Clinical Characteristics and Genetic Analysis of a Chinese Family with Birt–Hogg–Dubé Syndrome Harboring a Novel FLCN Gene Mutation

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Mutations in the FLCN gene are known causes, yet novel variants continue to be identified, expanding genotype–phenotype correlations. Methods We investigated a Chinese family with suspected BHDS. The proband was admitted to the Affiliated Cangnan Hospital of Wenzhou Medical University in October 2023. Comprehensive clinical evaluations and imaging studies were performed. Peripheral blood samples were collected from the proband and available family members after obtaining informed consent. Whole-exome sequencing (WES) was conducted to identify potential variants in the FLCN gene. Candidate variants were subsequently validated by Sanger sequencing and analyzed for co-segregation within the family. Pathogenicity was assessed using multiple bioinformatic prediction tools in accordance with the American College of Medical Genetics and Genomics (ACMG) guidelines. To evaluate the structural impact of the variant, a three-dimensional model of the folliculin protein was generated using SWISS-MODEL and visualized with PyMol. Results The proband presented with bilateral pulmonary cysts, a small left-sided pneumothorax, a left renal tumor, and suspected cutaneous lesions. Family screening revealed that several children had pulmonary cysts, some accompanied by pneumothorax, and two had previously undergone lobectomy. Whole-exome sequencing identified a novel heterozygous nonsense mutation in exon 11 of FLCN, c.1222C > T (p.Gln408Ter), which has not been previously reported. This mutation was also detected in one of the proband’s sons, while the proband’s husband and an asymptomatic daughter were wild type at this locus. Functional prediction indicates a loss of normal folliculin activity. Based on ACMG guidelines, the variant is classified as pathogenic. Conclusions The novel FLCN nonsense variant c .1222C > T (p.Gln408Ter) is the likely genetic basis of BHDS in this family. These findings expand the mutational spectrum of FLCN , inform genotype–phenotype relationships, and support improved diagnosis and genetic counseling of BHDS. Birt–Hogg–Dubé syndrome FLCN Genetic mutation Pedigree study Pulmonary cysts Renal tumor Figures Figure 1 Figure 2 Figure 3 Figure 4 Background Birt–Hogg–Dubé syndrome (BHDS; OMIM #135150), also known as Hornstein–Knickenberg syndrome, is a rare autosomal dominant disorder first described in 1977. It is clinically characterized by the triad of multiple pulmonary cysts with or without recurrent spontaneous pneumothorax, benign cutaneous lesions such as fibrofolliculomas or trichodiscomas, and an increased risk of renal neoplasms of various histological subtypes[ 1 ]. The disease is caused by germline mutations in the FLCN gene, a tumor suppressor located on chromosome 17p11.2, which encodes the folliculin protein involved in mTOR signaling, cell growth, and energy metabolism[ 2 ]. The phenotypic expression of BHDS is highly variable, even within the same family. Among the three hallmark features, pulmonary manifestations—particularly multiple bilateral lung cysts and spontaneous pneumothorax—are often the earliest and most common clinical signs. Cutaneous lesions and renal tumors are more frequently reported in Caucasian populations, whereas their prevalence appears to be lower in Asian patients, especially those of Chinese descent. This ethnic variation in clinical presentation may contribute to underdiagnosis or misdiagnosis in non-Western populations, particularly when skin or renal symptoms are absent[ 3 , 4 ]. To date, over 600 BHDS families have been identified worldwide, the majority originating from Europe and North America. In contrast, recognition of BHDS in China remains limited. By the end of 2021, only 287 confirmed patients from 143 families had been reported in the Chinese population, suggesting potential underreporting or misrecognition due to its nonspecific and heterogeneous presentation. Given the rarity of the disease and its overlapping features with other pulmonary or renal disorders, early and accurate diagnosis of BHDS is crucial for appropriate surveillance and clinical management. A definitive diagnosis relies on the identification of a