoxicity and Aging
SUMMARY
The accumulation of irreparable cellular damage restricts healthspan after acute stress or natural aging. Senescent cells are thought to impair tissue function, and their genetic clearance can delay features of aging. Identifying how senescent cells avoid apoptosis allows for the prospective design of anti-senescence compounds to address whether homeostasis can also be restored. Here, we identify FOXO4 as a pivot in senescent cell viability. We designed a FOXO4 peptide that perturbs the FOXO4 interaction with p53. In senescent cells, this selectively causes p53 nuclear exclusion and cell-intrinsic apoptosis. Under conditions where it was well tolerated in vivo, this FOXO4 peptide neutralized doxorubicin-induced chemotoxicity. Moreover, it restored fitness, fur density, and renal function in both fast aging XpdTTD/TTD and naturally aged mice. Thus, therapeutic targeting of senescent cells is feasible under conditions where loss of health has already occurred, and in doing so tissue homeostasis can effectively be restored.
Baar et al., 2017, Cell 169, 132–147 March 23, 2017

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JJay - 25-3-2017 at 15:36

Is it a carcinogen?

Metacelsus - 25-3-2017 at 15:46

I wouldn't be surprised if it was. Senescent cells that had p53 inactivated and failed to undergo apoptosis could quite likely cause cancer, depending on where and what kind of DNA damage they had.

[Edit: this is probably wrong; see post below]

[Edited on 3-26-2017 by Metacelsus]

phlogiston - 25-3-2017 at 17:30

After a cursory reading of the paper I think that is not what happens. They have developed a peptide that induces apoptosis in senescent cells.

It works by interfering with the interaction between FOXO4 and p53. It can enter the cells by means of a TAT domain.
The peptide is called "FOXO4 D-Retro-Inverso" (FOXO4-DRI) in the paper. The materials and methods has this to say about it:

Quote:

FOXO4 D-Retro-Inverso peptide development
FOXO4-DRI consists of the following amino acid sequence in D-Isoform:

H-ltlrkepaseiaqsileaysqngwanrrsggkrppprrrqrrkkrg-OH. MW: 5358.2 It was manufactured by Pepscan (Lelystad, the Netherlands) at > 95% purity and stored at −20°C in 1mg powder aliquots until used to avoid freeze-thawing artifacts. For in vitro experiments FOXO4-DRI was dissolved in PBS to generate a 2mM stock. For in vivo use, FOXO4-DRI was dissolved in PBS to generate a 5mg/ml stock solution, which was kept on ice until injection. Before injection the solution was brought to room temperature.

After a cursory reading of the paper I think that is not what happens. They have developed a peptide that induces apoptosis in senescent cells.

It works by interfering with the interaction between FOXO4 and p53. It can enter the cells by means of a TAT domain.
The peptide is called "FOXO4 D-Retro-Inverso" (FOXO4-DRI) in the paper. The materials and methods has this to say about it:

Quote:

So, you are after a peptide
Wasn't someone recently offering PyBOP on this forum?

[Edited on 26-3-2017 by phlogiston]

Metacelsus - 25-3-2017 at 19:55

You're right, I had the mechanism of action backwards. The drug inhibits FOXO4, causing activation of p53, not the other way around. This would make it unlikely to cause cancer.

I think the peptide is probably too long to synthesize as an amateur. You'd need an automated machine, and then HPLC to purify the product. I've made a hexapeptide by manual SPPS (as part of a class), and in that case my yield was only 63% after purification. Besides, you would need D-amino acids, which are harder to obtain.

[Edited on 3-26-2017 by Metacelsus]

Tsjerk - 26-3-2017 at 03:57

That is not just a peptide, I would call that a small protein. Maybe get some E. coli to make that would be easier.

JJay - 26-3-2017 at 13:42

If it can actually safely cure hair loss, it could be worth a fortune.

phlogiston - 26-3-2017 at 14:49

Tsjerk, note that the peptide/small protein is composed of D-amino acids.
As I am sure you know, E coli ribosomal protein synthesis would afford a peptide composed of L-amino acids.

It would a cool if it were possible someday to make an entire organism using the other stereoisomers than used by earth life, ie. D-amino acids, L-sugars, L-DNA, etc.
I've seen companies that can synthesize moderately large enzymes in an active form from D-amino acids. In fact, I'l see if I can dig up their name as they would probably be well suited to make the FOXO4-DRI peptide.

E coli, like many species of bacteria, does use certain D-amino acids (D-Ala and D-Glu) btw, mainly for its peptideglycan-based cell wall, but those peptides are not made by ribosomes, but by a dedicated pathway for synthesizing the components that make up the cell wall.

Tsjerk - 27-3-2017 at 09:01

Ah, I didn't notice that in my very quick scan. But ofcourse that is not going to work in E. coli indeed

Jekyll - 27-3-2017 at 13:00

Quote: Originally posted by phlogiston

I've seen companies that can synthesize moderately large enzymes in an active form from D-amino acids. In fact, I'l see if I can dig up their name as they would probably be well suited to make the FOXO4-DRI peptide.

Thanks much, I would greatly appreciate that!