SELECTIVE INHIBITORS OF C-FOS AND THEIR ANTIPROLIFERATIVE PROPERTIES

Abstract

Selective inhibitor of c-Fos and their antiproliferative properties The invention relates to selective inhibitor of c-Fos for use in the prevention and/or treatment of cancers and restenosis.

Claims

1. A selective inhibitor of c-Fos for use in the prevention and/or treatment of a cancer caused by or involving a mutation in the Ras/ERK pathway; and/or associated with an increased production of c-Fos; wherein said selective inhibitor of c-Fos is a peptide comprising: at least one cell penetrating sequence; and an amino acid sequence corresponding to a docking domain sequence of c-Fos comprising SEQ ID NO: 29 (FVFTYPEA).

2. The selective inhibitor of c-Fos for use according to claim 1, wherein said cancer caused by or involving a mutation in the Ras/ERK pathway is caused by or involves a mutation of Raf or Ras.

3. The selective inhibitor of c-Fos for use according to claim 2, wherein said cancer is selected in the group consisting of colon cancer, pancreatic cancer, melanoma, thyroid cancer, lung cancer and leukaemia, and ovary cancer.

4. The inhibitor peptide of c-Fos for use according to claim 1, wherein said cancer caused by or involving a mutation in the Ras/ERK pathway is caused by or involves a mutation of NF1.

5. The selective inhibitor of c-Fos for use according to claim 4, wherein said cancer is selected in the group consisting of glioma, juvenile myelomonocytic leukaemia and neurofibroma.

6. The inhibitor peptide of c-Fos for use according to claim 1, wherein said cancer associated with an increased production of c-Fos is selected in the group consisting of cervical hepatocarcinoma, pancreatic cancer, breast cancer, osteosarcoma, and endometrial cancer.

7. A selective inhibitor of c-Fos for use in a method of preventing metastasis, wherein said selective inhibitor of c-Fos is a peptide comprising: at least one cell penetrating sequence; and an amino acid sequence corresponding to a docking domain sequence of c-Fos comprising SEQ ID NO: 29 (FVFTYPEA).

8. A selective inhibitor of c-Fos for use for inhibiting and/or preventing proliferation of vascular smooth muscle cells on a stent, wherein said selective inhibitor of c-Fos is a peptide comprising: at least one cell penetrating sequence; and an amino acid sequence corresponding to a docking domain sequence of c-Fos comprising SEQ ID NO: 29 (FVFTYPEA); wherein said stent is used for treating a patient suffering from cardiovascular disease.

9. A selective inhibitor of c-Fos for use according to claim 8 wherein said stent is a drug-eluting stent.

10. A selective inhibitor of c-Fos for use in the prevention and/or treatment of neurofibromatosis, preferably in the prevention and/or treatment of peripheral nerve sheath tumors, wherein said selective inhibitor of c-Fos comprises at least one cell penetrating sequence, and an amino acid sequence corresponding to a docking domain sequence of c-Fos comprising SEQ ID NO: 29 (FVFTYPEA).

11. The selective inhibitor of c-Fos for use according to claim 1, wherein said amino acid sequence corresponding to a docking domain sequence of c-Fos is selected in the group consisting of: TABLE-US-00006 SEQIDNO:30 (SFVFTYPEAD), SEQIDNO:31 (SSFVFTYPEADS), SEQIDNO:32 (TSSFVFTYPEADSF), SEQIDNO:33 (YTSSFVFTYPEADSFP), SEQIDNO:34 (TYTSSFVFTYPEADSFP), SEQIDNO:35 (TTYTSSFVFTYPEADSFP), SEQIDNO:1 (CTTYTSSFVFTYPEADSFP), and SEQIDNO:39 (CTTYTSSFVFTYPEADSFPS).

12. The selective inhibitor of c-Fos for use according to claim 1, wherein said amino acid sequence corresponding to a docking domain sequence of c-Fos is SEQ ID NO: 39 (CTT YTS SF VFT YPEAD SFPS).

13. The selective inhibitor of c-Fos for use according to claim 1, wherein said cell penetrating sequence is chosen in the group consisting of: HIV-TAT sequence (SEQ ID NO: 2); Penetratin (SEQ ID NO: 3); an amino acid sequence of 7 to 11 arginine (SEQ ID NO: 4 to 8); a X7/11R sequence of 7 to 25 amino acids comprising 7 to 11 arginine randomly positioned in the sequence such as SEQ ID NO: 9 to 12; and a sequence derived from DPVs (SEQ ID NO: 13 to 17).