pathogenic germline variant in the FLCN gene through molecular genetic testing. Here, we present the clinical and genetic analysis of a Chinese family affected by BHDS, in which we identified a novel heterozygous nonsense mutation in FLCN. Our findings contribute to the expanding mutation spectrum of FLCN and provide further insights into the genotype–phenotype correlations in BHDS. Methods Study objects A 75-year-old female patient (the proband), who was highly suspected of having Birt–Hogg–Dubé (BHD) syndrome, was admitted to Cangnan Hospital of Wenzhou Medical University in October 2023. The proband and her available family members were enrolled in this study. Ethical approval was obtained from the Medical Ethics Committee of Cangnan Hospital of Wenzhou Medical University (Approval No. 2024025). Written informed consent was obtained from all participants prior to inclusion in the study. Diagnostic Criteria for BHD syndrome According to the 2023 Chinese Expert Consensus on the Diagnosis and Management of Birt–Hogg–Dubé (BHD) Syndrome[ 5 ], the diagnostic criteria for BHD in China include both clinical and genetic components.Clinical diagnostic criteria are as follows:① Multiple pulmonary cysts: bilateral, basally located lung cysts without other apparent cause, with or without spontaneous primary pneumothorax; ② At least five fibrofolliculomas or trichodiscomas on the face or neck; ③ Renal tumors: early-onset (< 50 years), multifocal or bilateral tumors, or tumors of the chromophobe/oncocytoma type. Genetic criteria include: ① Pathogenic germline mutation in the FLCN gene; ② A first-degree relative with BHD. A definitive diagnosis of BHD can be made when two clinical criteria are met, or when at least one clinical and one genetic criterion are satisfied. Clinical data collection Detailed clinical data of the proband and her family members were collected, including general condition, medical history, pulmonary findings, renal involvement, skin lesions, diagnosis, treatment, and follow-up information. Pulmonary involvement was assessed by chest CT, and renal abnormalities were evaluated using abdominal CT or MRI. Imaging findings were reviewed by two senior radiologists, and consensus was reached. Skin lesions were examined by an experienced dermatologist and confirmed by pathological biopsy when necessary. After written informed consent was obtained, 4 mL of peripheral venous blood was collected into EDTA anticoagulant tubes and sent to a collaborating laboratory for genetic analysis. FLCN gene variation testing Genomic DNA was extracted from blood samples using the magnetic bead method according to the manufacturer’s protocol. Whole-exome sequencing (WES) was performed for the proband. DNA was first fragmented and a library was prepared. The Agilent V6 probe was used to hybridize and capture all coding regions, including flanking intronic sequences, of the FLCN gene. High-throughput sequencing was then used to identify mutations. The target region coverage was 99.6%, with an average sequencing depth of 161.0, and 96.0% of the bases covered at > 20× depth. Sanger sequencing (details available upon request) was used to validate the variants identified by WES in the proband and other family members. Pathogenicity analysis of the gene mutations Sequencing results were compared to RefSeq NM_144997.7. Variants were filtered using stringent criteria and annotated with ANNOVAR, integrating information from dbSNP, ClinVar, gnomAD, and OMIM. Pathogenicity was assessed according to the standards and guidelines of the American College of Medical Genetics and Genomics (ACMG) for the interpretation of genetic variants[ 6 ]. Additionally, a three-dimensional (3D) structural model of the folliculin protein was generated and analyzed using SWISS-MODEL and PyMol software to predict the potential impact of the mutation on protein function. Results Clinical Findings The proband was a 75-year-old female from Cangnan County, Zhejiang Province, China, with a medical history of hypertension and type 2 diabetes mellitus. She had no history of smoking or exposure to pneumotoxic agents. In 2018, a chest computed tomography (CT) scan performed at an outside hospital revealed "multiple bilateral pulmonary bullae"; however, no further diagnostic workup or treatment was pursued at that