The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target

Upregulation of CD11b on Eosinophils in Aspirin Induced Asthma

Background: Although a challenge test using non-steroidal anti-inflammatory drugs (NSAIDs) is crucial for diagnosis of aspirin-induced asthma (AIA), it also has drawbacks in terms of possible side effects. Therefore, alternative in-vitro diagnostic methods for AIA are awaited. Methods: Nineteen stable non-AIA patients (9 males and 10 females; mean age, 49.4 ± 4.8 years), and 20 AIA patients (9 males and 11 females; mean age, 51.1 ± 4.8 years) were enrolled in this study. CD11b and CD16 expressions on the peripheral-blood granulocytes after administration of aspirin and different concentrations of PGE2 in vitro were examined using flowcytometry. Results: Aspirin induced a significant increase in CD11b expression on eosinophils (CD16 negative granulocytes) in 19 AIA patients and one non-AIA patient. Increase in CD11b expression on eosinophils by aspirin administration was suppressed by PGE2 in a dose-dependent manner. Conclusions: The measurement of CD11b expression on peripheral-blood eosinophils showed very high sensitivity and specificity of (−95%) in diagnosing AIA. Although this method requires laboratory facilities for flowcytometry, it may be very useful in diagnosis of AIA without side effects. In addition, PGE2 may be involved in regulation of CD11b expression on eosinophils by aspirin administration

Pharmacokinetics of Gefitinib in a Patient with Non-Small Cell Lung Cancer Undergoing Continuous Ambulatory Peritoneal Dialysis

A 72-year-old man undergoing continuous ambulatory peritoneal dialysis (CAPD) for chronic renal failure and who had undergone right upper lobectomy for lung adenocarcinoma (pT2aN0M0) 2 years ago was admitted for recurrence of lung cancer presenting as multiple brain metastases. An epidermal growth factor receptor mutation analysis of his lung cancer revealed a deletion of 15 nucleotides (E746-A750) in exon 19. After whole-brain radiotherapy, we started daily administration of 250 mg gefitinib under the continuation of CAPD and performed a pharmacokinetic analysis. We speculated that the plasma concentration of gefitinib reached the steady state at least by day 16 after the start of gefitinib (626.6 ng/ml at trough level). On day 46, the plasma concentration was 538.4 ng/ml at trough level and the concentration in the peritoneal dialysis fluid was 34.6 ng/ml, suggesting that CAPD appeared to have little effect on the pharmacokinetics of gefitinib. During gefitinib therapy, there were no significant adverse events except for grade 2 diarrhea. Gefitinib could be safely administered to a patient undergoing CAPD

Flow cytometric analysis of lymphocyte profiles in mediastinal lymphadenopathy of sarcoidosis.

Lymphocyte profiles in mediastinal lymph nodes may reflect the immune status of patients with sarcoidosis. Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is useful for the diagnosis of diseases with mediastinal lymphadenopathy including sarcoidosis. The purpose of this study was to determine lymphocyte profiles of lymph nodes in sarcoidosis by analyzing EBUS-TBNA samples. We prepared single cell suspensions from EBUS-TBNA samples of mediastinal lymph nodes from patients with sarcoidosis or lung cancer and analyzed surface markers (CD3, CD4, CD8, CD19, CD25) and FoxP3 expression in the resultant lymphocytes using flow cytometry. We studied 26 patients with sarcoidosis and 16 with lung cancer with mediastinal lymph node metastases. In sarcoidosis, the CD4/CD8 ratio was significantly more elevated in lymph nodes than in bronchoalveolar lavage fluid (P<0.001), although both were strongly correlated. The CD4/CD8 ratio was significantly higher in stage I than in stage II both in the BAL fluid and lymph nodes. When compared with lung cancer lymph node metastasis, the CD4/CD8 ratio was significantly higher in sarcoidosis, whereas the CD3/CD19 ratio was significantly higher in lung cancer. The proportion of regulatory T cells (CD4+, CD25+, FoxP3 high) did not differ between sarcoidosis and lung cancer samples. Lymphocyte profiles in mediastinal lymphadenopathy can be analyzed by flow cytometry of EBUS-TBNA samples. These findings might help elucidate the immunopathology of sarcoidosis

