American Association for Cancer Research
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FIGURE 5 from SHP2 Inhibition with TNO155 Increases Efficacy and Overcomes Resistance of ALK Inhibitors in Neuroblastoma

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posted on 2023-12-27, 14:20 authored by Ivette Valencia-Sama, Lynn Kee, Gabriella Christopher, Michael Ohh, Mehdi Layeghifard, Adam Shlien, Madeline N. Hayes, Meredith S. Irwin

Dual SHP2/ALK inhibition resensitizes lorlatinib-resistant ALKF1174L cells and reduces tumor regrowth. A, Kelly cells were treated with increasing doses of lorlatinib (0.1–5.0 µmol/L) or DMSO over 90 days to select for subpopulations of lorlatinib-resistant (Kelly-LR) or -sensitive (Kelly-S) cells. Cell lines were maintained under chronic exposure to lorlatinib or DMSO. Created with Cell viability (alamarBlue) and IC50 was assessed in Kelly-S and Kelly-LR cells treated with increasing concentrations of lorlatinib alone (B) or lorlatinib plus TNO155 (C) for 72 hours. A maximum of 150 µmol/L of either or both drugs was serially diluted (1:3) to assess wide-range dose efficacy (0–150 µmol/L, doses of 0.0076, 0.0228, 0.0685, 0.205, 0.617, 1.851, 5.555, 16.666, 50, and 150 µmol/L). D, Western immunoblots of Kelly-S and Kelly-LR cells treated with TNO155 (1.5 µmol/L), lorlatinib (1 µmol/L), or combination treatment for 6 hours. Tumor growth (E) and Kaplan–Meier survival curves (F) of Kelly xenografts treated with two rounds of low-dose vehicle control (n = 4), TNO155 (n = 3), lorlatinib (n = 5), or combination treatment (Combo, n = 4) for 4 weeks each, with a 3 week drug break in between each round. **, P < 0.01; ***, P < 0.001.


Gouvernement du Canada | Canadian Institutes of Health Research (IRSC)

Sick Kids Neuroblastoma Research

James Fund

Curtis Chow Memorial Fund

Lilah's Fund

Sebastian's Superheroes



Survival rates among patients with high-risk neuroblastoma remain low and novel therapies for recurrent neuroblastomas are required. ALK is commonly mutated in primary and relapsed neuroblastoma tumors and ALK tyrosine kinase inhibitors (TKI) are promising treatments for ALK-driven neuroblastoma; however, innate or adaptive resistance to single-agent ALK-TKIs remain a clinical challenge. Recently, SHP2 inhibitors have been shown to overcome ALK-TKI resistance in lung tumors harboring ALK rearrangements. Here, we have assessed the efficacy of the SHP2 inhibitor TNO155 alone and in combination with the ALK-TKIs crizotinib, ceritinib, or lorlatinib for the treatment of ALK-driven neuroblastoma using in vitro and in vivo models. In comparison to wild-type, ALK-mutant neuroblastoma cell lines were more sensitive to SHP2 inhibition with TNO155. Moreover, treatment with TNO155 and ALK-TKIs synergistically reduced cell growth and promoted inactivation of ALK and MAPK signaling in ALK-mutant neuroblastoma cells. ALK-mutant cells engrafted into larval zebrafish and treated with single agents or dual SHP2/ALK inhibitors showed reduced growth and invasion. In murine ALK-mutant xenografts, tumor growth was likewise reduced or delayed, and survival was prolonged upon combinatorial treatment of TNO155 and lorlatinib. Finally, we show that lorlatinib-resistant ALK-F1174L neuroblastoma cells harbor additional RAS-MAPK pathway alterations and can be resensitized to lorlatinib when combined with TNO155 in vitro and in vivo. Our results report the first evaluation of TNO155 in neuroblastoma and suggest that combinatorial inhibition of ALK and SHP2 could be a novel approach to treating ALK-driven neuroblastoma, potentially including the increasingly common tumors that have developed resistance to ALK-TKIs. These findings highlight the translatability between zebrafish and murine models, provide evidence of aberrant RAS-MAPK signaling as an adaptive mechanism of resistance to lorlatinib, and demonstrate the clinical potential for SHP2/ALK inhibitor combinations for the treatment of ALK-mutant neuroblastoma, including those with acquired tolerance or potentially resistance to ALK-TKIs.