time. In October 2023, the patient presented to our hospital with a one-week history of dull, left-sided chest pain. Physical examination revealed no signs of respiratory distress, and her oxygen saturation was 96% on room air. High-resolution computed tomography (HRCT) of the chest revealed numerous thin-walled cysts of varying shapes and sizes distributed bilaterally, predominantly in the lower lobes, along with a small, left-sided spontaneous pneumothorax (Fig. 1A, 1B). The largest cyst, measuring 52 × 39 × 36 mm, was located in the dorsal segment of the right lower lobe (red arrow). The intervening lung parenchyma appeared normal, with no evidence of nodules, ground-glass opacities, septal thickening, or honeycombing. Laboratory tests, including inflammatory markers, tumor markers, rheumatoid factor, antinuclear antibodies, anti-SSA/SSB antibodies, and anti-neutrophil cytoplasmic antibodies (ANCA), were within normal limits. Pulmonary function testing was deferred due to the presence of pneumothorax. A detailed medical history revealed that one of the patient’s sons had previously been diagnosed with pulmonary cysts and recurrent pneumothorax, raising suspicion of Birt–Hogg–Dubé syndrome (BHDS). Further imaging evaluation revealed a well-circumscribed, oval-shaped mass in the left kidney measuring 30 × 23 × 28 mm (Fig. 1C, red circle) and a low-density lesion in the right paracolic sulcus measuring 41 × 26 × 28 mm (Fig. 1D, red circle). Due to the patient’s advanced age and elevated serum creatinine, contrast-enhanced CT or MRI was not performed. Suspected fibrofolliculomas were also observed on the facial skin (Fig. 1E, 1F, red arrows), further supporting the clinical diagnosis of BHDS. Although kidney and skin biopsies were recommended, the patient declined both procedures. She was treated conservatively with high-flow oxygen, resulting in resolution of the pneumothorax. No recurrence was observed during subsequent outpatient and telephone follow-up. A detailed family history was obtained. The proband’s parents and one brother had passed away many years ago, and their medical histories were unknown. Her husband (I-2) was in good health. The couple had five children, and the family pedigree is shown in Fig. 2 A. Among them, four children (II-1, II-2, II-3, and II-4) reported a history of pulmonary cysts, with no evidence of renal tumors or cutaneous lesions.Individuals II-1 and II-3 resided in distant provinces and self-reported a history of pulmonary cysts. II-3 had previously undergone lobectomy due to recurrent pneumothorax. However, both individuals had lost their historical medical records and declined further evaluation or imaging.Individual II-2 had a prior chest CT scan that revealed multiple bilateral pulmonary cysts (Fig. 2 B–C). He had experienced more than four episodes of spontaneous pneumothorax since the age of 40, all of which were treated conservatively with closed thoracic drainage.Individual II-4 had two episodes of spontaneous pneumothorax, the first at age 35 and the second five years later. He subsequently underwent wedge resection of the right upper lobe, which contained multiple emphysematous bullae. A previous chest CT scan showed right-sided pulmonary cysts and pneumothorax, while the left lung appeared normal (Fig. 2 D–E). Neither II-2 nor II-4 had any documented renal or skin involvement.The proband’s husband (I-2) and youngest daughter (II-5) were clinically unaffected and showed no signs of BHDS. None of the proband’s grandchildren reported symptoms suggestive of BHDS, and all declined further CT or MRI evaluation.To confirm the genetic diagnosis, peripheral blood samples were collected from the proband (I-1), her husband (I-2), and two children (II-4 and II-5) for free genetic testing after obtaining written informed consent. Unfortunately, individuals II-1, II-2, II-3, and all grandchildren declined genetic testing due to concerns such as potential psychological stress and the lack of curative treatment for BHDS. Genetic test results Whole-exome sequencing (WES) of the proband identified a novel heterozygous nonsense mutation, c.1222C > T (p.Gln408Ter), in exon 11 of the FLCN gene. This variant has not been previously reported in the literature or in any public population databases. The same heterozygous variant was detected