MET TKD mutations in NSCLC cohort of Caris Life Sciences. (A) Flowchart of the NSCLC subgroup in the Caris Life Sciences dataset. (B) Prevalence of MET TKD mutations in NSCLC cases in the Caris Life Sciences dataset (MET TKD mutations in 280 unique patients). (C) Lollipop plot of the oncogenic / likely oncogenic MET TKD mutations detected in NSCLC cases in the Caris Life Sciences dataset (35 oncogenic / likely oncogenic MET TKD mutations in 35 unique patients). (D) concurrent driver alterations (information available for 24 cases, unavailable for 11 cases) and (E) concurrent MET amplification status (information available for 33 cases, unavailable for 2 cases) of NSCLC cases harboring oncogenic / likely oncogenic MET TKD mutations in the Caris Life Sciences dataset.

Concurrent driver mutations according to oncogenic / likely oncogenic MET TKD mutations detected in cohort #2.

Equilibration of interactions that drives ATP affinity in the presence MET R1170Q mutant. (A) Representative structure of ATP bound MET R1170Q with the regions of interest highlighted [hinge site (green), and mutation site (blue)]. (B) Mutation at position 1170 facilitates the formation of a hydrogen bonding network. (C) Interatomic distances for non solvent mediated hydrogen bonding.

BACKGROUND:While PDZ-LIM domain-containing protein (PDLIM2) suppresses lung cancer, its clinical significance and therapeutic potential remain to be fully explored. METHODS:Next-generation sequencing and immunohistochemistry of programmed death ligand 1 (PD-L1) were performed on lung adenocarcinoma (LUAD) tissues from 15,765 patients. PDLIM2 gene therapy was tested in syngeneic LUAD mouse models using intravenous nano-complexed plasmid DNA, alone or with chemoimmunotherapy. RESULTS:PDLIM2-high tumors were more common in primary/local biopsies versus metastatic samples (73.1% vs 53.0%; p < 0.001). High PDLIM2 expression was linked to lower mutation rates in RB1, TP53, SMARCA4, STK11, and KEAP1, but increased EGFR mutations (all p < 0.01). PDLIM2-high tumors also showed increased immune cell infiltration, T cell-inflamed scores, and PD-L1 positivity (all p < 0.008). High PDLIM2 was associated with improved survival (24.1 vs 18.1 months; p < 0.001, HR = 0.84) and remained prognostic in multivariate analysis (HR = 0.88, 95% CI 0.83-0.93), as well as with longer pembrolizumab time, especially with platinum-based therapy (p = 0.012, HR = 0.867). In two LUAD mouse models, nanoPDLIM2 improved median overall survival versus chemoimmunotherapy alone (10-12.5 vs 8-9 days; N = 8; p = 0.0001, 0.0003). CONCLUSION:PDLIM2 is a promising prognostic biomarker in LUAD linked to immune-receptive tumors. Preclinically, PDLIM2-based therapy enhances chemoimmunotherapy efficacy, though its predictive role warrants further evaluation.

Single trajectory generalized Born and surface area solvation (MM/GBSA) calculated free energy of ATP binding into the orthosteric pocket.

List of missense mutations with unknown biologic function and their pathogenicity scores using all three in silico tools in cohort #1

Concurrent genomic drivers in cases with oncogenic / likely oncogenic MET tyrosine kinase domain (TKD) mutations and co-occurring MET gene amplification in cohort #2.

Introduction Small cell lung cancer (SCLC) and extrapulmonary neuroendocrine (NE) tumors are aggressive malignancies with limited treatment options. Seizure-related homolog 6 (SEZ6) is a potential therapeutic target, but its expression in these tumors remains poorly understood. Lineage plasticity contributes to resistance in non-small cell lung cancer (NSCLC), where some cases can undergo SCLC-transformation after targeted therapy. We aimed to characterize SEZ6 expression across NE tumors and presumed NSCLC-to-SCLC transformations. Methods DNA and RNA sequencing were performed for SCLC, NSCLC, and NE samples. Samples were stratified by SEZ6 RNA expression quartiles and classified into subtypes based on ASCL1, NEUROD1, and POU2F3 expression. Significance was tested using the Mann-Whitney U test. Real-world overall survival was obtained from insurance claims data, with p-values calculated using the log-rank test. Paired samples for NSCLC-to-SCLC transformation were identified by sequential biopsies classified as NSCLC followed by SCLC. Results RNA sequencing was performed on 1318 SCLC and 2218 NE samples. Median SEZ6expression was higher in SCLC (39.7 transcripts per million (TPM)) than in NE tumors (20.8 TPM, p<0.0001) and NSCLC (1.34 TPM, p<0.001). Among NE tumors, median SEZ6 expression was highest in prostate (52.0 TPM, p=0.0016 vs SCLC) and lowest in adrenal gland tumors (1.2 TPM, p<0.0001 vs SCLC). In SCLC,SEZ6expression was positively correlated with ASCL1(p=0.44, p<0.0001) andNEUROD1(p=0.16, p<0.0001) expression but notPOU2F3(p=-0.04, p=0.1253). Median survival was longest in SEZ6-Q2 for both SCLC and NE (13.0 mos. and 33.7 mos., respectively). NSCLC-to-SCLC transformation samples showed numerically higher SEZ6 expression post-transformation (median: 86.2 vs 2.4 TPM). Conclusions SEZ6expression is higher in SCLC than in NE tumors, with notable heterogeneity by subtype, warranting consideration of expanded use of SEZ6-directed therapy.Translational Relevance Statement:This study establishes SEZ6 as a promising therapeutic target in small cell lung cancer (SCLC) and transformed non-small cell lung cancer (NSCLC), demonstrating its significantly elevated expression compared to neuroendocrine (NE) tumors and NSCLC. The positive correlation of SEZ6 expression with NE lineage markers, particularly in ASCL1 and NEUROD1 subtypes, highlights its role as a lineage-specific marker, guiding the development of SEZ6-targeted antibody-drug conjugates (ADCs). Additionally, the increased SEZ6 expression following NSCLC-to-SCLC transformation suggests that SEZ6-targeted therapies could address resistance mechanisms in transformed tumors. Importantly, the association between high SEZ6 expression and shorter survival indicates that integrating SEZ6 status into diagnostic workflows could help stratify patients by risk and guide therapeutic decision-making. The findings from this study will inform future clinical trials, aiming to implement SEZ6-targeted treatments as part of precision oncology strategies for aggressive NE malignancies.

Additional information regarding the methods used in this study

Detailed genomic characteristics for each case of NSCLC with oncogenic / likely oncogenic MET tyrosine kinase domain (TKD) mutations in cohort #1.

Clinicopathologic and genomic characteristics of 171 cases of cancers other than NSCLC harboring oncogenic / likely oncogenic MET TKD mutations in cohort #2.

Study flowchart. Cohort #1 (multi-institutional cohort): GENIE V.11.0, China Pan-cancer (OrigiMed2020, Nature 2022), The Cancer Genome Atlas (TCGA) Pan-Cancer Atlas studies, Dana-Farber Cancer Institute (DFCI) and NSCLC from Memorial Sloan Kettering Cancer Center (MSKCC); cohort #2: Foundation Medicine Pan-Cancer cohort.

Comparison of clinicopathologic characteristics between NSCLCs with oncogenic / likely oncogenic MET tyrosine kinase domain (TKD) mutations without other concurrent drivers and NSCLCs with MET exon 14 alterations in the Caris Life Sciences cohort.

Detailed clinicopathologic and genomic characteristics of the 61 cases with a malignancy other than NSCLC harboring an oncogenic / likely oncogenic MET TKD mutation in cohort #1.

Sensitivity pattern of MET TKD mutations in 293T cells. (A) ERK-mediated transcriptional activation assessed by luciferase assay in 293T cells co-transfected with MET WT or TKD mutant constructs, an ERK activation reporter vector (pGL4.33), and a control co-reporter vector (pRL-TK). The lower panel shows western blot analysis of transfected samples. (B) Western blot analysis of MET and ERK1/2 phosphorylation in response to 1 μM crizotinib. MET TKD mutants showing sensitivity and resistance to crizotinib are colored in green and orange, respectively.

Background/Objectives: Overexpression of transferrin receptor (TFR1) is common in cancer and may be associated with inferior treatment outcomes. Due to these patterns and TFR1's essential role in iron metabolism, the protein has been targeted for cytotoxic drug delivery. More recently, increased TFR1 expression has been linked to tumor microenvironment (TME) infiltration by immune effectors in selected tumors, but a comprehensive assessment of the genomic landscape associated TFRC (the gene encoding TFR1) expression has not been conducted. Methods: By utilizing a pan-cancer database of 93,248 patients with whole-exome and whole-transcriptome sequencing, we assessed TFRC-associated multiomic patterns. Results: We found that high TFRC expression correlates with significantly worse overall survival in multiple common solid tumor types, a higher tumor mutational burden (TMB), an increase in infiltrating effector cells with upregulated immune checkpoint markers within the TME, and increased frequency of specific high-risk genomic alterations. Further assessment in cell line models revealed increased susceptibility to cytotoxic T cells when iron metabolism is elevated, despite upregulation of the checkpoint ligand PD-L1. Conclusions: High TFRC expression, therefore, indicates worse clinical risk across multiple common tumor types but potentially increased susceptibility to cytotoxic immune effectors, informing the development of TFR1 biomarker-driven therapeutic strategies.

Frequency of MET tyrosine kinase domain (TKD) mutations in cohort #1 according to amino acid residue, out of a total of 711 MET TKD mutations.

Type and frequency of MET TKD mutations across cancer types in cohort #1. (A) Frequency of MET TKD mutations in various cancer types in cohort #1 according to OncoKB status; the numbers above each bar indicate the number of cases with MET TKD mutations (regardless OncoKB annotations) out of the total of cases of each cancer type. (B) Detailed oncogenic / likely oncogenic MET TKD mutations according to cancer type in cohort #1.

Prevalence of MET tyrosine kinase domain (TKD) mutations across cancer types in cohort #2.

Mutagenic primers used for mutagenesis.