Clinical Perspectives

Computational Mass Spectrometry Drives Translational Discoveries in Lung Cancer

May 14, 2020

Protomics Lab

In recent years, new therapy options for the treatment of advanced lung cancer have been realized through better understanding of its molecular underpinnings. In adenocarcinoma of the lung (ADC), identification of somatic gene mutations, gene amplifications, or gene fusions of oncogenes such as receptor tyrosine kinases has facilitated development of targeted agents with small molecule kinase inhibitors or antibody therapies. Mutations in the epidermal growth factor receptor (EGFR) or rearrangements creating fusion proteins of the EML4 and ALK tyrosine kinase now enable treatments with either EGFR or ALK tyrosine kinase inhibitors and improved patient outcomes 1-3. However, little inroads in targeted therapy have been made for squamous cell lung cancer (SCC), despite initial enthusiasm about targets including EGFR, fibroblast growth factor receptors (FGFR), discoidin domain receptors (DDR2), and phosphatidylinositide 3-kinases (PI3K) 4. In contrast, use of immune checkpoint antibody therapy has demonstrated durable tumor regressions in both ADC and SCC histologies with prolonged survival. This has led to approval of multiple antibodies targeting the PD-1/PD-L1 interaction for patients with advanced disease and now provides an alternative therapy beyond conventional cytotoxic chemotherapy for patients with advanced SCC 5-8.

Genomic technologies have provided important insights into the molecular underpinnings of SCC. The Cancer Genome Atlas (TCGA) identified recurrent mutations in genes associated with cell cycle and apoptosis (TP53, CDKN2A, RB1), antioxidant gene expression (NFE2L2, KEAP1), phosphatidylinositide 3-kinase signaling (PIK3CA, PTEN), and epigenetic signaling (MLL2) 9. This study also found high level changes in chromosome gain and loss associated with severe genomic instability. Subclasses of SCC have also been defined using transcriptomic data 9-11. In addition to tumor autonomous features, patterns of infiltrating immune cell types have been associated with tumor progression and patient prognosis 12,13. Based on these results, newer studies such as NCI Molecular Analysis for Therapy Choice (MATCH) are attempting to capitalize on improved molecular knowledge of SCC and are employing precision medicine to approach targets including PI3K, CDK4/6, FGFR, MET, and PD-L1.

To provide additional insights into SCC, Stewart et al. reported an integrated analysis incorporating expression proteomics analysis of protein abundance alongside DNA copy number analyses, somatic mutations, and mRNA expression via RNA sequencing in 108 surgically resected SCC (from Moffitt's Total Cancer Care® Protocol) with accompanying clinical outcome data, evaluation of tumor pathology, and other clinically relevant data 14. The incorporation of mass spectrometry-based proteomic data was a critical new addition as protein abundance can correlate poorly with corresponding mRNA abundance 15-18. The study by Stewart et al. leveraged prior deep genomic and transcriptomic studies of SCC allowing a focused examination of the SCC proteome and its relationship to previously observed genomic or transcriptomic subgroups 9,11.

Paul Stewart, PhD
Paul Stewart, PhD

In their study, Stewart et al. determined that the SCC tumors could be grouped into 3 subtypes based on their protein expression patterns, named Inflamed, Redox, and Mixed. The Inflamed subtype accounted for 40% of the cohort, and tumors in this subtype had higher levels of proteins associated with immune cells, especially neutrophils or myeloid cells, and an active inflammatory response. Based on RNA data, they discovered that the Inflamed subtype also had a high proportion of other immune cells, including memory B-cells and monocytes, and was associated with higher levels of PD-1 than the other two subtypes.

Results Inflamed, Redox, Mixed

The Redox comprised 47% of the cohort. These tumors were characterized by higher levels of proteins that are associated with oxidation-reduction cellular signaling pathways. The Redox subtype also had a higher number of genetic and chromosomal alterations that are known to be involved in SCC development. Using these data as guides, they identified new vulnerabilities that could be possible future therapeutic targets.

The final subtype, Mixed, represented only 13% of the tumors and only displayed an increased level of four proteins. The authors did not find any significant chromosomal alterations in this subtype but did learn that the mixed group had more mutations in the APC gene and had a greater infiltration of stromal cells than the other subtype.

The analysis showed that the three subtypes did not correspond to better or worse patient outcomes. However, tertiary lymph node structures, more commonly found in the Inflamed subtype, were associated with better outcomes. "These findings are in line with the general lack of agreement of prognostic signatures in SCC but now strongly suggest that an active immune response, indicated by tertiary lymph node structures, is associated with better outcomes. We hope to better understand this in future studies and determine how to exploit this knowledge for new therapy," said Eric Haura, M.D., senior author, director of the Lung Cancer Center of Excellence, and Associate Center Director of Clinical Science at Moffitt.

The team at Moffitt hopes that their results will lead to an improved understanding of SCC and highlight potential therapeutic targets for each subtype. Ongoing studies are examining metabolic targets for treatment of the Redox group. "Our results show SCC can be thought of as a disease with three subtypes, the bulk (87%) of which are associated with either immune infiltration (Inflamed) or oxidation-reduction (Redox) biology. This line of thinking is compelling, because it indicates that the majority of patients could benefit from therapies directed against immune cell types (Inflamed) or metabolic modulation of tumor intrinsic pathways (Redox)," Haura added.

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