RASopathies are a group of genetically heterogeneous developmental disorders caused by germline variants affecting the RAS/MAPK signaling pathway. These disorders display overlapping clinical features and diverse molecular mechanisms. This study aimed to evaluate the clinical and molecular spectrum of patients diagnosed with RASopathies, with a particular focus on novel and rare variants. A retrospective, multicenter study was conducted on patients with clinically suspected RASopathy and diagnosed by targeted Next Generation Sequencing analysis between 2021 and 2024. Variants were classified according to ACMG criteria, and clinical data were reviewed for genotype-phenotype correlations. Among 23 patients (14 males, 9 females), 15 (65.2%) had Noonan syndrome, five (21.7%) Neurofibromatosis Type 1, two (8.7%) Cardio-facio-cutaneous syndrome, and one (4.3%) Neurofibromatosis-Noonan syndrome. The most frequent clinical findings were craniofacial dysmorphism (91.3%), musculoskeletal anomalies (82.6%), and cutaneous features (78.3%). A total of 24 heterozygous variants were identified in seven genes: PTPN11 (45.8%), NF1 (25%), LZTR1 (12.5%), and RASA2, SOS1, MAP2K1, and BRAF (each 4.2%). Four novel variants were detected (PTPN11 c.853 + 4A>G, RASA2 p.E71D, NF1 p.W784Mfs*10, MAP2K1 p.A106T). This study highlights the clinical and molecular heterogeneity of RASopathies and expands the variant spectrum with novel and rare pathogenic alterations. The identification of new variants, particularly in rarely implicated genes such as RASA2, underlines the diagnostic value of comprehensive NGS-based testing and the need for individualized, multidisciplinary clinical management.

Data on clinical manifestations of neurofibromatosis-Noonan syndrome (NF-NS) remain heterogeneous, with limited validated descriptions. This study aims to better define the clinical and molecular features of NF-NS and compare them with existing literature. Secondary objectives include evaluating inter-rater diagnostic agreement among experienced clinicians and assessing the utility of deep-learning algorithms (Face2Gene® [F2G]). Additionally, we assess the prevalence of congenital heart malformations (CHM) in NF-NS compared to 'classic' neurofibromatosis type 1 (NF1). A 9-year, prospective, monocentric study was conducted, involving patients with NF1 pathogenic variants (PVs) and Noonan syndrome-like facial phenotype (NSLFP). Twenty-six patients were enrolled. NSLFP was categorized as 'suggestive' in 69% of cases and 'typical' in 31%. The presence of at least two facial abnormalities (e.g., low-set ears, downslanted palpebral fissures, hypertelorism, and ptosis) was consistently observed in 'typical' cases. Inter-rater concordance was substantial (0.65 [95% CI = 0.56; 0.74]), while concordance between clinicians and F2G was almost perfect at (0.821 [CI 95% = 0.625; 1.000]). Missense NF1 PVs were observed in 38.5% of cases. Apart from NSLP and a high frequency of pectus excavatum (62.5%), no significant differences in anthropometric, dermatological, neurological, skeletal, or ocular clinical features were observed between NF-NS and 'classic' NF1. CHM were found in 19.2% of NF-NS patients, with pulmonic stenosis present in 7.7%. NF-NS is a distinct phenotypic variant of NF1, marked by NSLP with consistent facial features -, and frequent pectus excavatum. F2G demonstrated high diagnostic concordance, reinforcing its clinical utility. Given the elevated risk of CHM, especially pulmonic stenosis, proactive cardiovascular assessment similar to other RASopathies is recommended for NS-NF patients, regardless of NF1 PV type.