Feasibility of tissue re-biopsy in non-small cell lung cancers resistant to previous epidermal growth factor receptor tyrosine kinase inhibitor therapies

Abstract Background When epidermal growth factor receptor (EGFR) gene mutation-positive non-small cell lung cancer (NSCLC) acquires resistance to the initial tyrosine kinase inhibitor (TKI) treatment, reassessing the tumor DNA by re-biopsy is essential for further treatment selection. However, the process of TKI-sensitive tumor re-progression and whether re-biopsy is possible in all cases of acquired resistance to EGFR-TKI remain unclear. Methods We retrospectively analyzed data from 69 consecutive patients with EGFR gene mutation-positive advanced NSCLC who had been treated with EGFR-TKI and exhibited disease relapse after initial disease remission. The relapsing lesions were identified at the time of RECIST-progressive disease (PD) and clinical-PD (when the attending physician judged the patient as clinically relapsing and stopped EGFR-TKI therapy). We determined the potential re-biopsy methods for each relapsing lesion and evaluated their feasibility according to difficulty and invasiveness criteria as follows: category A, accessible by conventional biopsy techniques; category B, difficult (but possible) to biopsy and accessible with invasive methods; and category C, extremely difficult to biopsy or inaccessible without using highly invasive methods, including surgical biopsy. Results The total feasibility rate of re-biopsy (category A or B) was 68% at RECIST-PD and 84% at clinical-PD, and the most common accessible relapsing lesions were primary tumors at RECIST-PD and pleural effusion at clinical-PD. All relapsing lesions at primary sites (categories A and B) were assessed as having the potential for re-biopsy. However, re-biopsy for metastasis was assessed as difficult in a substantial proportion of the study population (42 and 20% category C at RECIST-PD and clinical-PD, respectively). Conclusions Re-biopsy of relapsing disease is feasible in many cases, although it may present difficulties in cases with, e.g., metastatic relapsing lesions. To facilitate treatment strategies in NSCLC patients with relapse after EGFR-TKI therapy, re-biopsy should be standardized with the use of simpler and more reliable methods

A simple method of bronchial occlusion with silicone spigots (Endobronchial Watanabe Spigot; EWS) using a curette

Background: Bronchial occlusion with an Endobronchial Watanabe Spigot (EWS) has been shown to be useful in managing prolonged bronchopleural fistulas and intractable hemoptysis. EWS bronchial occlusion using a curette is less technically demanding. This retrospective study evaluated the clinical utility and simplicity of this method. Methods: A total of 18 consecutive patients (15 men, 3 women, aged 47–85 years) who underwent bronchial occlusion using an EWS from April 2012 to August 2014 were evaluated. The method involves sticking the tip of a curette into an EWS to the first joint, allowing it to be turned in any direction or at any angle. The time required to occlude the target bronchus was measured on routinely recorded digital videos. Other parameters evaluated included success rates, complications, and clinical outcomes. Results: Of the 18 patients, 11 underwent bronchial occlusion for intractable pneumothorax, 5 for postoperative bronchopleural fistula, two for intractable empyema, and one for hemoptysis. Each patient required 1–7 EWSs (median 4). Target bronchi included the right upper ( n = 8), left upper ( n = 5), right lower ( n = 2), left lower ( n = 2), and right middle ( n = 1) bronchi. The success rate of EWS insertion into the target bronchus was 100%. Time per EWS occlusion ranged from 65–528 sec (median 158.5 sec). Of the 62 insertions, 36 (58.1%) were completed within 3 min, and 58 (93.5%) within 5 min. Successful outcomes were observed in 15 (83.3%) of the 18 patients. Conclusions: EWS bronchial occlusion using a curette is a simple method for managing intractable bronchopleural fistulas in daily clinical settings