14. The selective inhibitor of c-Fos for use according to claim 1, wherein said peptide has the sequence SEQ ID NO: 26 (GRKKRRQRRRPPCTTYTSSFVFTYPEADSFP) or SEQ ID NO: 36 (GRKKRRQRRRPPCTTYTSSFVFTYPEADSFPS).

15. A pharmaceutical composition for use for preventing and/or treating: a cancer selected in the group consisting of colon cancer, pancreatic cancer, melanoma, ovary cancer, lung cancer, thyroid cancer, leukaemia, juvenile myelomonocytic leukaemia, glioma, neurofibroma, cervical hepatocarcinoma, breast cancer, osteosarcoma and endometrial cancer; and/or neurofibromatosis, preferably neurofibromatosis I, wherein, said pharmaceutical composition comprises: a) at least one selective inhibitor of c-Fos as defined in claim 1; b) a nucleic acid encoding said peptide; or c) an expression vector comprising said nucleic acid.

Description

FIGURES

[0170] FIG. 1: Schema summarizing the Ras-ERK signalling pathway and mutations in human cancers.

[0171] FIG. 2: Concept of the Pepsignal.

[0172] Panel A depicts the mechanism of action of classical MEK inhibitors used so far. They act upstream of ERK and block the activation of all ERK substrates without discrimination.

[0173] Panel B illustrates the particularity of the pepsignal that are the only bio-molecule acting downstream from ERK. They mimic the DEF docking domain of a given substrate towards ERK, and as such have a very specific inhibitory impact of a given substrate of interest.

[0174] FIG. 3: The peptide of the invention TAT-DEF-c-Fos, blocks serum-induced proliferation of vSMC cells whereas the peptide TAT-DEF-Elk-1 does not.

[0175] On day one, 10 000 cells per well were grown in the presence of 10% serum. After one cycle of cell division (around 32 hours), the cells were starved (0.1% serum), then they were grown in 10% FBS (serum) in the presence or not of either the TAT-DEF-Elk-1, the TAT-DEF-c-Fos or classical inhibitors of cell proliferation (U0126, an ERK inhibitor; VIVIT or Paclitaxel) in the presence of BrdU for 32 h to label dividing cells. On the bases of our previous results in neuronal cells (see Lavaur et al., 2007) three different doses of pepsignal were tested: from 3 to 12 M. Note that the TAT-DEF-c-Fos but not TAT-DEF-Elk-1 is efficient in the inhibition of vSMC proliferation; n=3; One-way ANOVA; Dunett post hoc test; ** p<0.01; *** p<0.001 when compared to the group treated with 10% serum

[0176] FIG. 4: Efficacy of the peptide of the invention TAT-DEF-c-Fos on the inhibition of serum-induced proliferation of vSMC cells: comparison of single and multiple applications.

[0177] Using the same protocol as in FIG. 3, cells were grown in the presence of 10% FBS (serum) after withdrawal. The peptide TAT-DEF-c-Fos was applied once (6 or 12 M) on day one (first day after withdrawal) and proliferation was tested using a BrdU assay 24, 48 or 72 hours later. When indicated, multiple applications of the peptide were done (m) every 24 hours. Note that the TAT-DEF-c-Fos12 M is not reported herein; n=3; Two-way ANOVA; Bonferroni post hoc test; *** p<0.001 when compared to the group treated with 10% serum.

[0178] FIG. 5: TAT-DEF-c-Fos is devoid of anti-proliferative properties on HCAEC.

[0179] Using the same protocol than in FIG. 3, cells were grown in the presence of 10% serum after withdrawal. The peptide TAT-DEF-c-Fos was applied at a single dose one day after withdrawal. Proliferation was tested using a BrdU assay; n=3; One-way ANOVA; Dunett post hoc test; * p<0.05; when compared to the group treated with 10% serum

[0180] FIG. 6: Antiproliferative properties of TAT-DEF-c-Fos on NIH3T3 cells

[0181] On day one, 10 000 cells per well were grown in the presence of 10% serum. After one cycle of cell division (around 22 hours), the cells were starved (0.1% serum), then they were grown in 10% serum in the presence or not of the TAT-DEF-c-Fos (1,3,6, or 12 M) or classical inhibitors of cell proliferation (U0126, an ERK inhibitor; VIVIT or Paclitaxel) in the presence of BrdU for 3 h to labelled dividing cells. Bars are the mean % of BrdU positive cells obtained from two experiments performed with 2 different stainings of NIH3T3 cells.