in her fourth child (II-4) (Fig. 3 A). Her husband (I-2) and phenotypically normal daughter (II-5) showed the wild-type sequence at the corresponding locus (Fig. 3 B). Pathogenicity Assessment of the FLCN c.1222C > T Variant The FLCN c.1222C > T (p.Gln408Ter) variant has not been reported in individuals with FLCN-related disorders and is absent from population databases such as gnomAD. This nonsense mutation introduces a premature stop codon at amino acid position 408, which is expected to result in early termination of the folliculin protein or activation of nonsense-mediated mRNA decay, thereby causing loss of protein function. The patient’s clinical features are consistent with Birt–Hogg–Dubé syndrome, and the ClinVar database classifies this variant as pathogenic. According to the American College of Medical Genetics and Genomics (ACMG) guidelines, the variant meets the following criteria for pathogenicity: PVS1 (null variant in a gene where loss-of-function is a known mechanism), PM2_Supporting (absent from population databases), and PP4 (patient's phenotype is highly specific for a disease with a single genetic etiology) [ 6 ]. The folliculin protein contains two major functional domains: the Longin/N-terminal domain (residues Lys105–Cys265) and the C-terminal domain (residues Pro344–Met566) [ 7 ]. The identified nonsense mutation leads to truncation of the protein beyond residue 408, resulting in loss of the majority of the C-terminal domain, which plays a critical role in cellular metabolic signaling pathways. To further assess the functional consequences of the mutation, a three-dimensional (3D) structural model of the wild-type and mutant folliculin proteins was generated using SWISS-MODEL and visualized in PyMol. The model demonstrated that the p.Gln408Ter mutation results in truncation of the C-terminal domain, significantly disrupting the structural integrity of the protein. Discussion The exact prevalence of Birt-Hogg-Dubé syndrome (BHDS) remains unclear due to its clinical heterogeneity and variable penetrance. Müller ME et al. estimated the prevalence of BHDS in the general population to be approximately 2 cases per million, with no gender difference, by applying Bayes’ theorem to epidemiological data on spontaneous pneumothorax[8]. Based on this estimate, there would be approximately 2,800 BHDS patients in the Chinese population of 1.4 billion. In recent years, more cases and studies from China have been reported, reflecting increased awareness of BHDS[3, 9, 10]. The typical clinical manifestations of BHDS involve the lungs, skin, and kidneys. Among these, pulmonary involvement is the most common and usually the earliest manifestation. Over 80% of patients develop multiple bilateral thin-walled pulmonary cysts, often distributed subpleurally and parasternally with a basilar predominance. These cysts vary widely in shape and number, while the surrounding lung parenchyma remains normal[11, 12]. Most patients are asymptomatic unless cyst rupture leads to pneumothorax. The risk of pneumothorax in BHDS patients is approximately 50 times higher than in the general population, typically occurring in the third or fourth decade of life[13]. In Chinese patients, 58–71% have experienced pneumothorax, with a high tendency for recurrence[9, 14]. Skin manifestations in BHDS include fibrofolliculomas, trichodiscomas, and acrochordons, typically developing in the fourth decade of life and increasing with age[15]. Fibrofolliculomas are characteristic of the syndrome, appearing as small, pale, flesh-colored, smooth, dome-shaped papules most commonly located on the face, neck, and upper trunk. The most serious complication of BHDS is renal tumors, which occur in approximately 30% of patients, with a median age at diagnosis of 48 years[16]. These tumors are often bilateral, multifocal, and slow-growing. BHDS patients have a 7-fold increased risk of developing renal tumors, and this risk increases with age[13]. The most commonly reported histopathologic subtype is the hybrid oncocytic tumor, which displays features of both chromophobe renal cell carcinoma and renal oncocytoma[12, 16]. Potential racial differences in BHDS clinical manifestations have been observed. Several studies suggest that Asian patients have a higher incidence of pulmonary involvement but a