High Shear Stress Reduces ERG Causing Endothelial-Mesenchymal Transition and Pulmonary Arterial Hypertension.

Pathological high shear stress (HSS, 100 dyn/cm 2 ) is generated in distal pulmonary arteries (PA) (100-500 µm) in congenital heart defects and in progressive PA hypertension (PAH) with inward remodeling and luminal narrowing. Human PA endothelial cells (PAEC) were subjected to HSS versus physiologic laminar shear stress (LSS, 15 dyn/cm 2 ). Endothelial-mesenchymal transition (EndMT), a feature of PAH not previously attributed to HSS, was observed. H3K27ac peaks containing motifs for an ETS-family transcription factor (ERG) were reduced, as was ERG-Krüppel-like factors (KLF)2/4 interaction and ERG expression. Reducing ERG by siRNA in PAEC during LSS caused EndMT; transfection of ERG in PAEC under HSS prevented EndMT. An aorto-caval shunt was preformed in mice to induce HSS and progressive PAH. Elevated PA pressure, EndMT and vascular remodeling were reduced by an adeno-associated vector that selectively replenished ERG in PAEC. Agents maintaining ERG in PAEC should overcome the adverse effect of HSS on progressive PAH.

An increased relative eosinophil count as a predictive dynamic biomarker in non-small cell lung cancer patients treated with immune checkpoint inhibitors.

BACKGROUND: An increased relative eosinophil count (REC) has potential as a predictive biomarker for a beneficial clinical response and outcome to cancer immunotherapies. Therefore, the present study investigated the impact of an increased posttreatment REC on the prognosis of non-small cell lung cancer (NSCLC) patients treated with immune checkpoint inhibitors (ICIs). METHODS: We retrospectively reviewed all 151 patients diagnosed with NSCLC and treated with ICI monotherapy and blood test data between March 2016 and August 2021 at National Hospital Organization Kochi Hospital and Tokushima University. RESULTS: A total of 151 patients with a mean age of 69 years were included. REC after 4 weeks of initial ICI monotherapy was higher than pretreatment REC in 87 patients but not in 64. REC after 4 weeks of the ICI treatment with and without an increased REC were 4.4 and 1.8%, respectively (p < 0.001). Disease control rates (DCR) were significantly higher in patients with than in those without an increased REC (84% vs. 47%, p < 0.001). The median overall survival (OS) of lung cancer patients with or without an increased REC were 674 and 234 days, respectively. A Kaplan-Meier univariate analysis revealed a significant difference in OS between the two groups (p < 0.001). A Cox proportional regression analysis identified an increased REC as an independent predictor of OS (p = 0.003). CONCLUSION: ICI-treated NSCLC patients with an increased REC after 4 weeks of treatment had a better DCR and prognosis than the other patients examined.

Potential of fluoropyrimidine to be an immunologically optimal partner of immune checkpoint inhibitors through inducing immunogenic cell death for thoracic malignancies.

BACKGROUND: Immune checkpoint inhibitors (ICIs) are a revolutionary paradigm in the treatment of thoracic malignancies and chemoimmunotherapy is a current standard care in this field. Chemotherapeutic agents are known to induce not only direct cytotoxic effects on tumor cells but also immune modulating effects, such as stimulating immunogenic cell death (ICD). Currently, either pemetrexed (PEM) or taxane plus platinum are combined with ICIs for patients with non-small cell lung cancer (NSCLC); however, it is still unknown whether these agents are immunologically optimal partners for ICIs. METHODS: To determine the immunologically optimal chemotherapeutic agent, we first evaluated the ability of several chemotherapeutic agents, including platinum, PEM, taxane, and 5-fluorouracil (5-FU) to induce ICD using several thoracic tumor cell lines in vitro. ICD was evaluated by the cell surface expression of calreticulin (CRT) and adenosine-triphosphate (ATP) secretion. We further performed an antitumor vaccination assay in vivo. RESULTS: 5-FU induced cell surface expression of CRT and ATP secretion most efficiently among the several chemotherapeutic agents. This effect was enhanced when it was combined with platinum. In the antitumor vaccination assay in vivo, we found that vaccination with dying-AB1-HA (a murine malignant mesothelioma cell line) cells treated with 5-FU, but neither PEM nor PTX, reduced the tumor growth of living-AB1-HA cells inoculated 1 week after vaccination by recruiting CD3+ CD8+ T cells into the tumor microenvironment. CONCLUSION: Our findings indicate that fluoropyrimidine can be an immunologically optimal partner of ICIs through the induction of ICD for thoracic malignancies.