[0182] FIG. 7: TAT-DEF-C-Fos toxicity on vSMC

[0183] Cell viability was tested using an Hoechst staining at the end of the indicated time after 10% serum application. A) the peptide was applied at 3; 6 or 12 M. the cells were fixed 24 hours after the serum application. B) The cells were fixed 24, 48 or 72 hours after serum application. When indicated, multiple applications of the peptide were done (m) every 24 hours. Note that the TAT-DEF-c-Fos12m is not reported, as it was toxic at this dose after multiple applications. The number of Hoechst positive cells was counted after acquisition of the image and automatized counting was performed, using a dedicated software (Image Pro). Statistics: Two way ANOVA followed by a bonferroni test *p<0.05; **p<0.01; ***p<0.001 when compared to 24 hours

[0184] FIG. 8: Inhibitory role of TAT-DEF-c-Fos peptides in the wound-healing test in vSMC.

[0185] The peptide was applied once, at the same time as 10% serum. The invasiveness was calculated as the percentage of cells over the size of the lesion, 24, 48 and 72 hours post-lesion. Note the lack of effect of the global MEK inhibitor (U0126), note also the inhibitory role of the TAT-DEF-c-Fos peptide at 12 M whatever the time point chosen.

[0186] FIG. 9: Comparative effects TAT-DEF-c-Fos and Rapamycin on MPNST from NF1 patients

[0187] Human cell lines from malignant peripheral nerve sheat tumors (MPNST) were treated with either TAT-DEF-c-Fos at increasing doses, or as a control, with Rapamycin, an inhibitor of the mTOR pathway that is classically used as an inhibitor of proliferation on this cell line. The TAT-DEF-c-Fos peptide inhibited proliferation of MPNST at very low doses when compared to Rapamycin (IC50 for TAT-DEF-c-Fos around 10 M instead of 10 mM for Rapamycin). This indicates that the peptide is highly efficient on the inhibition of proliferation on this malignant cell line.

[0188] FIG. 10: Inhibitory effects of 4 different peptides on sNF96.2

[0189] 4 different peptides are tested on malignant Peripheral Nerve Sheath Tumors (MPNST) cell lines, as follows: [0190] Peptide TAT-DEF-c-Fos; [0191] Peptide TAT-DEF-Elk-1; [0192] Peptide TAT-DEF-JunB; and [0193] Peptide Penetratin-DEF-c-Fos.

EXAMPLES

Example 1

Anti-Proliferative Property of TAT-DEF-c-Fos Peptide on vSMC, MPNST from NF1 Patients and NIH3T3 Cell Lines

[0194] Methods

[0195] Cell Lines

[0196] Primary vascular smooth muscle cells (vSMC) (ATCC CRL-1999) were grown in DMEM F12K medium supplemented 0.05 mg/ml ascorbic acid; 0.01 mg/ml insulin; 0.01 mg/ml transferrin; 10 ng/ml sodium selenite; 10% (v/v) heat-inactivated FBS (Fetal Bovine Serum), 100 U/ml penicillin, and 100 g/ml streptomycin, 10 mM HEPES.

[0197] Primary human coronary arterial endothelial cells (HCAEC) (Promocell C12222) were grown in Endothelial Cell Growth Medium MV2 (Promocell).

[0198] NIH 3T3 mouse embryonic fibroblast cells were grown in DMEM supplemented with 10% (v/v) heat-inactivated FBS, 100 U/ml penicillin, and 100 g/ml streptomycin.

[0199] Malignant Peripheral Nerve Sheath Tumors (MPNST: from NF1 patients) were obtained from Michel Videaud's laboratory. 2000 cells were grown in the presence of 15% serum after withdrawal.

[0200] Cultures were maintained at 37 C. in humidified 95% air and 5% CO2

[0201] Pharmaceutical Compounds

[0202] The compounds used by the inventors are as follows: [0203] U0126 was from Tocris. Paclitaxel was from Millipore.