lower prevalence of skin lesions and renal tumors compared to Caucasian patients[4, 17]. BHDS also exhibits considerable phenotypic heterogeneity; patients may present with isolated or combined pulmonary, skin, or renal symptoms, and disease severity can vary significantly even within the same family[18]. In this family, we identified five individuals with varying degrees of lung involvement, four of whom had a history of pneumothorax. Except for the elderly proband, no skin or kidney abnormalities were detected in other affected family members. These findings further confirm that the lungs are the most commonly involved organ in Chinese BHDS patients and suggest that the development of BHDS-related features is age-dependent, underscoring the need for long-term follow-up. In addition to the classic manifestations, colorectal polyps and carcinomas have also been reported in BHDS. Our proband had a neoplasm in the right paracolic sulcus; however, the association between such lesions and BHDS remains controversial and warrants further investigation[19, 20]. The underlying cause of Birt-Hogg-Dubé syndrome (BHDS) is a germline loss-of-function mutation in the FLCN gene, which encodes the protein folliculin. Folliculin is expressed in various tissues, including the lungs, skin, and kidneys[21]. Although the exact function of folliculin remains incompletely understood, it is believed to regulate cell growth, proliferation, and survival through interactions with the adenosine monophosphate-activated protein kinase (AMPK) and mechanistic target of rapamycin (mTOR) signaling pathways[22, 23]. The FLCN gene is located on chromosome 17p11.2, contains 14 exons, and encodes a protein of 579 amino acids; the translation initiation codon is located in exon 4. Currently, FLCN is the only gene known to be associated with BHDS, and the identification of a pathogenic mutation in this gene remains the gold standard for diagnosis[24]. Due to clinical suspicion of BHDS, FLCN gene testing was performed in this family. The results revealed a heterozygous nonsense mutation, c.1222C>T, in exon 11 of the FLCN gene, confirming the diagnosis of BHDS. As of April 2024, a total of 335 unique public DNA variants in FLCN have been documented in the Leiden Open Variation Database (https://databases.lovd.nl/shared/genes/FLCN). These variants include small deletions, small insertions, small indels, nonsense mutations, splice-site mutations, missense mutations, gross deletions, gross insertions, and regulatory variants. The most frequently reported mutation is c.1285dupC/delC, a cytosine duplication/deletion in a C8 tract within exon 11[25, 26]. Genotypic variation exists among different racial populations, which may explain the atypical phenotypes observed in Asian BHDS patients compared with Caucasians. By the end of 2021, a total of 63 distinct FLCN mutations had been identified in Chinese patients[3]. The previously reported mutational hotspot, c.1285dupC/delC, remains the most common variant in Chinese patients, with a similar frequency (25%)[3]. Despite this shared hotspot, the FLCN mutation spectrum in Chinese BHDS patients shows notable differences from that in other populations. More than half of the mutations identified in Chinese individuals have not been previously reported in other races[14]. The novel heterozygous nonsense mutation c.1222C>T identified in this study has not been reported in the literature. Some genotypes have been suggested to be associated with specific clinical manifestations[27, 28], but the exact genotype-phenotype correlation in BHDS remains to be fully established. Conclusion this study described the clinical and genetic characteristics of a Chinese family with BHDS. A novel nonsense mutation, c.1222C>T (p.Gln408Ter), was identified in exon 11 of the FLCN gene, providing a molecular basis for clinical diagnosis and genetic counseling in this family and expanding the known FLCN mutation spectrum in BHDS. Based on these findings, BHDS should be considered in patients with multiple lung cysts and a family history of pneumothorax, and FLCN genetic testing is recommended. Abbreviations BHDS Birt–Hogg–Dubé syndrome WES Whole-exome sequencing HRCT High-resolution computed tomography AMPK Adenosine monophosphate-activated protein kinase Declarations Ethics approval and