Antipodoplanin antibody enhances the antitumor effects of CTLA-4 blockade against malignant mesothelioma by natural killer cells.

Combination immunotherapy with multiple immune checkpoint inhibitors (ICIs) has been approved for various types of malignancies, including malignant pleural mesothelioma (MPM). Podoplanin (PDPN), a transmembrane sialomucin-like glycoprotein, has been investigated as a diagnostic marker and therapeutic target for MPM. We previously generated and developed a PDPN-targeting Ab reagent with high Ab-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). However, the effects of anti-PDPN Abs on various tumor-infiltrating immune cells and their synergistic effects with ICIs have remained unclear. In the present study, we established a novel rat-mouse chimeric anti-mouse PDPN IgG2a mAb (PMab-1-mG2a ) and its core-fucose-deficient Ab (PMab-1-mG2a -f) to address these limitations. We identified the ADCC and CDC activity of PMab-1-mG2a -f against the PDPN-expressing mesothelioma cell line AB1-HA. The antitumor effect of monotherapy with PMab-1-mG2a -f was not sufficient to overcome tumor progression in AB1-HA-bearing immunocompetent mice. However, PMab-1-mG2a -f enhanced the antitumor effects of CTLA-4 blockade. Combination therapy with anti-PDPN Ab and anti-CTLA-4 Ab increased tumor-infiltrating natural killer (NK) cells. The depletion of NK cells inhibited the synergistic effects of PMab-1-mG2a -f and CTLA-4 blockade in vivo. These findings indicated the essential role of NK cells in novel combination immunotherapy targeting PDPN and shed light on the therapeutic strategy in advanced MPM.

Suspected Aspergillus nodule histologically consistent with pulmonary nodular lymphoid hyperplasia:a case report.

Aspergillus nodules (AN) are an unusual form of chronic pulmonary aspergillosis. On the other hand, pulmonary nodular lymphoid hyperplasia (PNLH) is classified as a reactive pulmonary lymphoproliferative disorder. A 65-year-old male was referred to our hospital due to a nodule in the left upper lobe. Histologically, a mixture of prominent lymphoid follicular formation, and hyaline necrosis were observed. Grocott staining revealed morphological forms of Aspergillus spp. in the necrosis. The final clinical diagnosis was suspected AN histologically consistent with PNLH. This case suggests that there may be PNLH cases in which local infection with Aspergillus contributes to its pathophysiology. J. Med. Invest. 70 : 499-502, August, 2023.

Reduced FOXF1 links unrepaired DNA damage to pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a progressive disease in which pulmonary arterial (PA) endothelial cell (EC) dysfunction is associated with unrepaired DNA damage. BMPR2 is the most common genetic cause of PAH. We report that human PAEC with reduced BMPR2 have persistent DNA damage in room air after hypoxia (reoxygenation), as do mice with EC-specific deletion of Bmpr2 (EC-Bmpr2-/-) and persistent pulmonary hypertension. Similar findings are observed in PAEC with loss of the DNA damage sensor ATM, and in mice with Atm deleted in EC (EC-Atm-/-). Gene expression analysis of EC-Atm-/- and EC-Bmpr2-/- lung EC reveals reduced Foxf1, a transcription factor with selectivity for lung EC. Reducing FOXF1 in control PAEC induces DNA damage and impaired angiogenesis whereas transfection of FOXF1 in PAH PAEC repairs DNA damage and restores angiogenesis. Lung EC targeted delivery of Foxf1 to reoxygenated EC-Bmpr2-/- mice repairs DNA damage, induces angiogenesis and reverses pulmonary hypertension.