TABLE-US-00004 TAT-DEF-c-Fos: sequence (SEQIDNO:26) GRKKRRQRRRPPCTTYTSSFVFTYPEADSFP; and sequence (SEQIDNO:36) GRKKRRQRRRPPCTTYTSSFVFTYPEADSFPS; TAT-DEF-Elk-1: sequence (SEQIDNO:27) GRKKRRQRRRPPSPAKLSFQFPSSGSAQVHI; and VIVIT: sequence (SEQIDNO:28) MAGPHPVIVITGPHEE.

[0204] TAT-DEF-Elk-1 and TAT-DEF-c-FOS peptides were synthesized under their L conformation using Fmoc solid-phase peptide synthesizer by Genecust.

[0205] Peptides were purified by HPLC (>99%) and analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Stock solution (1 mM) were prepared in sterilized water and stored until use at 20 C.

[0206] Proliferation and Toxicity Assay

[0207] 10 000 cells per well were grown in the presence of complete medium. After one cycle of cell division (around 32 hours for HCAEC and vSMC, 24 hours for NIH3T3), the cells were starved for one other cycle (0.1% serum or nutriment supplement for HCAEC). Then they were grown in complete medium in the presence or not of each of the Pepsignal (3, 6 or 12 M), the MEK inhibitor U0126 (10 M), or classical inhibitors of cell proliferation Paclitaxel (10 nM), VIVIT (100 M) for 24 to 72 hours. Proliferation was analyzed after BrdU incorporation, which allows the visualization of cells under division. For HCAEC and vSMC, BrdU was added with inhibitor. For NIH3T3, it was added only during the last three hours. Cells were fixed in 4% paraformaldehyde.

[0208] Toxicity was analyzed by Hoechst staining after fixation, which allows the labeling of all nuclei, and hence their quantification. Only the nuclei with an intact nucleus were taken into account. Image-Pro Plus software (Media Cybernetics) was used in order to quantify BrdU positive cells. The percentage of positive cells was reported to the total number of Hoechst-positive cells.

[0209] Wound Healing Assay

[0210] Cells were cultured in 24-well plates as a confluent monolayer (30000 cells plated per well). The monolayer was starved in 0,1% serum (or nutriment supplement for HCAEC) for 24 hours and wounded in a line across the well with a 200-l pipette tip, then incubated with 10% serum (or nutriment supplement for HCAEC), in the presence or absence of U0126 (10 M), Paclitaxel (10 nM/l), VIVIT (100 M) or TAT-DEF-c-FOS (3,6,12 M) for 24 to 72 hours. Cells were fixed in 4% paraformaldehyde then stained with cresyl violet dye (1% in methanol). Pictures were taken to visualize the marked wound location. The wound healing effect was measured using the NIH ImageJ program and expressed as percentage of recovery of the lesion.

[0211] Results

[0212] 1) Anti-Proliferative Properties of TAT-DEF-c-Fos on vSMCs

[0213] The inventors first tested and compared the anti-proliferative properties of TAT-DEF-Elk-1, TAT-DEF-c-Fos, classical inhibitors of ERK, and classical inhibitors of cell proliferation on vSMC (FIG. 3). They used a classical protocol of cell proliferation induced by serum (10%) and BrdU incorporation. On the bases of our previous results in neuronal cells (see Lavaur et al., 2007) three different doses of pepsignal were tested: from 3 to 12 M. The inventors found that TAT-DEF-c-Fos (but not TAT-DEF-Elk-1) had anti-proliferative properties at 3, 6 and 12 M. Since results were more reproducible at the doses of 6 and 12 M, other experiments were carried on with 6 or 12 M. At 12 M, this anti-proliferative effect was comparable to classical inhibitors of ERK, or inhibitors of cell proliferation classically used on this cell line (FIG. 3c).

[0214] Having established that a unique application of the TAT-DEF-c-Fos was efficiently inhibiting the proliferation of vSMC, the inventors decided to determine its anti-proliferative properties after multiple cycles. In this experiment, single and multiple applications of the TAT-DEF-c-Fos peptide were tested. The inventors found that the TAT-DEF-c-Fos peptide was as efficient at single and multiple applications at 6 M for inhibiting cell proliferation (FIG. 4). At the dose of 12 M, one application of the peptide was necessary and sufficient to obtain rapidly (as soon as 24 hours) an inhibition of proliferation. At this dose, multiple applications were toxic.

