Inhibition of cancer cell growth and c-Myc transcriptional activity by a c-Myc helix 1-type peptide fused to an internalization sequence.
c-Myc is a nuclear protein with important roles in cell transformation, cell proliferation, and gene transcription. It has been previously shown that a 14-amino acid (aa) modified peptide (H1-S6A,F8A) derived from the helix 1 (H1) carboxylic region of c-Myc can interfere in vitro with specific c-Myc DNA binding. Here, we have linked the above Myc-derived 14-aa peptide to a 16-aa sequence from the third helix of Antennapedia (Int). It has been repeatedly reported that this 16-aa Antennapedia peptide is able to cross mammalian cell membranes and to work as a vector for short peptides. Using fluorescent (dansylated or rhodaminated) peptides, we have shown that the fusion peptide with the Antennapedia fragment (Int-H1-S6A,F8A) but not the c-Myc derived fragment alone (H1-S6A,F8A) was capable of internalization inside MCF-7 human breast cancer cells. Int-H1-S6A,F8A and H1-S6A,F8A were the only two peptides capable of inhibiting coimmunoprecipitation of the c-Myc/Max heterodimer in vitro. We have treated (continuously for 10-11 days) MCF-7 cells with four different peptides: Int, H1-S6A,F8A, Int-H1-S6A,F8A, and Int-H1wt [a peptide differing from Int-H1-S6A,F8A by 2 aa (S6 and F8) in the H1 region]. In intact MCF-7 cells, Int-H1-S6A,F8A was the only active peptide capable of inducing the following biological effects: (a) inhibition of cloning efficiency on plates; (b) inhibition of cell growth and induction of apoptosis in subconfluent/confluent cells; and (c) inhibition of transcription of two c-Myc-regulated genes (ODC and p53). Int-H1-S6A,F8A was active in the 1-10 microM range. Int-H1-S6A,F8A may represent a lead molecule for peptidomimetic compounds that have a similar three-dimensional structure but are more resistant to peptidases and, therefore, suitable for in vivo treatment of experimentally induced tumors.
Giorello
L
L
Department of Experimental Oncology, Istituto Nazionale per la Ricerca sul Cancro, Genoa, Italy.
Clerico
L
L
Pescarolo
M P
MP
Vikhanskaya
F
F
Salmona
M
M
Colella
G
G
Bruno
S
S
Mancuso
T
T
Bagnasco
L
L
Russo
P
P
Parodi
S
S
D000595:Amino Acid Sequence; D051943:Antennapedia Homeodomain Protein; D017209:Apoptosis; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D050976:Basic-Leucine Zipper Transcription Factors; D002448:Cell Adhesion / Q000187:drug effects; D002452:Cell Count; D002455:Cell Division / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000187:drug effects / Q000378:metabolism; D016259:Genes, myc / Q000187:drug effects; D018398:Homeodomain Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D016271:Proto-Oncogene Proteins c-myc / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D013997:Time Factors; D014157:Transcription Factors; D014407:Tumor Cells, Cultured / Q000378:metabolism; D014410:Tumor Stem Cell Assay
D051943:Antennapedia Homeodomain Protein; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; C499330:MAX protein, human; C067850:Myc associated factor X; D009687:Nuclear Proteins; D010446:Peptide Fragments; D016271:Proto-Oncogene Proteins c-myc; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors