Multiplexed quantitative proteomics in small-cell lung cancer: stage-linked DLL3 distribution and an independent prognostic signal of thymidine phosphorylase.

Background: The regulatory approval of the delta-like ligand 3 (DLL3)-directed bispecific T-cell engager tarlatamab has positioned DLL3 as a clinically actionable target in small-cell lung cancer (SCLC), yet whether DLL3 is independently prognostic alongside other therapy-relevant proteins in real-world SCLC cohorts remains unknown. We addressed this using standardized mass spectrometry-based absolute quantitation. Methods: We applied selected reaction monitoring-mass spectrometry (SRM-MS) to archival formalin-fixed paraffin-embedded tumor tissue from 100 patients with SCLC treated with first-line platinum/etoposide (2005-2015), quantifying nine therapy-relevant proteins (DLL3, ASCL1, EZH2, SLFN11, TYMP, MGMT, CD56, TUBB3, TOPO1) in absolute molar units. Associations with tumor stage, platinum-sensitivity status, and overall survival (OS) were analyzed. Results: SRM-MS yielded interpretable measurements in all 100 tumors. DLL3 was tightly stage-linked, detected in 76% of extensive-stage versus 29% of limited-stage tumors (area under the receiver operating characteristic curve 0.78; P<0.001). A four-class DLL3/ASCL1 model stratified OS (P=0.004); however, stage-stratified analyses showed that this signal was confined to limited-stage disease and largely driven by a small DLL3-/ASCL1- subgroup (n=9, median OS 25.8 months). In multivariable Cox regression, detectable TYMP emerged as an independent predictor of shorter OS. Conclusions: Multiplexed SRM-MS is clinically applicable to routinely archived FFPE SCLC tissue and provides a quantitative framework that may help address the unmet clinical need for biomarker-guided patient selection in SCLC, particularly in the era of DLL3-directed therapy.