[0215] Because anti-proliferative properties of a compound may be highly specific of a cell line and/or organ, depending on the extracellular stimuli and the environment, the inventors decided to test the efficacy of TAT-DEF-c-Fos on serum-induced proliferation of endothelial cells (HCAEC) (FIG. 5).

[0216] On this cell line, the classical inhibitors (VIVIT, Paclitaxel) of proliferation and ERK inhibitors (U0126) were efficient to inhibit HCAEC cell proliferation, but the TAT-DEF-c-Fos peptide was devoid of effect whatever the dose. This data thus indicates that the anti-proliferative properties of TAT-DEF-c-Fos are cell-specific.

[0217] 2) Anti-Proliferative Properties of TAT-DEF-c-Fos on Malignant NF1

[0218] The inventors tested the efficacy of TAT-DEF-c-Fos on malignant NF1 cell proliferation (FIG. 9). For this purpose, the inventors used Human cell lines from malignant peripheral nerve sheat tumors (MPNST) from NF1 patients.

[0219] The MPNST cell lines were derived from a recurrent mass associated with peripheral nerve sheat and diagnosed as MPNST (Malignant Peripheral Nerve Sheath Tumor) in patients meeting NF1 diagnostic criteria. The cells were derived from numerous passages of primary tumor material in culture until they were a homogenous Schwann cell-like population which displayed a clonal morphology immunopositive for both cytoplasmic Schwann cell markers S100 and p75.

[0220] As a control, the inventors chose Rapamycin, an inhibitor of the mTOR pathway that is classically used as an inhibitor of proliferation on this cell line. The TAT-DEF-c-Fos peptide inhibited proliferation of MPNST at very low doses when compared to Rapamycin (IC50 for TAT-DEF-c-Fos around 10 M instead of 10 mM for Rapamycin). This indicates that the peptide is highly efficient on the inhibition of proliferation on this malignant cell line.

[0221] 3) Anti-Proliferative Properties of TAT-DEF-c-Fos on Fibroblasts

[0222] Next the inventors tested the efficacy of TAT-DEF-c-Fos peptide on fibroblasts (NIH3T3 cells) (FIG. 6). On this cell line, the efficacy of the TAT-DEF-c-Fos peptide was found at low doses (from 1 to 12 M), while the ERK inhibitor U0126 failed to inhibit proliferation. This data indicates the specificity of ERK activity towards c-Fos on fibroblast proliferation.

[0223] 4) The TAT-DEF-c-Fos is Devoid of Toxicity

[0224] Classical inhibitors of proliferation, a MEK inhibitor or the TAT-DEF-c-Fos peptide at increasing doses were applied on vSMCs, in the presence of 10% serum, on a single application (FIG. 7A) and toxicity was evaluated after PFA fixation and Hoechst staining. Counting of viable cells was performed and expressed as number of Hoechst positive cells (showing nuclear labelling integrity). Contrasting with the classical inhibitor of cell proliferation on this cell line (VIVIT) that showed toxicity (decreased number of viable cells), no toxicity was observed in the presence of TAT-DEF-c-Fos at the doses used for inhibiting cell proliferation (from 3 to 12 M) (see FIG. 3B). The inventors also evaluated the toxicity of TAT-DEF-c-Fos after multiple applications (FIG. 7B). On this experiment, the peptide was applied every 24 hours in the presence of serum. The cells were fixed and stained with Hoechst at 24, 48 or 72 hours. At 6 M, the peptide did not alter the number of Hoechst positive cells whatever the number of applications. At 12 M, it showed toxicity after 48 and 72 hours.

[0225] 5) The TAT-DEF-c-Fos has Anti-Invasive Properties on vSMC

[0226] Invasive properties were studied using the wound healing protocol.

[0227] The percentage of lesion recovery was measured after cresyl violet staining of vSMC cells (FIG. 8). At 24 hours, the reference compound VIVIT showed a significant inhibition of invasiveness (around 30% recovery versus 78% in the presence of serum only). This inhibition was sustained since it was still significant after 78 hours of serum application (35% recovery versus 110% in the presence of serum only). However due to its toxic effect, this inhibition may be related to cell death.

