Research Papers:

The CNGRC-GG-D(KLAKLAK)2 peptide induces a caspase-independent, Ca2+-dependent death in human leukemic myeloid cells by targeting surface aminopeptidase N/CD13

Sandrine Bouchet, Ruoping Tang, Fanny Fava, Ollivier Legrand and Brigitte Bauvois _

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Oncotarget. 2016; 7:19445-19467. https://doi.org/10.18632/oncotarget.6523

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Sandrine Bouchet1,2, Ruoping Tang3,4, Fanny Fava3,4, Ollivier Legrand3,4, Brigitte Bauvois1

1Centre de Recherche des Cordeliers, INSERM UMRS1138, Sorbonne Universités UPMC Paris 06, Université Paris Descartes Sorbonne Paris Cité, Paris, France

2Assistance Publique des Hôpitaux de Paris, Paris, France

3Centre de Recherche de Saint-Antoine, INSERM UMRS 938, Service d’Hématologie, Hôpital St Antoine, Paris, France

4Sorbonne Universités UPMC Paris 06, Paris, France

Correspondence to:

Brigitte Bauvois, e-mail: [email protected]

Keywords: calcium, leukemia, metalloproteinase, necrosis, superoxide radical

Received: July 30, 2015     Accepted: November 16, 2015     Published: December 09, 2015


The CD13 antigen’s binding site for the Asn-Gly-Arg (NGR) motif enables NGR-containing chemotherapeutic drugs to be delivered to CD13-positive tumours. Human CD13-positive acute myeloid leukemia (AML) cells proliferate abnormally and escape death. Here, we show that the CNGRC-GG-D(KLAKLAK)2 peptide induces death in AML cell lines (U937, THP-1, NB4, HL-60) and primary blood cells from AML patients. Cell death was characterized as a caspase-independent mechanism, without DNA fragmentation, but phosphatidylserine externalization and membrane disruption. Our results demonstrate in U937 cells that (i) the NGR-peptide triggers the loss of mitochondrial potential(ΔΨm) and generates superoxide anion (O2), (ii) N-acetyl-L-cysteine (NAC) and extra/intracellular Ca2+ chelators (BAPTA) prevent both O2− production and cell death, (iii) the Ca2+-channel blocker nifedipine prevents cell death (indicating that Ca2+ influx is the initial death trigger), and (iv) BAPTA, but not NAC, prevents ΔΨm loss (suggesting O2 is a mitochondrial downstream effector). AML cell lines and primary blasts responding to the lethal action of NGR-peptide express promatrix metalloproteinase-12 (proMMP-12) and its substrate progranulin (an 88 kDa cell survival factor). A cell-free assay highlighted proMMP-12 activation by O2. Accordingly, NGR-peptide’s downregulation of 88 kDa progranulin protein was prevented by BAPTA and NAC. Conversely, AML blast resistance to NGR-peptide is associated with the expression of a distinct, 105 kDa progranulin isoform. These results indicate that CNGRC-GG-D(KLAKLAK)2 induces death in AML cells through the Ca2+-mitochondria-O2.-pathway, and support the link between proMMP-12 activation and progranulin cleavage during cell death. Our findings may have implications for the understanding of tumour biology and treatment.

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