Interstitial lung diseases (ILDs) are rare and severe respiratory conditions that may ultimately result in pulmonary fibrosis (PF). The objective of this study was to present the results of molecular diagnosis of early-onset ILD (from neonates to young adults < 50 years) in a reference genetic diagnostic laboratory. DNAs from 699 index cases and 190 relatives were studied over 6 years by Sanger and/or targeted next generation sequencing of surfactant-related genes and other genes involved in early-onset ILD. Pathogenic/likely pathogenic variants were evidenced for 62 patients (8.9%). The genes most frequently involved were SFTPA2 (13/62), followed by ABCA3 (12/62) and SFTPC (10/62). Among index cases for whom precise clinical data were available (n = 542), indications associated with a high molecular diagnostic yield were pulmonary alveolar proteinosis (61.5%, 8/13; p < 0.0007); family history of ILD/PF and lung cancer (36.8%, 7/19; p = 0.0132) and newborns > 32 weeks gestation with neonatal respiratory distress (14.8%, 9/61). The proportion of positive molecular investigations culminated in two age groups over the lifespan: 23.3% (7/30) in children aged 1 to 10 years, and 18.3% (15/82) in adults aged 30 to 40 years. Over the 6-year period, 190 relatives were subjected to testing in order to perform segregation studies (n = 123) and/or predictive testing (n = 79). This study highlights the specific patient's characteristics associated with a high or low molecular diagnostic yield in clinical practice. Furthermore, it emphasises the importance of establishing a molecular diagnosis in order to provide genetic counselling to the family.

The transcription factor NK2 homeobox1 (NKX2-1), associated with brain lung thyroid syndrome, regulates the transcription of surfactant proteins, thyroglobulin (TG) and thyroid peroxidase (TPO). This study explored the pathogenicity of three NKX2-1 variants (p.(Tyr214Cys), p.(Arg165Trp) and p.(Gly147Ala)) that were identified in three infants with lethal forms of childhood interstitial lung disease. HEK293T cells were co-transfected with expression plasmids of NKX2-1 (wild-type (WT) and variants) and PAX8, along with reporter plasmids containing the promoters of SFTPB, SFTPC, TG and TPO). Protein expression was analyzed by western blotting and immunofluorescence. Luciferase assays were performed to evaluate the activation of different promoters. Surfactant protein and NKX2-1 expression were also assessed on patient lung biopsies using immunohistochemistry. All three mutant proteins exhibited nuclear localization. Protein expression was altered in the p.(Tyr214Cys) and p.(Arg165Trp) variants located in NKX2.1 homeodomain. The p.(Tyr214Cys) variant failed to transactivate the tested promoters and was associated with a lack of pro-SP-C and SP-C expression in lung biopsy whereas the p.(Arg165Trp) variant induced both gain- or loss-of-function effects on the tested promoters. Finally, the p.(Gly147Ala) variant transactivated all the promoters tested, as for the WT. Conclusion: Our results demonstrated the pathogenicity of two variants, p.(Tyr214Cys) and p.(Arg165Trp), located within the homeodomain of NKX2-1. Conversely, the p.(Gly147Ala) variant showed no pathogenic effects. To date, the p.(Tyr214Cys) variant is associated with the most severe respiratory phenotype reported for NKX2-1-related disorders. Further studies are needed to understand the specific mechanisms underlying the pathogenicity of NKX2.1 variants located in the homeodomain.