Pretreatment eosinophil counts as a predictive biomarker in non-small cell lung cancer patients treated with immune checkpoint inhibitors.

BACKGROUND: The peripheral blood eosinophil count prior to treatment has potential as a predictive biomarker for a beneficial clinical response to cancer immunotherapies. Therefore, the present study investigated the impact of the eosinophil count on overall survival (OS) in non-small cell lung cancer (NSCLC) patients treated with immune checkpoint inhibitors (ICI). METHODS: We retrospectively reviewed all patients diagnosed with NSCLC and treated with ICI monotherapy between March 2016 and August 2021 at National Hospital Organization Kochi Hospital and Tokushima University. RESULTS: A total of 166 patients were included. Fifty-five patients had an eosinophil count of less than 100 cells/µL (Eo < 100). Nighty-eight patients had an eosinophil count of 100 cells/µL or more, but less than 500 cells/µL (100 ≤ Eo < 500). Thirteen patients had an eosinophil count of 500 cells/µL or more (Eo ≥500). The median OS of all lung cancer patients was 476 days. The median OS of lung cancer patients with Eo <100, 100 ≤ Eo <500, and Eo ≥500 was 339, 667, and 143 days, respectively. A Kaplan-Meier univariate analysis showed a significant difference in OS between these three groups (p < 0.001). A Cox proportional regression analysis identified 100 ≤ Eo <500 (p = 0.04), ECOG PS score ≥ 2 (p = 0.02), tumor size ≥5 cm (p = 0.02), and PD-L1 ≥ 1% (p = 0.01) as independent predictors of OS. CONCLUSION: OS was significantly longer in ICI-treated NSCLC patients with a pretreatment eosinophil count of 100 ≤ Eo <500 than in the other patients and, thus, has potential as a new predictive biomarker.

Slowly Expanding 18 F-FDG PET-Positive Irregular Opacities in the Lung Due to Diffuse Lymphoid Hyperplasia Preceding Rheumatoid Arthritis.

ABSTRACT: Pulmonary diffuse lymphoid hyperplasia (DLH), a nonneoplastic lymphoproliferative disorder (LPD), is extremely rare, and no PET/CT findings have been reported for pulmonary DLH. We observed slowly expanding irregular opacities with 18 F-FDG accumulation (SUV max , 3.64) in the right lower lobe of a 51-year-old asymptomatic man. The patient underwent video-assisted thoracoscopic biopsy on suspicion of malignant lesions. Histologically, no neoplastic cells were present, and the lesion was consistent with DLH. Six months later, the patient developed rheumatoid arthritis. DLH should be considered in the differentiation of PET-positive irregular opacities, even in the absence of known immune abnormalities.

Identification of fibrocyte cluster in tumors reveals the role in antitumor immunity by PD-L1 blockade.

Recent clinical trials revealed that immune checkpoint inhibitors and antiangiogenic reagent combination therapy improved the prognosis of various cancers. We investigated the roles of fibrocytes, collagen-producing monocyte-derived cells, in combination immunotherapy. Anti-VEGF (vascular endothelial growth factor) antibody increases tumor-infiltrating fibrocytes and enhances the antitumor effects of anti-PD-L1 (programmed death ligand 1) antibody in vivo. Single-cell RNA sequencing of tumor-infiltrating CD45+ cells identifies a distinct "fibrocyte cluster" from "macrophage clusters" in vivo and in lung adenocarcinoma patients. A sub-clustering analysis reveals a fibrocyte sub-cluster that highly expresses co-stimulatory molecules. CD8+ T cell-costimulatory activity of tumor-infiltrating CD45+CD34+ fibrocytes is enhanced by anti-PD-L1 antibody. Peritumoral implantation of fibrocytes enhances the antitumor effect of PD-L1 blockade in vivo; CD86-/- fibrocytes do not. Tumor-infiltrating fibrocytes acquire myofibroblast-like phenotypes through transforming growth factor ß (TGF-ß)/small mothers against decapentaplegic (SMAD) signaling. Thus, TGF-ßR/SMAD inhibitor enhances the antitumor effects of dual VEGF and PD-L1 blockade by regulating fibrocyte differentiation. Fibrocytes are highlighted as regulators of the response to programmed death 1 (PD-1)/PD-L1 blockade.

