PMC9659814_01
The treatment scheme and representative CT scan images during treatment. (A) Treatment scheme. (B) The representative CT scan of during treatment courses. ADC, adenocarcinoma; SD, stable disease; PR, partial response.
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IVB lung adenocarcinoma was diagnosed on the basis of the combination of pathological results of biopsy specimen and CT ( Figures 1A, B ). Unfortunately, the lorlatinib treatment had to be discontinued due to the trachea metastasis after 5 months therapy ( Figures 1A, B ). DNA NGS was performed on the metastasis tumor specimen after resistance to the lorlatinib, and the novel reciprocal ROS1-CD74 fusion was detected again, which was also confirmed by FISH result ( Figures 1A ,  2B ).
Identification of a novel ROS1 fusion. (A) DNA NGS analysis of the genomic ROS1-CD74 novel fusion. (B) FISH analysis showed fused yellow signals (negative signal), single green signals (positive signal), and single red signals (positive signal) in the patient's specimen. (C) RNA NGS analysis of the dual CD74-ROS1 fusions at transcript level. (D) Possible schematic diagram of ROS1-CD74 fusion detected by DNA-based NGS, but CD74-ROS1 fusions were identified by RNA-based NGS. NGS, next-generation sequencing; TKI, tyrosine kinase inhibitor; FISH, fluorescence in situ hybridization.
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A novel reciprocal ROS1 rearrangement generated by intron 28 of ROS1 and intron 6 of CD74 was identified by sequencing ( Figure 2A ). TP63 p.H247R (variant allele frequency (VAF), 19.49%), and ACVR2A p.V433del (variant allele frequency (VAF), 16.38%). The gene fusion was confirmed by FISH with ROS1 break-apart probe ( Figure 2B ). Interestingly, dual functional fusions of CD74-ROS1 (E6:E32) and CD74-ROS1 (E6:E35) were presented at the transcriptional level by targeted RNA sequencing (Fusioncapture , Genetron Health) ( Figure 2C ), which was confirmed by Sanger's sequencing ( Figure 2D ). DNA NGS was performed on the metastasis tumor specimen after resistance to the lorlatinib, and the novel reciprocal ROS1-CD74 fusion was detected again, which was also confirmed by FISH result ( Figures 1A ,  2B ).