Childhood interstitial lung disease (chILD) encompasses a heterogeneous group of rare disorders characterized by respiratory distress, hypoxemia, exercise intolerance, and distinctive radiological findings. Despite the variable age of onset, these conditions often present with overlapping symptoms and variable progression, even with identical genetic mutations. Surfactant protein deficiencies fall under the category of chILD, with Surfactant Protein-C (SP-C) deficiency posing significant diagnostic challenges due to its rarity and the variable severity of clinical presentation. We present the case of a 15-year-old male from Senegal who recently arrived in Italy, presenting with severe respiratory distress and hypoxemia. The patient, born full-term, had a long history of chronic cough, recurrent respiratory distress, and poor growth since early infancy. Upon hospitalization, he tested positive for SARS-CoV-2 and exhibited signs of chronic respiratory failure and severe malnutrition. An extensive diagnostic work-up, including a chest CT scan, revealed small cystic-like air spaces and diffuse ground-glass opacities. Whole-exome sequencing confirmed the diagnosis of SP-C deficiency by identifying a heterozygous missense mutation (c.218t>C, Ile73Thr) in the third exon of the SFTPC gene. Treatment with steroids, azithromycin and hydroxychloroquine was initiated. Despite pharmacological treatments, the patient remained oxygen dependent due to the severity of this condition and required long-term bilevel non-invasive ventilatory support. This case provides insight into the natural course of untreated child, specifically SP-C deficiency, enhancing our understanding of its manifestations and progression. The lack of standardized treatments underscores the critical need for increased awareness among physicians of this rare but potentially life-threatening condition, enabling early diagnosis and timely therapeutic interventions.

Childhood interstitial lung diseases (chILD) are rare, chronic lung diseases characterized by symptoms such as tachypnea, dyspnea, hypoxemia, crackles, and diffuse parenchymal abnormalities on chest imaging. To evaluate the etiologic spectrum, clinical presentation, management, and outcomes of chILD at a Polish referral center. We retrospectively reviewed data from patients (0-18 years) diagnosed with chILD, admitted to the Department of Pediatric Pulmonology and Allergy, Medical University of Warsaw, from June 2009 to February 2024, classified according to the chILD-EU categorization system. A total of 275 patients (65.5% male) were included, with a median age at diagnosis of 13 months (range: 1-221). Persistent tachypnea of infancy (PTI)/neuroendocrine cell hyperplasia of infancy (NEHI) was the most common diagnosis (52.4%), followed by disorders related to systemic diseases (11.3%) and related to exposures (10.2%). 13.8% of diseases remained undefined. The predominant symptoms included crackles (81.5%), dyspnea (72.7%) and tachypnea (68.3%). All children underwent chest computed tomography. Bronchoscopy, genetic testing, and lung biopsy were performed in 46.2%, 34.9%, and 21.4% of cases, respectively. Most children (92.7%) received some form of treatment, including inhaled bronchodilators/steroids (68.8%), systemic steroids (26.5%), long-term macrolides (16.3%), and immunosuppressants (11.6%). Oxygen supplementation and nutritional support were required in 50.5% and 29.8% of patients, respectively. At a median follow-up of 31.5 months, 92.9% of patients achieved clinical improvement or stabilization, and 6.2% deteriorated, including seven deaths. The 5-year survival rate was 95.66%. This study highlights the significant diversity within chILD, with PTI/NEHI being the most common condition.

Persistent tachypnea of infancy (PTI) or neuroendocrine cell hyperplasia of infancy (NEHI) is a form of childhood interstitial lung disease (chILD) that predominantly affects young children. Although it is one of the most common chILDs, no unified diagnostic approach specific to this condition exists. Are the clinical presentation and the diagnostic approach different in patients with PTI/NEHI among European countries? This was a European multicenter, retrospective, observational study. Data on clinical characteristics and diagnostic strategies in patients with PTI/NEHI were analyzed and compared across participating countries. The study included 378 children with PTI/NEHI from 17 countries (63.5% male, 97.4% White) who received a diagnosis at a median age of 9 months (interquartile range, 6-13 months). The most common baseline symptoms were tachypnea, chest retractions, crackles on auscultation, hypoxemia, and failure to thrive. High-resolution CT (HRCT) imaging was performed in all patients, with most undergoing chest radiography, echocardiography, and immunology tests. Lung biopsy was carried out in 23.5% of patients, with a decreasing trend over time and variation by country; its use was associated with longer diagnostic delay. Histopathologic examination showed a hyperplasia of pulmonary neuroendocrine cells in 52.8% of patients. Genetic testing was rare, and its application varied significantly among countries. Additional investigations that do not have an established role, such as assessment for gastroesophageal reflux disease and OSA, infant pulmonary function tests, and lung ultrasound, were limited to single countries. Diagnosis of PTI/NEHI relies on clinical symptoms and HRCT imaging results, with lung biopsies less commonly performed. Differences exist among countries regarding the number and type of investigations. A need exists for guidelines that will standardize the diagnostic approach.