Non-small cell lung cancer with EGFR (L858R and E709X) and CNNB1 mutations responded to afatinib.

Lung cancer with complex epidermal growth factor receptor (EGFR) and CTNNB1 comutations is rare, and the efficacy of tyrosine kinase inhibitors (TKIs) is generally poor. Here, we encountered a lung cancer patient with complex EGFR (L858R and E709X) and CTNNB1 comutations who successfully responded to afatinib. A 78-year-old woman visited our hospital with a cough and bloody sputum that had worsened over the past year. She had multiple mass shadows in both lungs and nodular shadows in the bronchi. The patient was diagnosed with lung adenocarcinoma cT4N3M1c stage IVB. A genetic analysis of the primary tumor using the Oncomine Dx target test multi-CDx system revealed positivity for EGFR (L858R and E709X) and CTNNB1 mutations. The expression of programmed death ligand 1 (22C3 clones) in tumor cells was negative by immunostaining. The patient was treated with afatinib as first-line therapy and achieved clinical improvement and a partial response and is continuing treatment 1 year later. Case reports of lung cancer patients with EGFR/CTNNB1 comutations are rare, and TKIs are not considered to be effective. We herein present the first case report of lung cancer with the co-occurrence of uncommon and complex EGFR (L858R and E709X) and CTNNB1 mutations that was successfully treated with afatinib.

Late-onset acute type 1 diabetes mellitus 7 months after discontinuation of pembrolizumab against lung cancer.

Immune-related adverse events (irAEs) occur in rare cases, even after the completion of immune checkpoint inhibitor (ICI) therapy. We encountered a lung cancer patient diagnosed with acute-onset type 1 diabetes mellitus (DM) 7 months after the cessation of ICI. A 68-year-old woman was referred to our hospital for chest abnormalities. She was diagnosed with lung adenocarcinoma cT4N2M1c, stage IVB. Immunostaining showed that the expression of programmed death ligand 1 in tumor cells was negative. A genetic analysis using the Oncomine Dx Target Test Multi-CDx System revealed that the primary tumor was positive for ERBB2. Combined immunotherapy with carboplatin, pemetrexed, and pembrolizumab was performed as first-line therapy, followed by maintenance therapy with pemetrexed plus pembrolizumab, which was successful. After the seventh course, maintenance therapy was stopped because only the primary tumor showed local enlargement. Local chest radiotherapy (66 Gy/33 Fr) was performed, and the patient was followed up. HbA1c was 4.9% 3 months after the completion of pembrolizumab, and dry mouth and polyuria occurred after 5 months. Seven months later, the patient developed diabetic ketoacidosis with a blood glucose of 348 mg/dL and an HbA1c of 11.3%. Antiglutamic acid decarboxylase antibodies were negative and urinary C-peptide was 9.3 µg/day. The patient was diagnosed with acute-onset type 1 diabetes and received insulin therapy. There has been no case report of type 1 diabetes diagnosed 7 months after the last administration of an ICI. These results indicate that irAE needs to be considered even after the cessation of ICI.

Solitary Ground-Glass Nodule Mimicking Lung Cancer due to Focal Progression of Usual Interstitial Pneumonia.