[0228] Paclitaxel had a significant but transient effect on vSMC invasiveness. Whatever the doses used, the classical MEK inhibitor failed to block invasiveness. By contrast, the inventors found a significant effect of the TAT-DEF-c-Fos peptide at 12 M at the different time points (FIG. 8). Despite a tendency at 24 hours, the TAT-DEF-c-Fos peptide had no significant effect on cell invasiveness at 6 M.

[0229] Conclusion:

[0230] The TAT-DEF-c-Fos peptide has anti-proliferative properties on vSMC, PNMT and NIH3T3 cell lines.

[0231] Its efficacy is dose-dependent, but it becomes toxic at higher doses after multiple applications.

[0232] The TAT-DEF-c-Fos inhibits invasiveness.

[0233] The TAT-DEF-c-Fos peptide has specific properties when compared to TAT-DEF-Elk-1 or global MEK inhibitors.

Example 2

Comparative Properties of TAT-DEF-c-Fos, TAT-DEF-Elk-1, TAT-DEF-JunB, Penetratin-DEF-c-Fos on Proliferation of Malignant NF1 Cells (MPNST)

[0234] Method

[0235] The inventors have further tested 4 different peptides in the Peripheral Nerve Sheath Tumors (MPNST) cell lines sNF96.2, which was deposited to the ATCC under the reference ATCC CRL-2884.

[0236] The compounds used by the inventors are as follows:

TABLE-US-00005 PeptideTAT-DEF-c-Fos: (SEQIDNO:36) GRKKRRQRRRPPCTTYTSSFVFTYPEADSFPS; PeptideTAT-DEF-Elk-1: (SEQIDNO:27) GRKKRRQRRRPPSPAKLSFQFPSSGSAQVHI; PeptideTAT-DEF-JunB: (SEQIDNO:37) GRKKRRQRRRPPTTPTPPGQYFYPRGGGSGGGAG; PeptidePenetratin-DEF-c-Fos: (SEQIDNO:38) RQIKIWFWNRRMKWKKPPCTTYTSSFVFTYPEADSFPS.

[0237] Cells were cultured at DayO in a complete medium (DMEM with 10% of fetal bovine serum). Then they were deprived at Day2 (24 hours in a medium comprising DMEM with 0.1% fetal bovine serum followed by 24 hours in DMEM alone).

[0238] At Day 4, the cells were incubated with a dose escalation regimen of pepsignal peptides from 2 to 100 M.

[0239] The proliferation tests were carried out 96 hours later.

[0240] Proliferation was analysed by a MTT assay. The MTT assay is a colorimetric assay for assessing cell viability. The viability is measured by the metabolic activity if the cells, which is determined by the activity of a mitochondrial succinate dehydrogenate. Typically, the cell media goes from yellow to blue and the intensity of the coloration is proportional to the number of viable cells. Coloration is determined by atomic absorption spectrometry at 570 nm.

[0241] Results

[0242] The inventors performed three independent experiments, each point being performed in duplicate for each experiment.

[0243] The results are illustrated on FIG. 10 and summarized as follows:

[0244] The IC50 of TAT-DEF-c-Fos is approximatively of 10 M (FIG. 10). At a concentration of 100 M, the peptide shows total inhibition.

[0245] The peptide Penetratin-DEF-c-Fos shows anti proliferative properties on MPNST (FIG. 10), with however lower efficacy than the TAT-DEF-c-Fos peptide (approximatively 66% of inhibition at a concentration of 100 M).

[0246] The peptide TAT-DEF-JunB also shows anti proliferative properties, with lower efficacy than the TAT-DEF-c-Fos peptide.

[0247] Finally, the peptide TAT-DEF-Elk1 has low anti-proliferative properties (FIG. 10). The inhibition of proliferation does not reach 50%, even at higher doses.

[0248] Conclusion

[0249] In conclusion, the peptides TAT-DEF-c-Fos and Penetratin-DEF-c-Fos both show an inhibitory effect on MPNST. However, TAT-DEF-c-Fos shows higher efficacy.

[0250] In addition, TAT-DEF-Junb and TAT-DEF-Elk-1 shows effects, which are well below the effects conveyed by TAT-DEF-c-Fos.

[0251] Finally, the results of the inventors show that a peptide bearing a TAT sequence