Contemporary and emerging technologies for research in children's rare and interstitial lung disease.

Although recent decades have seen the identification, classification and discovery of the genetic basis of many children's interstitial and rare lung disease (chILD) disorders, detailed understanding of pathogenesis and specific therapies are still lacking for most of them. Fortunately, a revolution of technological advancements has created new opportunities to address these critical knowledge gaps. High-throughput sequencing has facilitated analysis of transcription of thousands of genes in thousands of single cells, creating tremendous breakthroughs in understanding normal and diseased cellular biology. Spatial techniques allow analysis of transcriptomes and proteomes at the subcellular level in the context of tissue architecture, in many cases even in formalin-fixed, paraffin-embedded specimens. Gene editing techniques allow creation of "humanized" animal models in a shorter time frame, for improved knowledge and preclinical therapeutic testing. Regenerative medicine approaches and bioengineering advancements facilitate the creation of patient-derived induced pluripotent stem cells and their differentiation into tissue-specific cell types which can be studied in multicellular "organoids" or "organ-on-a-chip" approaches. These technologies, singly and in combination, are already being applied to gain new biological insights into chILD disorders. The time is ripe to systematically apply these technologies to chILD, together with sophisticated data science approaches, to improve both biological understanding and disease-specific therapy.

A pediatric case of primary ciliary dyskinesia caused by novel copy number variation in PIH1D3.

An 11-month-old boy with productive cough was referred to our hospital. He had nasal obstruction immediately after birth, and wheezing, wet cough, and rhinorrhea were observed daily after the neonatal period. Clinical and imaging findings revealed secretory otitis media, chronic sinusitis, and bronchiectasis. Primary ciliary dyskinesia was suspected. Transmission electron microscopy of nasal cilia showed defects of the outer and inner dynein arms. Genetic examinations of the family revealed copy number variation in PIH1 domain-containing 3 (PIH1D3) in the proband and mother. This is the first report of a Japanese patient with primary ciliary dyskinesia caused by copy number variation in PIH1D3.

Multicellular modeling of ciliopathy by combining iPS cells and microfluidic airway-on-a-chip technology.

Mucociliary clearance is an essential lung function that facilitates the removal of inhaled pathogens and foreign matter unidirectionally from the airway tract and is innately achieved by coordinated ciliary beating of multiciliated cells. Should ciliary function become disturbed, mucus can accumulate in the airway causing subsequent obstruction and potentially recurrent pneumonia. However, it has been difficult to recapitulate unidirectional mucociliary flow using human-derived induced pluripotent stem cells (iPSCs) in vitro and the mechanism governing the flow has not yet been elucidated, hampering the proper humanized airway disease modeling. Here, we combine human iPSCs and airway-on-a-chip technology, to demonstrate the effectiveness of fluid shear stress (FSS) for regulating the global axis of multicellular planar cell polarity (PCP), as well as inducing ciliogenesis, thereby contributing to quantifiable unidirectional mucociliary flow. Furthermore, we applied the findings to disease modeling of primary ciliary dyskinesia (PCD), a genetic disease characterized by impaired mucociliary clearance. The application of an airway cell sheet derived from patient-derived iPSCs and their gene-edited counterparts, as well as genetic knockout iPSCs of PCD causative genes, made it possible to recapitulate the abnormal ciliary functions in organized PCP using the airway-on-a-chip. These findings suggest that the disease model of PCD developed here is a potential platform for making diagnoses and identifying therapeutic targets and that airway reconstruction therapy using mechanical stress to regulate PCP might have therapeutic value.

Core-shell hydrogel microfiber-expanded pluripotent stem cell-derived lung progenitors applicable to lung reconstruction in vivo.

Lung transplantation is the only treatment available for end-stage lung diseases; however, donor shortage is a global issue. The use of human pluripotent stem cells (hPSCs) for organ regeneration is a promising approach. Nevertheless, methods for the expansion of isolated hPSC-derived lung progenitors (hLPs) for transplantation purposes have not yet been reported. Herein, we established an expansion system of hLPs based on their three-dimensional culture in core-shell hydrogel microfibers, that ensures the maintenance of their bipotency for differentiation into alveolar and airway epithelial cells including alveolar type II (AT2) cells. Further, we developed an efficient in vivo transplantation method using an endoscope-assisted transtracheal administration system; the successful engraftment and in vivo differentiation of hLPs into alveolar epithelial cells (incorporated into the alveoli) was observed. Importantly, expanded hLPs in the context of microfibers were successfully transplanted into the murine lungs, opening avenues for cell-based therapies of lung diseases. Therefore, our novel method has potential regenerative medicine applications; additionally, the high-quality hLPs and AT2 cells generated via the microfiber-based technology are valuable for drug discovery purposes.