Idiopathic pulmonary fibrosis is often associated with lung cancer, but early malignant lesions mixed with fibrous lesions are not always easy to diagnose. A 78-year-old woman was referred to our hospital due to a ground-glass nodule in the left upper lobe detected on chest high resolution computed tomography during follow-up of chronic idiopathic interstitial pneumonia. Pathological examination of the resected specimen revealed that the ground-glass nodule was locally progressed usual interstitial pneumonia (UIP). It should be noted that focal progression of UIP may occur and present with ground-glass nodule mimicking lung cancer, even if lesions in other areas remain unchanged. Moreover, in such cases, recognition of nodular lesions by the gross findings on the pleural surface and palpation during surgical resection are difficult and require precise marking.

Early mortality factors in immune checkpoint inhibitor monotherapy for advanced or metastatic non-small cell lung cancer.

PURPOSE: Immune checkpoint inhibitors (ICI) are a promising treatment, but may cause hyperprogressive disease and early death. The present study investigated early mortality factors in ICI monotherapy for lung cancer. PATIENTS AND METHODS: We retrospectively reviewed all patients diagnosed with advanced or metastatic non-small cell lung cancer (NSCLC) and treated with ICI monotherapy (nivolumab, pembrolizumab, and atezolizumab) between March 2016 and August 2021 at National Hospital Organization Kochi Hospital and Tokushima University. Early death was defined as patients who died within 60 days of ICI treatment. RESULTS: A total of 166 patients were included. The majority of patients (87%) had an Eastern cooperative oncology group (ECOG) Performance status (PS) of 0/1. There were 21 early deaths. Significant differences were observed in ECOG PS, the histological type, liver metastasis, tumor size, the white blood cell count, neutrophils (%), lymphocytes (%), the neutrophil-to-lymphocyte ratio in serum (sNLR), C-reactive protein (CRP), and albumin between the groups with or without early death. Univariate logistic regression analyses identified ECOG PS score ≥ 2, liver metastasis, tumor size ≥ 5 cm, neutrophils ≥ 69%, lymphocytes < 22%, sNLR ≥ 4, CRP ≥ 1 mg/dl, and albumin < 3.58 g/dl as significant risk factors for early death. A multivariate logistic regression analysis revealed that liver metastasis (Odds ratio [OR], 10.3; p = 0.008), ECOG PS score ≥ 2 (OR, 8.0; p = 0.007), and a smoking history (OR, 0.1; p = 0.03) were significant risk factors for early death. CONCLUSION: Liver metastases, ECOG PS score ≥ 2, and a non-smoking history are early mortality factors in ICI monotherapy for advanced or metastatic NSCLC.

Classifications of moderate to severe asthma phenotypes in Japan and analysis of serum biomarkers: A Nationwide Cohort Study in Japan (NHOM Asthma Study).

BACKGROUND: Asthma is a heterogeneous disease, and phenotyping can facilitate understanding of disease pathogenesis and direct appropriate asthma treatment. This nationwide cohort study aimed to phenotype asthma patients in Japan and identify potential biomarkers to classify the phenotypes. METHODS: Adult asthma patients (n = 1925) from 27 national hospitals in Japan were enrolled and divided into Global Initiative for Asthma (GINA) steps 4 or 5 (GINA 4, 5) and GINA Steps 1, 2, or 3 (GINA 1-3) for therapy. Clinical data and questionnaires were collected. Biomarker levels among GINA 4, 5 patients were measured. Ward's minimum variance hierarchical clustering method and tree analysis were performed for phenotyping. Analysis of variance, the Kruskal-Wallis, and chi-square tests were used to compare cluster differences. RESULTS: The following five clusters were identified: 1) late-onset, old, less-atopic; 2) late-onset, old, eosinophilic, low FEV1; 3) early-onset, long-duration, atopic, poorly controlled; 4) early-onset, young, female-dominant, atopic; and 5) female-dominant, T1/T2-mixed, most severe. Age of onset, disease duration, blood eosinophils and neutrophils, asthma control questionnaire Sum 6, number of controllers, FEV1, body mass index (BMI), and hypertension were the phenotype-classifying variables determined by tree analysis that assigned 79.5% to the appropriate cluster. Among the cytokines measured, IL-1RA, YKL40/CHI3L1, IP-10/CXCL10, RANTES/CCL5, and TIMP-1 were useful biomarkers for classifying GINA 4, 5 phenotypes. CONCLUSIONS: Five distinct phenotypes were identified for moderate to severe asthma and may be classified using clinical and molecular variables (Registered in UMIN-CTR; UMIN000027776.).

