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FOXO4-DRI or Proxofim is a research peptide being used for education and research uses only. The foxo4-DRI human trials are finding it to be effective in reducing aging!
What is the FOXO4-DRI Experience?
The research peptide FOXO4 is a cell-penetrating peptide shown to selectively induce apoptosis of senescent cells thereby reversing the effects of aging in mice.
The FOXO4-M peptide is a modified version of the already studied FOXO4DRI, a cell-penetrating peptide (CPP) that has been shown to selectively induce apoptosis of senescent cells in mice. However, the cell-penetrating peptide DRI is toxic for the cells in the brain of rats and the salt form has to be administered in the free form. In an attempt to overcome this limitation, we have developed a modified version of DRI, named FOXO4-M, in which we have removed the free salt form, and the modified version is administered in the form of its amino acid residue version.
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We show that FOXO4-M is not toxic for the cells in the brain, contrary to the already studied FOXO4DRI. Moreover, FOXO4-M penetrates cells and the whole mouse body. In an in vitro senescence assay, FOXO4-M induces senescent cells to revert the senescence phenotype and revert the effect of aging. Importantly, FOXO4-M decreases the expression of several aging-associated proteins in the brain in vivo, and, in agreement with this finding, FOXO4-M increases life span in mice.
FOXO4-DRI human trials
In a previous study we have identified DRI, a peptide composed of 21 amino acids (AA) with a cell-penetrating capacity in vitro, which induces apoptosis of senescent cells and reverses the effect of aging in mice. Here, we describe FOXO4DRI, a novel chemically synthesized version of DRI (18AA) that is devoid of the salt form and that penetrates cells in vitro. Moreover, the peptide does not induce oxidative stress in vitro in contrast to DRI. Here, we report on the in vivo activity of the peptide, which induces apoptosis of senescent cells and reverses the effects of aging in mice.
We present FOXO4DRI, a new DRI isoform that is devoid of the salt form and is not toxic to cultured cells.
What is the FOXO4 DRI sequence?
In our study, we identified a peptide, FOXO4DRI, that induces apoptosis of senescent cells in vitro and reduces expression of proteins associated with aging in mice, including p16INK4A and p21WAF1. Importantly, FOXO4DRI decreased the lifespan of mice, an effect that could be reversed by co-administration of a synthetic antioxidant compound.
In a previous study, we identified a peptide (DRI, 1) consisting of 21 AAs, which is able to selectively induce apoptosis of senescent cells in vitro. Moreover, in vivo treatment with DRI restores the level of proliferation and reverses the effect of aging in mice. DRI acts by penetrating cells in culture and it is not toxic for the cells in the brain. Notably, DRI is an unstructured peptide, and salt formation is essential for its activity in vivo.
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In the study described above, the DRI-induced reversal of effects of aging in mice was achieved by administrating the peptide in the free form. However, the free peptide forms aggregates when injected, which is required for its penetration into cells and the whole body. These aggregates are toxic for the cells and are difficult to control. Therefore, we aimed to develop a modified version of DRI (1), namely DRI.
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Our aim was to remove the salt form of the peptide by modifying it with a hydrophobic AA at the C-terminus. Additionally, we deleted the 3 AA (A-L) from the N-terminus in the following version. Finally, we decided to use the CPP, Pep17, and, as the final product, we synthesized FOXO4DRI-A.
To investigate whether FOXO4DRI-A could enter cells in vitro and induce senescence, we transfected a human embryonic kidney cell line (HEK 293) with a plasmid expressing FOXO4 fused with EGFP. The cells were treated with various concentrations of FOXO4DRI-A, and senescence was assessed using the senescence β-galactosidase assay. FOXO4DRI-A-induced senescence of the cells in a dose-dependent manner. At 100 μM concentration of FOXO4DRI-A, the induction of senescence was significant. We then transfected the cells with a plasmid expressing and quantified the expression of p21 using immunoblot.
As expected, p21 was decreased by FOXO4DRI-A at the same concentration (100 μM) that induced senescence in the HEK 293 cells. These findings indicate that FOXO4DRI-A penetrates the cells and induces senescence.
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In our study, we have identified a novel peptide, named FOXO4DRI, which penetrates the cells and induces senescence of the cells in culture, and reverses the effect of aging in mice. In the present study, we modified the peptide and developed a novel peptide, FOXO4DRI-A, which has similar activity as the peptide.