A Phase II Study of Cisplatin Plus Gemcitabine followed by Maintenance Gemcitabine for Advanced Squamous Non-Small-Cell Lung Cancer: Kyoto Thoracic Oncology Research Group 1302.

BACKGROUND: There has been no study so far on gemcitabine continuous maintenance therapy targeting only squamous non-small-cell lung cancer (NSCLC) patients. This study aimed to assess the efficacy and safety of cisplatin plus gemcitabine followed by maintenance gemcitabine for chemotherapy- naïve Japanese patients with advanced squamous NSCLC. METHODS: The patients received 4 cycles of gemcitabine (1,000 mg/m2, days 1 and 8) and cisplatin (80 mg/m2, day 1) every 3 weeks, followed by gemcitabine alone as maintenance therapy every 3 weeks until disease progression or unacceptable toxicity. The primary end point of the study was progression-free survival (PFS) from the date of registration. RESULTS: From May 2013 to October 2018, 26 patients were enrolled, and 25 patients received ≥1 cycle of planned treatment. Eighteen patients (69.2%) received 4 cycles of cisplatin plus gemcitabine, and 16 patients (61.5%) received ≥1 cycle of maintenance gemcitabine. The median PFS from the date of registration was 5.3 months (95% CI 2.9-7.3 months). In 16 patients who received ≥1 cycle of maintenance gemcitabine, the median PFS from the date of maintenance gemcitabine initiation was 3.8 months (95% CI 2.3-5.2 months). Their median overall survival from the date of registration was 11.9 months (95% CI 7.5-26.5 months). During the maintenance therapy, adverse events (AEs) were mostly Common Terminology Criteria for AE grade 1. CONCLUSIONS: While this trial did not meet the primary endpoint, the sufficient efficacy and feasibility of gemcitabine maintenance therapy were suggested.

A phase I/II trial of weekly nab-paclitaxel for pretreated non-small-cell lung cancer patients without epidermal growth factor receptor mutations and anaplastic lymphoma kinase rearrangement.

AIM: We investigated the efficacy, safety and optimal schedule of nanoparticle albumin-bound paclitaxel monotherapy as second- or third-line treatment for non-small-cell lung cancer patients without epidermal growth factor receptor mutation and anaplastic lymphoma kinase rearrangement. METHODS: Patients with pretreated advanced non-small-cell lung cancer without epidermal growth factor receptor mutation and anaplastic lymphoma kinase rearrangement were included. The patients were administered 100 mg/m2 of nanoparticle albumin-bound paclitaxel on days 1, 8, 15 and 22 (level 0) or on days 1, 8 and 15 (level -1) every 4 weeks during phase I of the trial. The primary endpoint was objective response rate. The estimated objective response rate was 15% and the threshold was 5% with an α error of 0.05 and ß error of 0.2 in phase II. RESULTS: The recommended schedule was determined as level -1 in phase I. The characteristics of the 55 patients enrolled in phase II were as follows: median age = 66 years, male/female = 40/15, second/third line = 34/21 and adenocarcinoma/squamous cell carcinoma/large cell carcinoma/others = 34/17/2/2. Objective response rate was 7.3% (95% confidence interval, 2.0-17.6%). Median progression-free survival was 3.4 months. Treatment-related grade 3 or 4 toxicities were neutropenia (36.4%), febrile neutropenia (5.5%) and pulmonary infection (3.6%). Three patients had grade 2 pneumonitis and one treatment-related death occurred due to adult respiratory distress syndrome. CONCLUSION: This study failed to meet predefined primary endpoints for pretreated patients with advanced non-small-cell lung cancer without epidermal growth factor receptor mutation and anaplastic lymphoma kinase rearrangement.

In Vitro Disease Modeling of Hermansky-Pudlak Syndrome Type 2 Using Human Induced Pluripotent Stem Cell-Derived Alveolar Organoids.

