Peptides derived from the dependence receptor ALK are proapoptotic for ALK-positive tumors

Abstract

ALK is a receptor tyrosine kinase with an oncogenic role in various types of human malignancies. Despite constitutive activation of the kinase through gene alterations, such as chromosomal translocation, gene amplification or mutation, treatments with kinase inhibitors invariably lead to the development of resistance. Aiming to develop new tools for ALK targeting, we took advantage of our previous demonstration identifying ALK as a dependence receptor, implying that in the absence of ligand the kinase-inactive ALK triggers or enhances apoptosis. Here, we synthesized peptides mimicking the proapoptotic domain of ALK and investigated their biological effects on tumor cells. We found that an ALK-derived peptide of 36 amino acids (P36) was cytotoxic for ALK-positive anaplastic large-cell lymphoma and neuroblastoma cell lines. In contrast, ALK-negative tumor cells and normal peripheral blood mononuclear cells were insensitive to P36. The cytotoxic effect was due to caspase-dependent apoptosis and required N-myristoylation of the peptide. Two P36-derived shorter peptides as well as a cyclic peptide also induced apoptosis. Surface plasmon resonance and mass spectrometry analysis of P36-interacting proteins from two responsive cell lines, Cost lymphoma and SH-SY5Y neuroblastoma, uncovered partners that could involve p53-dependent signaling and pre-mRNA splicing. Furthermore, siRNA-mediated knockdown of p53 rescued these cells from P36-induced apoptosis. Finally, we observed that a treatment combining P36 with the ALK-specific inhibitor crizotinib resulted in additive cytotoxicity. Therefore, ALK-derived peptides could represent a novel targeted therapy for ALK-positive tumors.

Crizotinib Synergizes with Chemotherapy in Preclinical Models of Neuroblastoma

“…Additionally, synthetic peptides corresponding to the proapoptotic domain of ALK caused p53-mediated cytotoxicity in ALCL and neuroblastoma cells (31). In the same study, ALK peptides interacted with proteins that have been previously reported to interact with the p53 gene and protein.…”

Anaplastic Lymphoma Kinase (ALK) Receptor Tyrosine Kinase: A Catalytic Receptor with Many Faces

“…One crucial characteristic of ALK is that it is a so-called dependence receptor [ 39 , 40 , 41 ] ( Figure 2 ). Without ligand binding to activate its kinase activity, ALK can be cleaved by caspase-3 during apoptosis [ 39 , 40 , 41 ]. In the juxtamembrane region of ALK, there is a caspase-3 cleavage site (amino acids 1160–1163: DELD) ( Figure 2 ).…”

“…Elevated caspase-3 activity can cleave this ALK at this cleavage site, which releases an intracellular ALK fragment (about 60 kDa) into the cytoplasm. This caspase-dependent cleavage of ALK enhances apoptosis through the exposure of a pro-apoptotic segment (addiction/dependence domain, ADD) within the ALK juxtamembrane region [ 39 , 40 , 41 ]. ALK mutant D1160N abolishes the caspase-3 cleavage at this cleavage position, which also abrogates the ALK-mediated enhancement of apoptosis [ 39 , 40 , 41 ].…”

“…This caspase-dependent cleavage of ALK enhances apoptosis through the exposure of a pro-apoptotic segment (addiction/dependence domain, ADD) within the ALK juxtamembrane region [ 39 , 40 , 41 ]. ALK mutant D1160N abolishes the caspase-3 cleavage at this cleavage position, which also abrogates the ALK-mediated enhancement of apoptosis [ 39 , 40 , 41 ]. Moreover, people have found that synthesized peptides mimicking the proapoptotic domain of ALK caused cytotoxicity to ALK-positive ALCL and neuroblastoma (NB) cell lines [ 41 ].…”

“…ALK mutant D1160N abolishes the caspase-3 cleavage at this cleavage position, which also abrogates the ALK-mediated enhancement of apoptosis [ 39 , 40 , 41 ]. Moreover, people have found that synthesized peptides mimicking the proapoptotic domain of ALK caused cytotoxicity to ALK-positive ALCL and neuroblastoma (NB) cell lines [ 41 ]. This cytotoxic effect was found to be due to caspase-dependent apoptosis [ 41 ].…”