consent to participate This study was approved by the Medical Ethics Committee of Cangnan Hospital of Wenzhou Medical University (No. 2024025). Written informed consent was obtained from the proband and family members. Consent for publication Not applicable. Competing interests The authors declares no competing interests. Funding This study was supported by Hainan Provincial Natural Science Foundation of China, No.823RC571 and Hainan Provincial Health Industry Research Projects of China, No.22A200023. Availability of data and materials The data analyzed in this study are not subject to any license restrictions. Requests for access should be directed to [email protected] . The datasets generated during the study are available in public repositories. Acknowledgments We thank the patients and their families for their cooperation in this study. Authors’ contributions MKJ and DXX designed the study and performed the research. HSJ and CZFanalyzed the data and wrote the manuscript. HSJ and ZL collected the data. All authors discussed the results and revised the manuscript. References Menko FH, van Steensel MA, Giraud S, Friis-Hansen L, Richard S, Ungari S, Nordenskjold M, Hansen TV, Solly J, Maher ER et al : Birt-Hogg-Dube syndrome: diagnosis and management . Lancet Oncol 2009, 10 (12):1199-1206. 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Cite Share Download PDF Status: Published Journal Publication published 19 Jan, 2026 Read the published version in BMC Medical Genomics → Version 1 posted Editorial decision: Revision requested 14 Oct, 2025 Editor assigned by journal 13 Oct, 2025 Submission checks completed at journal 13 Oct, 2025 First submitted to journal 12 Oct, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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(Hainan Affiliated Hospital of Hainan Medical University)","correspondingAuthor":false,"prefix":"","firstName":"CHEN","middleName":"","lastName":"Zefu","suffix":""},{"id":529472211,"identity":"defea6c8-34d0-4f53-a13b-a7f53eb392ab","order_by":2,"name":"Lin ZHANG","email":"","orcid":"","institution":"The Affiliated Cangnan Hospital of Wenzhou Medical University","correspondingAuthor":false,"prefix":"","firstName":"Lin","middleName":"","lastName":"ZHANG","suffix":""},{"id":529472215,"identity":"a97f8363-91b6-4cb1-afb3-5cd14f2de0cc","order_by":3,"name":"DING Xiuxiu","email":"","orcid":"","institution":"Hainan General Hospital (Hainan Affiliated Hospital of Hainan Medical University)","correspondingAuthor":false,"prefix":"","firstName":"DING","middleName":"","lastName":"Xiuxiu","suffix":""},{"id":529472216,"identity":"120657de-4995-4109-a964-fdd636a94136","order_by":4,"name":"MIU Keji","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAt0lEQVRIiWNgGAWjYDACHoaEA4l/bOTY2NsPEK0l8cDDhjRjPp4zCURrYT74sOFQ4jwJBwPidOj2HHhwIHHHgfQ2CYYEhh8V2whrMTvbAPTLmTu5bdKNBxh7ztwmQst5oPcT2J7ltskcSGBmbCNey+F0NokEAyK1gB3WdjiBBC1nDgBtOZNm2AYM5IPE+eVMTvLHHxU28vLt7Qcf/KggQgswYhLgzAPEqAcCdmIVjoJRMApGwYgFADUDR0j3YmFFAAAAAElFTkSuQmCC","orcid":"","institution":"The Affiliated Cangnan Hospital of Wenzhou Medical University","correspondingAuthor":true,"prefix":"","firstName":"MIU","middleName":"","lastName":"Keji","suffix":""}],"badges":[],"createdAt":"2025-10-12 15:08:27","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7841541/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7841541/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s12920-026-02311-y","type":"published","date":"2026-01-19T15:59:15+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":93571142,"identity":"9a2d9c9d-a1d2-4560-aa22-15847eca6349","added_by":"auto","created_at":"2025-10-15 09:02:47","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":1685575,"visible":true,"origin":"","legend":"","description":"","filename":"ClinicalCharacteristicsandGeneticAnalysisofaChineseFamilywithBirtHoggDubeSyndromeHarboringaNovelFLCNGeneMutation2.docx","url":"https://assets-eu.researchsquare.com/files/rs-7841541/v1/eb6c330ef7d4b9844a472d49.docx"},{"id":93571141,"identity":"b74f6f2d-6fef-45bc-b02c-36c63eaf0650","added_by":"auto","created_at":"2025-10-15 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1","display":"","copyAsset":false,"role":"figure","size":711096,"visible":true,"origin":"","legend":"\u003cp\u003eRepresentative manifestations in lung, abdomen and skin observed in the patient.