Kang DH, et al. Immune Netw. 2020 May;20(3):e27. https://doi.org/10.4110/in.2020.20.e27

Background: Immune checkpoint inhibitors (ICIs) have become the standard of care for a variety of cancers, including non-small cell lung cancer (NSCLC). In this study, we investigated the frequency of pseudoprogression and hyperprogression in lung cancer patients treated with ICIs in the real world and aimed to discover a novel candidate marker to distinguish pseudoprogression from hyperprogression soon after ICI treatment. Methods: This study included 74 patients with advanced NSCLC who were treated with PD-1/PD-L1 inhibitors at Chungnam National University Hospital (CNUH) between January 2018 and August 2020. Chest X-rays were examined on day 7 after the first ICI dose to identify changes in the primary mass, and the response was assessed by computed tomography (CT). We evaluated circulating regulatory T (Treg) cells using flow cytometry and correlated the findings with clinical outcomes. Results: The incidence of pseudoprogression was 13.5%, and that of hyperprogression was 8.1%. On day 7 after initiation of treatment, the frequency of CD4⁺CD25⁺CD127^loFoxP3⁺ Treg cells was significantly decreased compared with baseline (P = 0.038) in patients who experienced pseudoprogression and significantly increased compared with baseline (P = 0.024) in patients who experienced hyperprogression. In the responder group, the frequencies of CD4⁺CD25⁺CD127^loFoxP3⁺ Treg cells and PD-1⁺CD4⁺CD25⁺CD127^loFoxP3⁺ Treg cells were significantly decreased 7 days after commencement of treatment compared with baseline (P = 0.034 and P < 0.001, respectively). Conclusion: Circulating Treg cells represent a promising potential dynamic biomarker to predict efficacy and differentiate atypical responses, including pseudoprogression and hyperprogression, after immunotherapy in patients with NSCLC.

The extracellular matrix (ECM) exerts physiological activity, facilitates cell-to-cell communication, promotes cell proliferation and metastasis, and provides mechanical support for tumor cells. The development of solid tumors is often associated with increased stiffness. A stiff ECM promotes mechanotransduction, and the predominant transcription factors implicated in this phenomenon are YAP/TAZ, β-catenin, and NF-κB. In this study, we aimed to investigate whether YAP is a critical mediator linking matrix stiffness and PD-L1 in lung adenocarcinoma. We confirmed that YAP, PD-L1, and Ki-67, a marker of cell proliferation, increase as the matrix stiffness increases in vitro using the lung adenocarcinoma cell lines PC9 and HCC827 cells. The knockdown of YAP decreased the expression of PD-L1 and Ki-67, and conversely, the overexpression of YAP increased the expression of PD-L1 and K-67 in a stiff-matrix environment (20.0 kPa). Additionally, lung cancer cells were cultured in a 3D environment, which provides a more physiologically relevant setting, and compared to the results obtained from 2D culture. Similar to the findings in 2D culture, it was confirmed that YAP influenced the expression of PD-L1 and K-67 in the 3D culture experiment. Our results suggest that matrix stiffness controls PD-L1 expression via YAP activation, ultimately contributing to cell proliferation.