It has been challenging to generate in vitro models of alveolar lung diseases, as the stable culture of alveolar type 2 (AT2) cells has been difficult. Methods of generating and expanding AT2 cells derived from induced pluripotent stem cells (iPSCs) have been established and are expected to be applicable to disease modeling. Hermansky-Pudlak syndrome (HPS) is an autosomal recessive disorder characterized by dysfunction of lysosome-related organelles, such as lamellar bodies (LBs), in AT2 cells. From an HPS type 2 (HPS2) patient, we established disease-specific iPSCs (HPS2-iPSCs) and their gene-corrected counterparts. By live cell imaging, the LB dynamics were visualized and altered distribution, enlargement, and impaired secretion of LBs were demonstrated in HPS2-iPSC-derived AT2 cells. These findings provide insight into the AT2 dysfunction in HPS patients and support the potential use of human iPSC-derived AT2 cells for future research on alveolar lung diseases.

Serum albumin level as a potential marker for deciding chemotherapy or best supportive care in elderly, advanced non-small cell lung cancer patients with poor performance status.

BACKGROUND: There have been few data on the chemotherapy in elderly advanced non-small cell lung cancer (NSCLC) patients with poor performance status (PS), and usefulness of chemotherapy for such patients remains unclear. The objective of this study was to identify factors that predicted the survival benefit of chemotherapy. METHODS: All consecutive elderly patients (≥75 years) with advanced NSCLC, Eastern Cooperative Oncology Group PS ≥2, EGFR mutation wild type/unknown, and newly diagnosed from January 2009 to December 2012 at a tertiary hospital were retrospectively reviewed. RESULTS: We enrolled 59 patients, and 31 patients received at least one chemotherapy regimen (chemotherapy group). However, 28 patients received best supportive care (BSC) alone (BSC group). The proportion of PS 2 and serum albumin levels was significantly higher in the chemotherapy group than in the BSC group. In the chemotherapy group, log-rank testing did not show statistically significant differences in overall survival (OS) between the single-agent therapy group and carboplatin-based doublet therapy group; however, the OS of patients receiving chemotherapy for only 1 cycle (early termination) was significantly shorter than patients receiving chemotherapy for ≥2 cycles. Hypoalbuminemia was not only a risk factor for the early termination of chemotherapy but also an independent prognostic factor in the chemotherapy group. A receiver operating characteristic curve analysis showed that the best cut-off value was 3.40 g/dL. In patients with serum albumin levels ≥3.40 g/dL, OS was significantly better in the chemotherapy group than in the BSC group (p = 0.0156), however, patients with serum albumin levels <3.40 g/dL exhibited poor prognosis regardless of the presence or absence of chemotherapy. CONCLUSION: In the elderly NSCLC patients with poor PS, serum albumin levels may help identify certain patient populations more likely to receive a survival benefit of systemic chemotherapy.

Long-term expansion of alveolar stem cells derived from human iPS cells in organoids.

The stable expansion of tissue-specific stem cells in vitro has contributed to research on several organs. Alveolar epithelial type II (AT2) cells function as tissue stem cells in the lung, but robust models for studying human AT2 cells are lacking. Here we report a method for the efficient generation and long-term expansion of alveolar organoids (AOs) harboring SFTPC+ alveolar stem cells derived from human induced pluripotent stem cells (hiPSCs). hiPSC-derived SFTPC+ cells self-renewed, with transcriptomes and morphology consistent with those of AT2 cells, and were able to differentiate into alveolar epithelial type I (AT1)-like cells. Single-cell RNA-seq of SFTPC+ cells and their progenitors demonstrated that their differentiation process and cellular heterogeneity resembled those of developing AT2 cells in vivo. AOs were applicable to drug toxicology studies recapitulating AT2-cell-specific phenotypes. Our methods can help scientists overcome the limitations of current approaches to the modeling of human alveoli and should be useful for disease modeling and regenerative medicine.

Three-Arm Randomized Trial of Sodium Alginate for Preventing Radiation-Induced Esophagitis in Locally Advanced Non-Small Cell Lung Cancer Receiving Concurrent Chemoradiotherapy: The OLCSG1401 Study Protocol.

