Novel CAD-ALK gene rearrangement is drugable by entrectinib in colorectal cancer

Abstract

Background: Activated anaplastic lymphoma kinase (ALK) gene fusions are recurrent events in a small fraction of colorectal cancers (CRCs), although these events have not yet been exploited as in other malignancies.

Methods: We detected ALK protein expression by immunohistochemistry and gene rearrangements by fluorescence in situ hybridisation in the ALKA-372-001 phase I study of the pan-Trk, ROS1, and ALK inhibitor entrectinib. One out of 487 CRCs showed ALK positivity with a peculiar pattern that prompted further characterisation by targeted sequencing using anchored multiplex PCR.

Results: A novel ALK fusion with the carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD) gene (CAD-ALK fusion gene) was identified. It resulted from inversion within chromosome 2 and the fusion of exons 1-35 of CAD with exons 20-29 of ALK. After failure of previous standard therapies, treatment of this patient with the ALK inhibitor entrectinib resulted in a durable objective tumour response.

Conclusions: We describe the novel CAD-ALK rearrangement as an oncogene and provide the first evidence of its drugability as a new molecular target in CRC.

Figure 1

Histological, immunohistochemical, and fluorescence in situ hybridisation analyses of the primary right colonic tumour, lymph node, and liver metastasis of the case presented. Haematoxylin & eosin, immunohistochemical and FISH images of the primary colon tumour (A 1–3; N.M: normal mucosa, T: tumour), lymph node (B 1–3), and liver metastasis (C 1–3) of the patient presented in this report, showing malignant tumour (A1, B1, C1), ALK protein overexpression (A2, B2, C2) and ALK gene rearrangements (A3, B3, C3, white arrows). Original magnification of images: × 100 for haematoxylin & eosin and immunohistochemical staining, Insert × 400; × 630 for FISH analysis.

Figure 2

Identification of the CAD-ALK gene rearrangement. (A) The upper section shows a schematic representation of the CAD-ALK genomic DNA rearrangement and the resulting transcript. The sequence spanning the rearrangement junction is also shown. Exons are represented by coloured boxes, and introns are represented by lines: CAD in red and ALK in light blue. The lower section shows the functional domains conserved in the chimaeric CAD-ALK protein. (B) Characterisation of the CAD-ALK transcript by PCR. Agarose gel showing amplification with primers for the rearranged CAD-ALK chimaeric transcript, spanning CAD exon 35 to ALK exon 20. The tumour sample was compared with a negative control sample (U138-MG cell line, expressing ALK full length).

Figure 3

Computed tomography (CT) scans showing the objective tumour response to entrectinib. The baseline abdominal CT scan of March 2015 demonstrated liver involvement with the two largest lesions both in hepatic segment VII, measuring 27 and 33 mm in longest diameter, respectively (A, C arrows). At the first-response assessment, in April 2015, 4 weeks after the initiation of treatment, CT showed a RECIST partial response with an overall decrease in the sum of the target lesions of 38%, and lesions in segment VII displaying longest diameters of 15 and 22 mm (B, D arrows).