RASopathies are among the most prevalent genetic syndromes caused by variants in the Ras/MAPK signaling pathway, affecting various systems such as the heart, craniofacial features, skin, musculoskeletal system, hearing, and vision. They can also increase the risk of secondary malignancies. Despite clinical overlaps, distinguishing features are crucial for diagnosis, as different variants lead to distinct clinical implications. This study reviews the molecular and clinical characteristics of RASopathies, focusing on neurofibromatosis type 1 (NF1) and non-NF1 RASopathies. The study analyzed 76 patients referred to our outpatient clinic over a 6-year period, all of whom were clinically diagnosed with RASopathy and confirmed in most cases by molecular testing. Patient files, clinical photographs, and laboratory results were reviewed and analyzed. A targeted multigene next-generation sequencing panel test was performed, followed by Sanger sequencing for both confirmation and segregation analysis. Multiplex ligation-dependent probe amplification was conducted in a patient with normal sequence results but strong clinical suspicion, to identify potential deletions. We identified 44 pathogenic, 25 likely pathogenic variants, and 6 variants of uncertain significance based on American College of Medical Genetics and Genomics (ACMG) criteria. Among these, 14 novel variants were found - 13 in the NF1 gene and one in SOS1. NF1 variants were present in 51 cases. Additional variants, likely to represent clinically significant findings, were identified in PTPN11 (n = 11), RAF1 (n = 4), SOS1 (n = 3), RIT1 (n = 3), KRAS (n = 1), NRAS (n = 1), SOS2 (n = 1), and BRAF (n = 1). Diagnoses included 49 patients with NF1, 21 with Noonan syndrome, 2 with neurofibromatosis-Noonan syndrome, 2 with Noonan syndrome with multiple lentigines, and 1 with cardiofaciocutaneous syndrome. Here, 12% of NF1 variants were located in exon 21, 36% of PTPN11 variants in exon 3, and 75% of RAF1 variants in exon 7. RASopathies have a broad molecular and clinical spectrum, complicating diagnosis and management. Accurate clinical correlation and molecular analysis are essential, as different RASopathy syndromes can result from variants in the same genes, while the same syndrome may arise from different genetic alterations. This study identifies novel variants and emphasizes the need for precise diagnostic approaches in these complex disorders.

Background: Noonan syndrome (NS) is a genetically heterogeneous condition within the RASopathies spectrum, with distinctive craniofacial features, congenital heart defects, short stature, and variably present developmental delay. Most cases result from variants in genes regulating the RAS/MAPK pathway, with PTPN11 variants being the most frequent; the c.922A>G substitution being among the most commonly reported. Methods: This pilot study analyzed clinical and partial genetic features of NS in a cohort from Transylvania, evaluated in the Children's Emergency Clinical Hospital in Cluj-Napoca. Thirty-one patients fulfilling the Van der Burgt diagnostic criteria (twenty-two males, nine females) were included. Clinical data were systematically reviewed, and targeted molecular testing for the PTPN11 c.922A>G variant was performed. Results: Congenital heart defects were highly prevalent, with pulmonary stenosis representing the most frequent anomaly (54.8%). Craniofacial dysmorphism was observed in 76.7% of cases, cryptorchidism in 50% of the males, and short stature below the third percentile was described in 77.4% of patients. Genetic screening identified the PTPN11 c.922A>G variant in two individuals (6.45%). Additional diagnoses included Williams-Beuren syndrome and a 17q11.2 deletion consistent with Neurofibromatosis-Noonan syndrome, underscoring the clinical and genetic heterogeneity of the cohort. Comparison with international reports highlighted variability in phenotype and variant frequency. Future research directions include Sanger sequencing of key PTPN11 exons and the application of next-generation sequencing targeting all RAS pathway genes. Conclusions: This is the first Romanian cohort study on patients with a clinical suspicion of NS, providing insight into their evaluation. The findings reinforce the need for comprehensive molecular approaches, facilitating diagnostic precision and counseling strategies.

RASopathies are genetic disorders linked to the RAS/MAPK signaling pathway, including Noonan syndrome and related conditions. These disorders have overlapping features and variable phenotypes, making diagnosis challenging. This study examines the clinical and genetic characteristics of RASopathies in pediatric patients to improve diagnostic accuracy and identify novel pathogenic variants. A total of 23 patients, who were not related to each other and were clinically diagnosed with RASopathy, participated in the study. Patients underwent next-generation sequencing (NGS) panel analyses of 14 RASopathy genes. Pathogenic variants were found in 18 out of 23 patients (78%). The most common variants were in PTPN11 and SOS1. Novel variants were identified in KRAS and NF1 expanding the known genetic spectrum. Frequent clinical features included distinctive facial features, growth delays, and heart defects, notably pulmonary stenosis. Intellectual disability was more common in patients with PTPN11 variants, while SOS1 variants were associated with unique features such as previously unreported polydactyly and choanal atresia. Targeted NGS panels improve diagnosis of RASopathies, with a variant detection rate of 78%. Including the NF1 gene, even without signs of neurofibromatosis, enhances diagnostic success. This study adds to our understanding of genotype-phenotype relationships in RASopathies and highlights new clinical features tied to SOS1 variants. Comprehensive genetic testing supports earlier and more personalized care for patients with RASopathies.