A Molecular Epidemiological Analysis Of Programmed Cell Death Ligand-1 (PD-L1) Protein Expression, Mutations And Survival In Non-Small Cell Lung Cancer.

PURPOSE: To characterize programmed cell death ligand-1 (PD-L1) expression in relation to survival and gene mutation status in patients with advanced NSCLC. The study also explored the influence of tumor mutational burden (TMB) on PD-L1 expression and patient characteristics. PATIENTS AND METHODS: Adult patients with histologically or cytologically documented Stage IIIB/Stage IV/recurrent/progressive NSCLC, Eastern Cooperative Oncology Group performance status 0 to 3, and >2 lines of prior systemic treatment regimens were included in this retrospective analysis. Patients were treated from 1997 to 2015 at H. Lee Moffitt Cancer Center and Research Institute, Tampa, or at 7 community centers across the United States. PD-L1 expression level was determined using the VENTANA PD-L1 (SP263) Assay. EGFR and KRAS mutation status and ALK rearrangements were determined by targeted DNA sequencing; these were obtained from clinical records where targeted DNA sequencing was not performed. TMB was calculated as the total number of somatic mutations per sample. RESULTS: From a total of 136 patients included in the study, 23.5% had tumors with high PD-L1 expression (≥25%). There were no significant differences in patient characteristics, overall survival (OS), and progression-free survival (PFS) between patients with high PD-L1 expression (median OS: 39.5 months; median PFS: 15.8 months) vs low PD-L1 expression (<25%; median OS: 38.1 months; median PFS: 18.6 months). PD-L1 expression level correlated (P=0.05) with TMB and was consistent with The Cancer Genome Atlas data. CONCLUSION: In this retrospective analysis, survival outcomes of patients with advanced NSCLC were comparable by PD-L1 expression level. EGFR and KRAS mutation status were not found to be significantly associated with PD-L1 expression level, while TMB was weakly associated with PD-L1 expression level. Overall, PD-L1 expression level was not observed to be an independent prognostic biomarker in this cohort of patients with advanced NSCLC treated with chemotherapy.

Sequence, "subtle" alternative splicing and expression of the CYYR1 (cysteine/tyrosine-rich 1) mRNA in human neuroendocrine tumors.

BACKGROUND: CYYR1 is a recently identified gene located on human chromosome 21 whose product has no similarity to any known protein and is of unknown function. Analysis of expressed sequence tags (ESTs) have revealed high human CYYR1 expression in cells belonging to the diffuse neuroendocrine system (DNES). These cells may be the origin of neuroendocrine (NE) tumors. The aim of this study was to conduct an initial analysis of sequence, splicing and expression of the CYYR1 mRNA in human NE tumors. METHODS: The CYYR1 mRNA coding sequence (CDS) was studied in 32 NE tumors by RT-PCR and sequence analysis. A subtle alternative splicing was identified generating two isoforms of CYYR1 mRNA differing in terms of the absence (CAG- isoform, the first described mRNA for CYYR1 locus) or the presence (CAG+ isoform) of a CAG codon. When present, this specific codon determines the presence of an alanine residue, at the exon 3/exon 4 junction of the CYYR1 mRNA. The two mRNA isoform amounts were determined by quantitative relative RT-PCR in 29 NE tumors, 2 non-neuroendocrine tumors and 10 normal tissues. A bioinformatic analysis was performed to search for the existence of the two CYYR1 isoforms in other species. RESULTS: The CYYR1 CDS did not show differences compared to the reference sequence in any of the samples, with the exception of an NE tumor arising in the neck region. Sequence analysis of this tumor identified a change in the CDS 333 position (T instead of C), leading to the amino acid mutation P111S. NE tumor samples showed no significant difference in either CYYR1 CAG- or CAG+ isoform expression compared to control tissues. CYYR1 CAG- isoform was significantly more expressed than CAG+ isoform in NE tumors as well as in control samples investigated. Bioinformatic analysis revealed that only the genomic sequence of Pan troglodytes CYYR1 is consistent with the possible existence of the two described mRNA isoforms. CONCLUSION: A new "subtle" splicing isoform (CAG+) of CYYR1 mRNA, the sequence and the expression of this gene were defined in a large series of NE tumors.

Differential expression of alternatively spliced mRNA forms of the insulin-like growth factor 1 receptor in human neuroendocrine tumors.

The activation of the insulin-like growth factor 1/IGF1 receptor system (IGF1/IGF1R) is a critical event in the transformation and tumorigenicity processes in a wide variety of human tumors. The IGF1/IGF1R system has been recently studied in carcinoid tumors that often arise in the gastrointestinal tract; these tumors are characterized by hypersecretion of bioamines and neuropeptides, leading to functional tumor disease. Two alternatively spliced IGF1R mRNA transcripts have been described to differ by only three nucleotides (CAG) in the coding sequence, resulting in an amino-acid change from the originally described Thr-Gly to an Arg in the extracellular portion of the receptor beta subunit. In transfected Chinese hamster ovary cells, the form without CAG (CAG-) exhibited an approximate 2-fold increase in IGF1 stimulation of activities required for its mitogenic properties. In this study, we examine the relative expression of the two IGF1R mRNA isoforms by a semiquantitative RT-PCR approach using highly standardized conditions, beta-2 microglobulin (B2M) as a reference gene and gel imaging analysis. We analyzed a large series of human neuroendocrine tumors (32 samples) and 9 normal tissues. A significant higher expression of both isoforms in the tumor samples (approximately 2-fold increase) was found, while a constant CAG+/CAG- IGF1R mRNA isoforms of an approximate 3:1 ratio was observed in all tumoral and normal cell types studied. The phylogenetic study of the IGF1R locus in several species suggests that human IGF1R CAG- mRNA isoform is evolutionarily more recent compared to the IGF1R CAG+ mRNA isoform and it could be used by the splicing apparatus at this intron/exon junction with a lower efficiency. This study highlights the relevance of IGF1R mRNA expression in neuroendocrine tumor cells, and the constant presence of 'subtle' alternative splicing for the IGF1R locus.