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eA\u003c/strong\u003e and\u003cstrong\u003eB\u003c/strong\u003e. Chest HRCT scan revealedmultiple thin-walled cysts in both lungs of varying sizes; \u003cstrong\u003eC.\u003c/strong\u003e Abdominal CT scan revealed a left renal tumor (red circle); \u003cstrong\u003eD. \u003c/strong\u003ePelvic CT scan showed a neoplasm in the right paracolic sulcus (red circle); \u003cstrong\u003eE\u003c/strong\u003e and \u003cstrong\u003eF.\u003c/strong\u003eSuspected fibrofollicular tumors were observed on the patient’s facial skin (red arrows).\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7841541/v1/ce35fded24984228340163f7.png"},{"id":93571149,"identity":"c4edcc85-336c-466d-ab43-c391660fad08","added_by":"auto","created_at":"2025-10-15 09:02:47","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":360877,"visible":true,"origin":"","legend":"\u003cp\u003ePedigree and chest CT findings of the proband’s family.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eA.\u003c/strong\u003e Pedigree chart of the patient’s family in two generations; \u003cstrong\u003eB \u003c/strong\u003eand \u003cstrong\u003eC\u003c/strong\u003e. Previous chest CT images of the patient's second child (II-2); \u003cstrong\u003eD\u003c/strong\u003e and \u003cstrong\u003eE. \u003c/strong\u003ePrevious chest CT images of the patient's fourth child (II-4).\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-7841541/v1/00a468df2db0720b79ecedc8.png"},{"id":93573391,"identity":"090d3878-8d77-4ca0-b99c-ad633ffcade2","added_by":"auto","created_at":"2025-10-15 09:10:47","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":262056,"visible":true,"origin":"","legend":"\u003cp\u003eValidation of the FLCN c.1222C\u0026gt;T (p.Gln408Ter) variant by Sanger sequencing.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eA. \u003c/strong\u003eSanger sequencing showing the heterozygous FLCN c.1222C\u0026gt;T (p.Gln408Ter) variant in the proband and individual II-4. Arrow indicates the mutation site; \u003cstrong\u003eB.\u003c/strong\u003e Wild-type FLCN sequence in the proband’s husband and youngest daughter (II-5).\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7841541/v1/c40e1e90b65ae7b19df50da6.png"},{"id":93573392,"identity":"d9ce0f18-e97e-4025-b27d-804f69c5c07a","added_by":"auto","created_at":"2025-10-15 09:10:47","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":231103,"visible":true,"origin":"","legend":"\u003cp\u003eStructural modeling of the wild-type and mutant folliculin protein.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eA.\u003c/strong\u003e Predicted 3D structure of the full-length wild-type folliculin protein; \u003cstrong\u003eB.\u003c/strong\u003eTruncated folliculin protein structure due to c.1222C\u0026gt;T mutation, showing chain termination after residue Ser407.\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-7841541/v1/23820f705db868ab3a6d41ef.png"},{"id":101153351,"identity":"95a5ac0b-42e2-4038-a23f-fb5b166fc3b7","added_by":"auto","created_at":"2026-01-26 16:14:52","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3560952,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7841541/v1/a9966c42-ba69-4013-81f0-bcca88d039a4.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Clinical Characteristics and Genetic Analysis of a Chinese Family with Birt–Hogg–Dubé Syndrome Harboring a Novel FLCN Gene Mutation","fulltext":[{"header":"Background","content":"\u003cp\u003eBirt\u0026ndash;Hogg\u0026ndash;Dub\u0026eacute; syndrome (BHDS; OMIM #135150), also known as Hornstein\u0026ndash;Knickenberg syndrome, is a rare autosomal dominant disorder first described in 1977. It is clinically characterized by the triad of multiple pulmonary cysts with or without recurrent spontaneous pneumothorax, benign cutaneous lesions such as fibrofolliculomas or trichodiscomas, and an increased risk of renal neoplasms of various histological subtypes[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. The disease is caused by germline mutations in the FLCN gene, a tumor suppressor located on chromosome 17p11.2, which encodes the folliculin protein involved in mTOR signaling, cell growth, and energy metabolism[\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe phenotypic expression of