Background: Immune checkpoint inhibitors (ICIs) have transformed treatment and have provided significant clinical benefits and durable responses for patients with advanced non-small cell lung cancer (NSCLC). However, only a small percentage of patients respond to ICI treatment, and immune-related adverse events (irAEs) leading to treatment discontinuation remain challenging. Despite the recognized need for biomarkers to predict both the efficacy of ICIs and the risk of irAEs, such biomarkers are yet to be clearly identified. Methods: In this study, we performed single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) from 33 patients with NSCLC before ICIs treatment. To validate our findings, we reanalyzed public scRNA-seq data, conducted a cytometric bead array (CBA), and supported our findings with T-cell receptor sequencing. Results: While the immune response was more pronounced in patients with a favorable prognosis, the hypoxic pathway was more prominent in patients with primary resistance. Lymphocytes such as CD8 T cells, CD4 T cells, and natural killer cells were primarily involved in these pathways, with PRF1 and GZMB expression showing strong associations with favorable prognosis. In contrast, irAEs were mainly linked to myeloid cells, such as monocytes and macrophages. As irAE severity increased, inflammation and the TNF-NFKB1 pathway were more prominent. Specifically, increased expression of IL1B, CXCL8, and CXCL2 in monocytes and TNF in macrophages was closely associated with severe irAE through involvement in these pathways. Notably, the increase of PRF1 and GZMB expression showed a close association with both a favorable prognosis and a reduced severity of irAE, which was validated through CBA analysis. Moreover, the expression of these key markers varied according to prognosis and irAE severity regardless of patient background, such as programmed death-ligand 1 expression levels, tumor histology, or prior treatment regimens. Conclusions: This study identified biological pathways and key biomarkers associated with ICI prognosis and irAE severity using PBMC samples before treatment. These findings provide a foundation for improved therapeutic strategies that enhance clinical outcomes while minimizing ICI treatment-associated risks.

Recent advancements in spatial transcriptomics (ST) have revolutionized our understanding of the lung’s cellular organization and pathological alterations. By preserving the spatial distribution of gene expression, ST reveals localized immune niches, stromal–epithelial interactions, and disease-associated transcriptional “hotspots” that cannot be captured by conventional sequencing methods alone. In lung cancer, ST-based investigations have delineated distinct tumor microenvironments between tumor cores and invasive fronts, revealing prognostically significant gene signatures and identifying subpopulations with differential responses to immunotherapy and chemotherapy. Similarly, in chronic obstructive pulmonary disease, asthma, and idiopathic pulmonary fibrosis, ST has mapped the ecosystem, including immune cells, inflammatory mediators, and fibroblast subtypes, of discrete regions within diseased lung tissue, offering mechanistic insights into disease progression and tissue remodeling. In addition, a more recent ST study provides spatial information for where drugs act within tissues. This review highlights the emerging role of spatial transcriptomics in respiratory research, demonstrating its potential to refine disease classification, elucidate mechanisms of therapeutic resistance, and inform spatially guided personalized interventions in respiratory diseases.

One major challenge associated with lung cancer organoids (LCOs) is their predominant derivation from surgical specimens of patients with early-stage lung cancer. However, patients with advanced lung cancer, who are in need of chemotherapy, often cannot undergo surgery. Therefore, there is an urgent need to successfully generate LCOs from biopsy specimens. Conventional lung biopsy techniques, such as transthoracic needle biopsy and forceps biopsy, only yield small amounts of lung tissue, resulting in a low success rate for culturing LCOs from biopsy samples. Furthermore, potential complications, like bleeding and pneumothorax, make it difficult to obtain sufficient tissue. Another critical issue is the overgrowth of normal lung cells in later passages of LCO culture, and the optimal culture conditions for LCOs are yet to be determined. To address these limitations, we attempted to create LCOs from cryobiopsy specimens obtained from patients with lung cancer (n = 113). Overall, the initial success rate of establishing LCOs from cryobiopsy samples was 40.7% (n = 46). Transbronchial cryobiopsy enables the retrieval of significantly larger amounts of lung tissue than bronchoscopic forceps biopsy. Additionally, cryobiopsy can be employed for peripheral lesions, and it is aided via radial endobronchial ultrasonography. This study significantly improved the success rate of LCO culture and demonstrated that the LCOs retained characteristics that resembled the primary tumors. Single-cell RNA sequencing confirmed high cancer cell purity in early passages of LCOs derived from patients with advanced lung cancer. Furthermore, the three-dimensional structure and intracellular components of LCOs were characterized using three-dimensional holotomography. Finally, drug screening was performed using a specialized micropillar culture system with cryobiopsy-derived LCOs. LCOs derived from cryobiopsy specimens offer a promising solution to the critical limitations of conventional LCOs. Cryobiopsy can be applied to patients with lung cancer at all stages, including those with peripheral lesions, and can provide sufficient cells for LCO generation. Therefore, we anticipate that cryobiopsy will serve as a breakthrough strategy for the clinical application of LCOs in all stages of lung cancer.