A membrane penetrating peptide aptamer inhibits STAT3 function and suppresses the growth of STAT3 addicted tumor cells

Abstract

Cancer cells are characterized by the aberrant activation of signaling pathways governing proliferation, survival, angiogenesis, migration and immune evasion. These processes are partially regulated by the transcription factor STAT3. This factor is inappropriately activated in diverse tumor types. Since tumor cells can become dependent on its persistent activation, STAT3 is a favorable drug target. Here, we describe the functional characterization of the recombinant STAT3 inhibitor, rS3-PA. This inhibitor is based on a 20 amino acid peptide which specifically interacts with the dimerization domain of STAT3. It is integrated into a thioredoxin scaffold and fused to a protein transduction domain. Protein gel blot and immunofluorescence analyses showed that rS3-PA is efficiently taken up by cells via an endocytosis independent mechanism. Intracellularly, it reduces the phosphorylation of STAT3 and enhances its degradation. This leads to the downregulation of STAT3 target gene expression on the mRNA and protein levels. Subsequently, tumor cell proliferation, survival and migration and the induction of angiogenesis are inhibited. In contrast, normal cells remain unaffected. Systemic administration of rS3-PA at doses of 7.5 mg/kg reduced P-STAT3 levels and significantly inhibited tumor growth up to 35% in a glioblastoma xenograft mouse model.

The Inhibition of Stat5 by a Peptide Aptamer Ligand Specific for the DNA Binding Domain Prevents Target Gene Transactivation and the Growth of Breast and Prostate Tumor Cells

“…Recombinant rS3-PA is rapidly taken up by cultured cells upon addition to the growth medium and intracellularly interacts with its target domain. It causes the inhibition of Stat3 activation and dimerisation and reduces the viability and the growth of Stat3-dependent tumor cells [48]. The binding of S5-DBD-PA to the Stat5-DBD is expected to block the function of Stat5 and elicit similar phenotypes as rS3-PA (Figure 3b).…”

“…We do not a priori expect that the binding of S5-DBD-PA to the Stat5-DBD should interfere with the phosphorylation of Stat5. This is a consequence we can attribute to the Stat3-specific inhibitor rS3-PA which blocks the phosphorylation site [48]. The reduction of P-Stat5 could be due to enhanced proteasomal degradation of activated Stat5-dimers associated with S5-DBD-PA making S5-DBD-PA a potent inhibitor of Stat5 and Stat5 dependent tumor cells (Figure 7).…”

“…In order to create the complete S5-DBD-PA peptide aptamer construct, applicable for protein transduction, purification, nuclear translocation and proper detection, the hTRXΔcys5 scaffold with the Stat5-DBD specific 12mer PA sequence was cloned together with a NLS-sequence, the9-R PTD and the 6xHis tag, C-terminal of the Flag tag as previously described [48]. …”

Low PIAS3 Expression in Malignant Mesothelioma Is Associated with Increased STAT3 Activation and Poor Patient Survival

“…STAT3 inhibition can also be affected by peptides (5,17,38). Our study indicates that the uptake of a STAT3-inhibiting peptide into mesothelioma cells is effective and induces cell growth inhibition and apoptosis.…”

“…STAT3 is constitutively phosphorylated and thereby activated in many forms of cancer. The inhibition of STAT3 activity often results in apoptosis and cell death (5,6). STAT3 serves as a central integration point for multiple oncogenic signaling pathways to regulate genes involved in cell-cycle control, apoptosis, angiogenesis, tumor invasion, and metastasis.…”