S-1 eliminates MDSCs and enhances the efficacy of PD-1 blockade via regulation of tumor-derived Bv8 and S100A8 in thoracic tumor.

Myeloid-derived suppressor cells (MDSCs) have been known to play a pivotal role in the induction of immune tolerance, which limits the benefits of immune checkpoint inhibitors (ICIs). Recent studies revealed that several chemotherapeutic agents decreased tumor-infiltrating MDSCs. Therefore, combination therapy with cytotoxic chemotherapeutic agents and ICIs was approved for first-line treatment for lung cancer. However, the impact of chemotherapeutic agents on MDSCs and an optimal partner of ICIs has not been fully investigated in thoracic tumors, including lung cancer and malignant pleural mesothelioma. In the present study, we found that treatment with 5-FU and its oral formulation, S-1, suppressed tumor progression and inhibited the accumulation of MDSCs in thoracic tumor-bearing mice. Tumor-infiltrating T cells and dendritic cells were significantly expanded in S-1-treated mice. 5-FU suppressed the ability of tumor cells to recruit MDSCs, while it did not suppress the survival and differentiation of mouse MDSCs in vitro. We also revealed that 5-FU or S-1 significantly downregulated the expression of tumor-derived Bv8 and S100A8. The knockdown of Bv8 or S100A8 in tumor cells suppressed tumor growth and MDSC recruitment in vivo. Furthermore, in comparison with pemetrexed, administration of S-1 improved the synergistic therapeutic efficacy of anti-PD-1 antibodies with or without carboplatin. Our findings revealed a novel mechanism wherein S-1 primed a favorable tumor microenvironment to provide the rationale for combination therapy with S-1 and ICIs as the optimal therapy for thoracic cancer.

Transient and Recurrent Pulmonary Infiltrations Associated with Familial Mediterranean Fever.

Chest symptoms and pleural effusion due to serositis in familial Mediterranean fever (FMF) are occasionally misdiagnosed as acute pneumonia. However, the actual pulmonary involvement of FMF is extremely rare. A 67-year-old man was referred to our hospital due to repeated and transient anterior chest pain. Chest images revealed a moderate amount of pericardial fluid, slight bilateral pleural effusion, and infiltrations in both lower lung lobes. Colchicine treatment without antibiotics rapidly improved these symptoms and findings. Pericarditis, pleurisy and the response to colchicine indicated FMF. FMF should be considered as a causative disease of pulmonary infiltrations, especially if it occurs repeatedly.

Surgical resection combined with perioperative chemotherapy for a patient with locally recurrent, previously stage IV thymic small-cell carcinoma: A case report.

An 83-year-old Japanese man visited our hospital with dyspnea and general fatigue. Computed tomography (CT) revealed a tumor in the anterior mediastinum, bilateral pleural effusion, pericardial fluid, and multiple liver nodules. We performed a CT-guided tumor biopsy, and the patient was diagnosed with thymic small-cell carcinoma, Masaoka-Koga stage classification IVb. The patient received four cycles of carboplatin and etoposide, and all lesions disappeared on CT. However, after 6 months, CT revealed a recurrent tumor in the anterior mediastinum. After one cycle of rechallenge chemotherapy, we performed extended total thymectomy followed by another three cycles of chemotherapy. More than 2.5 years after the last chemotherapy session, the patient's carcinoma did not recur. Thus, this case suggests that salvage surgery may be a treatment option for local recurrence of thymic carcinoma after complete remission with chemotherapy, even in patients with stage IV cancer.