Ceritinib-Induced Regression of an Insulin-Like Growth Factor-Driven Neuroepithelial Brain Tumor

Abstract

The insulin-like growth factor (IGF) pathway plays an important role in several brain tumor entities. However, the lack of inhibitors crossing the blood-brain barrier remains a significant obstacle for clinical translation. Here, we targeted the IGF pathway using ceritinib, an off-target inhibitor of the IGF1 receptor (IGF1R) and insulin receptor (INSR), in a pediatric patient with an unclassified brain tumor and a notch receptor 1 (NOTCH1) germline mutation. Pathway analysis of the tumor revealed activation of the sonic hedgehog (SHH), the wingless and integrated-1 (WNT), the IGF, and the Notch pathway. The proliferation of the patient tumor cells (225ZL) was inhibited by arsenic trioxide (ATO), which is an inhibitor of the SHH pathway, by linsitinib, which is an inhibitor of IGF1R and INSR, and by ceritinib. 225ZL expressed INSR but not IGF1R at the protein level, and ceritinib blocked the phosphorylation of INSR. Our first personalized treatment included ATO, but because of side effects, we switched to ceritinib. After 46 days, we achieved a concentration of 1.70 µM of ceritinib in the plasma, and after 58 days, MRI confirmed that there was a response to the treatment. Ceritinib accumulated in the tumor at a concentration of 2.72 µM. Our data suggest ceritinib as a promising drug for the treatment of IGF-driven brain tumors.

Keywords: ATO; IGF; NOTCH1; SHH; WNT; ceritinib.

Figure 1

Histopathological features of the primary tumor. (A) HE staining showing small, round, blue tumor cells. (B) Epithelial antigens (EMA). (C) NeuN. (D) CD56. (E) Ki67. (F) L1Cam. Original magnification 200×.

Figure 2

Principal Component Analysis. Principal component analysis by genes based on 850 k DNA methylation analysis for different tumor entities commonly found in childhood. Samples of the index patient do not cluster together with ependymoma, RELA fusion-positive tumors (arrow heads), but rather form their own cluster (arrows). The kind of material used for the analysis is indicated (fresh frozen or formalin-fixed, paraffin-embedded (FFPE)).

Figure 3

Embryonal pathways are activated in the tumor relapse. GLI family zinc finger 2 (GLI2) (A), AXIN2 (B), hes family bHLH transcription factor 4 (HES4) (C) and insulin-like growth factor 2 (IGF2) (D) expression was analyzed by qRT-PCR in the relapse (no 225), the primary tumor cells isolated from the relapse (no 225ZL), and two normal brain regions (no 110, 111). After normalization to the housekeeping gene hypoxanthine phosphoribosyltransferase 1 (HPRT1), the relative quantification value was expressed as 2^-ΔΔCt. The qPCR experiments were carried out in biological triplicates. Data are represented as the mean ± standard deviation (SD). Statistical analyses were performed using t-tests compared with sample 111 (* p < 0.01, *** p < 0.001, **** p < 0.0001).

Figure 4

Detection of a notch receptor 1 (NOTCH1) germline mutation and a tumor protein P53 (TP53) somatic mutation. Sanger sequencing of the genomic DNA extracted from the blood and the relapse (no 225) was done with primers specific for the NOTCH1 (A) and the TP53 (C) mutation identified by RNA-seq. The arrow indicates the (NM_017617.3:c.689G > A) mutation rs121912651. The arrowhead indicates the rs121912651. (B) Structure of NOTCH1 as reported in cBioportal. The position of the Gly230Glu mutation is indicated. Green: EGF-like domains; Red: Calcium-binding EGF domains; Yellow: LNR Domain; Purple: NOD (NOTCH protein domain); Orange: NODP (NOTCH protein domain); Rose: Ankyrin repeats; Dark red: unknown function.

Figure 5

The primary tumor cells are sensitive to arsenic trioxide (ATO) and ceritinib. (A) 225ZL cells were grown for 15 days in the presence of ATO, vismodegib, linsitinib, or ceritinib. As control, the vehicle alone was used. The percentage of the growth compared to the control is shown. The proliferation experiments were carried out in biological duplicates. Data are represented as the mean ± standard deviation (SD). (B) 225ZL cells were stimulated after starvation with IGF2 in the presence or absence of 1 µM of ceritinib. The expression of IGF1R, insulin receptor (INSR), and the phosphorylated form of INSR (P-INSR) was analyzed by Western blot with specific antibodies. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as loading control. A representative experiment of two independent experiments is shown.

Figure 6

Personalized therapy protocol. Chemotherapy incorporated systemic (ctx) VA (vincristine and actinomycin-D, red) and VAd (adriamycin, blue) blocks, and intraventricular (via Ommaya) cytarabine (araC, yellow) and etoposide-Gry (eto, green). ATO was administered to coincide with the chemotherapy backbone as an intravenous (iv) treatment for 1 week (bright grey), and was subsequently switched to an oral ATO formulation (dark grey). The duration of ATO treatment lasted 68 days (9.7 weeks) in total. Ceritinib was given for 93 days. A metastatic lesion in the cerebellum was detected by MRI after 30 days of ceritinib treatment in combination with VA and intraventricular therapy (blue star). After 30 days of ceritinib monotherapy following the detection of the metastatic relapse, a response to the treatment was confirmed by MRI (red star). Surgical removal of the sanguineous necrotic area was performed (big blue arrow). A third relapse was detected by MRI (green star) two weeks after the detection of the response. The patient died 27 months after initial diagnosis and 3.5 months after the third relapse.

Figure 7

The target lesion is sensitive to the targeted therapy with ceritinib. MRI before therapy with ceritinib (A–C) showing a right sided solid mesiotemporal tumor (A, white arrow) with a cystic component, as well as a contrast media enriching metastatic lesion within the cerebellar folia (C, FLAIR post-contrast media, white arrow head). After 30 days of ceritinib (D–F), the metastatic lesion was regredient (F, FLAIR post-contrast media, white arrow head), and hemorrhagic transformation (E, T2*, black arrow) of the tumor was detected.