Antitumorigenic effects of a mutant of the heparin affin regulatory peptide on the U87 MG glioblastoma cell line

Abstract

Glioblastoma is the most common primary brain tumor in human adults. Since existing treatments are not effective enough, novel therapeutic targets must be sought. The heparin-binding growth factor, heparin affin regulatory peptide (HARP), also known as pleiotrophin (PTN), could potentially represent such a target. We have previously shown that a mutant protein, HARPD111-136, which lacks HARP's C-terminal 26 amino acids, acts as a dominant negative HARP effector by heterodimerizing with the wild-type growth factor. The aim of our study was to evaluate the potential inhibitory activity of HARPD111-136 on the U87 MG human glioblastoma cell line. By overexpressing the truncated form of HARP in stably established clones of U87 MG cells, we observed an inhibition of proliferation under both anchorage-dependent and anchorage-independent conditions. We confirmed these results in an in vivo subcutaneous tumor xenograft model. In addition, we found that HARPD111-136 inhibited cell proliferation in a paracrine manner. Analysis of key cellular pathways revealed a decrease of cell adhesion in U87 MG cells that overexpressed the mutant protein, which could explain this inhibitory effect. A replication-defective adenovirus model that encoded HARPD111-136 supported a putative antiproliferative role for the truncated protein in vitro and in vivo. Interestingly, HARPD111-136 was also able to abolish angiogenic activity in HUVEC proliferation and in a Matrigel plug assay. These results demonstrate that considering its antiproliferative and angiostatic effects, HARPD111-136 could be of great interest when used in conjunction with standard treatments.Glioblastoma is the most common primary brain tumor in adult humans, with a median survival time of 12 months from the time of diagnosis, despite using standard therapeutic approaches (surgery, radiation and chemotherapy). A proper understanding of the principal pathways implicated in glioblastoma progression is needed to identify potential therapeutic targets. The heparin-binding growth factor, heparin affin regulatory peptide (HARP), also known as pleiotrophin (PTN) and heparin-binding growth-associated molecule (HB-GAM), could represent 1 such potential target.HARP is a 136-amino acid secreted polypeptide that forms with the protein midkine (MK), a 2-member family of heparin-binding growth factors. HARP is a developmentally regulated cytokine that is highly expressed in the embryonic nervous system, but its expression drops markedly after the perinatal phase and during adulthood. However, HARP expression may be deregulated, and HARP has been shown to be overexpressed in various tumor cell lines and primary human tumors, 1-3 including glioblastomas. In vitro, HARP displays a variety of biological activities including mitogenesis, angiogenesis, neurite outgrowth and cell migration. 4 Transforming activity has also been demonstrated for HARP when it is overexpressed in nontumor cell lines, such as NIH 3T3. 5Two transmembrane proteins with intracellular catalytic domains have been descri...

HARPΔ111-136 enhances radiation-induced apoptosis of U87MG glioblastoma by induction of the proapoptotic protein CHOP

“…There was no difference between groups receiving RT alone or RT + Ad-CO1 indicating that the viral infection itself had no influence on the effect of the radiation therapy. In accordance with our previous data using intra-tumoral Ad-HARPΔ111-136 injection (19), a single systemic injection of Ad-HARPΔ111-136 inhibited tumor growth by 40% (p<0.01) when compared to control virus. A limited tumor growth was observed with the combination therapy, Ad-HARPΔ111-136 + RT.…”

“…The deleted HARP peptide HARPΔ111-136 was expressed in cells using an adenovirus as described previously (19). RT-PCR was performed on RNA transcripts extracted from U87MG cells infected for 48 h with Ad-HARPΔ111-136 or the control Ad-CO1 virus.…”

“…Ad-HARPΔ111-136 an E1-E3-deleted recombinant adenovirus that expresses the first 111 amino acids of the protein HARP was previously described (19). Adenovirus expressing no transgene (Ad-CO1) was used as control.…”

“…In a recent study (19) we demonstrated using the human glioblastoma cell line U87MG as an experimental model, that overexpression of this truncated protein inhibited in vitro and in vivo proliferation of this tumor cell line as well as in vivo angiogenesis. A replication-defective adenovirus encoding this truncated protein (Ad-HARPΔ111-136) was constructed and our results showed that direct intra-tumoral administration of Ad-HARPΔ111-136 inhibited growth and angiogenesis of established U87MG xenografts in nude mice.…”

The synthetic peptide P111-136 derived from the C-terminal domain of heparin affin regulatory peptide inhibits tumour growth of prostate cancer PC-3 cells

“…Previous studies suggested that the C-TSR domain and the C-terminus regions of HARP were particularly involved in the biological activities of HARP [25-28]. Thus, the C-terminus, composed of amino acids 111 through 136, was involved in the binding of HARP to the ALK receptor [25,26].…”