Concurrent chemoradiotherapy (CRT) is the standard of care for locally advanced non-small cell lung cancer (LA-NSCLC). However, this intensive therapy often causes severe esophagitis, which could deteriorate a patient's quality of life (QOL), leading to poor treatment compliance. Sodium alginate, approved in Japan for gastritis, is sufficiently highly viscous to remain in the esophageal mucosa, providing a protective effect in the esophagus. To investigate whether this compound has a preventive effect against severe esophagitis in patients receiving concurrent CRT, we plan a 3-arm randomized trial of sodium alginate with 2 different schedules versus water. The primary endpoint is set as the proportion of patients with grade ≥ 3 esophagitis using the Common Terminology Criteria for Adverse Events, version 4.0. With stratification by institute, performance status, and percentage of the esophageal volume receiving >35 Gy, the patients will be randomly assigned to 1 of the following groups: sodium alginate initiated concomitantly with CRT (group A), sodium alginate initiated soon after the development of extremely mild esophagitis during CRT (group B), or water administered throughout CRT (group C). Assuming that the proportion of grade ≥ 3 esophagitis would be 8% in groups A and B and 27% in group C, the required sample size would be 200 patients, with 70% power and 5% α. The secondary endpoints include QOL, the frequency of additional prescriptions of analgesics, treatment response, and survival. The results of the present study will clarify whether sodium alginate can prevent esophagitis in patients with LA-NSCLC undergoing CRT.

18 Cases of pulmonary Mycobacterium abscessus: Clinical difference depending on the presence or absence of Mycobacterium avium complex.

BACKGROUND AND OBJECTIVES: It can be difficult to treat respiratory tract infections caused by Mycobacterium abscessus (M. abscessus) as there is no established treatment strategy. Complications involving other nontuberculous mycobacterial infections such as Mycobacterium avium complex (MAC) are also commonly observed. METHODS: We investigated the clinical background and course of 18 cases of pulmonary M. abscessus infection treated over 8 years at Kurashiki Central Hospital. Radiological evaluation was performed using NICE scoring system, a method of semi-quantitative evaluation of imaging findings of pulmonary MAC infection. RESULTS: The mean age of the 18 patients (males, 6; females, 12) was 74.7 years. The median follow-up period was 1316 days (95% confidence interval; 720-1675 days), and 11 patients were concomitantly infected with pulmonary MAC. Among the patients that underwent antibacterial treatment for M. abscessus, there was one MAC-complication case and one non-MAC-complication case. All MAC-complication cases underwent antibacterial treatment including clarithromycin. Chest X-ray NICE scores for all cases were 8.50 ± 5.45 and 10.94 ± 6.03 at baseline and follow-up, respectively (p = 0.0063). For MAC-complication cases, scores were 8.36 ± 4.74 and 12.00 ± 6.02 at baseline and follow-up, respectively (p = 0.00818), and for non-MAC-complication cases, scores were 8.71 ± 6.82 and 9.29 ± 6.13 at baseline and follow-up, respectively (p = 0.356). MAC-complication cases were significantly further exacerbated than non-MAC-complication cases (p = 0.027). CONCLUSIONS: Some cases of pulmonary M. abscessus infection progressed well without undergoing antibacterial treatment. In particular, results suggested that the clinical course of MAC-complication and non-MAC-complication cases differs.

Interstitial lung disease induced by alectinib (CH5424802/RO5424802).

A 75-year-old woman with anaplastic lymphoma kinase (ALK)-rearranged Stage IV lung adenocarcinoma was administered the selective anaplastic lymphoma kinase inhibitor, alectinib, as a third-line treatment in a Phase 1-2 study. On the 102nd day, chest computed tomography showed diffuse ground glass opacities. Laboratory data revealed high serum levels of KL-6, SP-D and lactate dehydrogenase without any clinical symptoms. There was no evidence of infection. Marked lymphocytosis was seen in bronchoalveolar lavage fluid analysis, and transbronchial lung biopsy showed mild thickening of alveolar septa and lymphocyte infiltration. Interstitial lung disease was judged to be related to alectinib based on improvements in imaging findings and serum biomarkers after discontinuation of alectinib. To our knowledge, this is the first reported case of alectinib-induced interstitial lung disease. Alectinib is a promising drug for ALK-rearranged non-small cell lung cancer. Clinical trials of this selective anaplastic lymphoma kinase inhibitor will facilitate the meticulous elucidation of its long-term safety profile.