Technical CommentsSepsis

Comment on “ALK is a therapeutic target for lethal sepsis”

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Science Translational Medicine  12 Dec 2018:
Vol. 10, Issue 471, eaar4321
DOI: 10.1126/scitranslmed.aar4321
  • Fig. 1 Undetectable ALK expression in monocytes and macrophages.

    (A) ALK mRNA expression in tissues according to the GTEx project. TPM, transcripts per million. (B) ALK expression in murine cell lines [top: SP8 anti-ALK antibody (1:1000); bottom: F-12 anti-ALK antibody (1:100)]. 293T cells transfected with mouse Alk (293T_ALK) and EML4-ALK–expressing TeCla1 cells (positive controls). Untransfected 293T and KP1233 murine KRASG12D-driven NSCLC cells (negative controls). Arrows indicate the full-length (f.l.) ALK or the EML4-ALK fusion. (C) ALK expression in human cell lines [top: D5F3 anti-ALK antibody (1:1000); bottom: F-12 anti-ALK antibody (1:100)]. Human neuroblastoma cell lines SK-N-BE and SH-SY5Y (positive controls). Human B-lymphoma Raji and Ramos cell lines (negative controls). Arrows indicate the full-length ALK or the cleaved ALK. Asterisk indicates the nonspecific band detected by the F-12 antibody. (D) ALK expression in human cell lines [left: D5F3 anti-ALK (1:1000); right: F-12 anti-ALK antibody (1:100)]. THP-1 cells transduced with human ALK receptor (THP-1_ALK) (positive control). THP-1 cells transduced with empty retroviral vector (THP-1_vector) (negative control). H3122 and H2228 NSCLC cell lines carry the EML4-ALK (ex13-ex20) and the EML4-ALK (ex6-ex20) rearrangements that generate fusion proteins of 130 and 90 kDa, respectively. Karpas-299 is an NPM-ALK–rearranged cell line that generates a fusion protein of 80 kDa. Arrows indicate the full-length ALK or the cleaved ALK. Asterisk indicates the nonspecific band detected by the F-12 antibody. Top panels: Short exposure. Bottom panels: Long exposure. (E) Immunohistochemistry performed on an ALK-rearranged NSCLC case (D5F3 anti-ALK antibody) and on an ALCL case (4C5B8 anti-ALK antibody). Tumor cells shown by black arrows; macrophages shown by red arrows. Scale bar, 50 μm.

  • Fig. 2 Undetectable ALK mRNA expression in Raji and THP-1 cell lines.

    ALK mRNA expression in different tumor cell lines according to two different datasets in CCLE (https://portals.broadinstitute.org/ccle). (A) RNA sequencing (RNA-seq) data. (B) Gene expression profiling data. Raji, THP-1, and the ALK-positive neuroblastoma cell lines SH-SY5Y and SK-N-BE are indicated by arrows.

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