BHDS is highly variable, even within the same family. Among the three hallmark features, pulmonary manifestations\u0026mdash;particularly multiple bilateral lung cysts and spontaneous pneumothorax\u0026mdash;are often the earliest and most common clinical signs. Cutaneous lesions and renal tumors are more frequently reported in Caucasian populations, whereas their prevalence appears to be lower in Asian patients, especially those of Chinese descent. This ethnic variation in clinical presentation may contribute to underdiagnosis or misdiagnosis in non-Western populations, particularly when skin or renal symptoms are absent[\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eTo date, over 600 BHDS families have been identified worldwide, the majority originating from Europe and North America. In contrast, recognition of BHDS in China remains limited. By the end of 2021, only 287 confirmed patients from 143 families had been reported in the Chinese population, suggesting potential underreporting or misrecognition due to its nonspecific and heterogeneous presentation.\u003c/p\u003e\u003cp\u003eGiven the rarity of the disease and its overlapping features with other pulmonary or renal disorders, early and accurate diagnosis of BHDS is crucial for appropriate surveillance and clinical management. A definitive diagnosis relies on the identification of a pathogenic germline variant in the FLCN gene through molecular genetic testing. Here, we present the clinical and genetic analysis of a Chinese family affected by BHDS, in which we identified a novel heterozygous nonsense mutation in FLCN. Our findings contribute to the expanding mutation spectrum of FLCN and provide further insights into the genotype\u0026ndash;phenotype correlations in BHDS.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eStudy objects\u003c/h2\u003e\u003cp\u003eA 75-year-old female patient (the proband), who was highly suspected of having Birt\u0026ndash;Hogg\u0026ndash;Dub\u0026eacute; (BHD) syndrome, was admitted to Cangnan Hospital of Wenzhou Medical University in October 2023. The proband and her available family members were enrolled in this study. Ethical approval was obtained from the Medical Ethics Committee of Cangnan Hospital of Wenzhou Medical University (Approval No. 2024025). Written informed consent was obtained from all participants prior to inclusion in the study.\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eDiagnostic Criteria for BHD syndrome\u003c/h3\u003e\n\u003cp\u003eAccording to the 2023 Chinese Expert Consensus on the Diagnosis and Management of Birt\u0026ndash;Hogg\u0026ndash;Dub\u0026eacute; (BHD) Syndrome[\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e], the diagnostic criteria for BHD in China include both clinical and genetic components.Clinical diagnostic criteria are as follows:① Multiple pulmonary cysts: bilateral, basally located lung cysts without other apparent cause, with or without spontaneous primary pneumothorax; ② At least five fibrofolliculomas or trichodiscomas on the face or neck; ③ Renal tumors: early-onset (\u0026lt;\u0026thinsp;50 years), multifocal or bilateral tumors, or tumors of the chromophobe/oncocytoma type. Genetic criteria include: ① Pathogenic germline mutation in the FLCN gene; ② A first-degree relative with BHD. A definitive diagnosis of BHD can be made when two clinical criteria are met, or when at least one clinical and one genetic criterion are satisfied.\u003c/p\u003e\n\u003ch3\u003eClinical data collection\u003c/h3\u003e\n\u003cp\u003eDetailed clinical data of the proband and her family members were collected, including general condition, medical history, pulmonary findings, renal involvement, skin lesions, diagnosis, treatment, and follow-up information. Pulmonary involvement was assessed by chest CT, and renal abnormalities were evaluated using abdominal CT or MRI. Imaging findings were reviewed by two senior radiologists, and consensus was reached. Skin lesions were examined by an experienced dermatologist and confirmed by pathological biopsy when necessary. After written informed consent was obtained, 4 mL of peripheral venous blood was collected into EDTA anticoagulant tubes and sent to a collaborating laboratory for genetic analysis.\u003c/p\u003e\n\u003ch3\u003eFLCN gene variation testing\u003c/h3\u003e\n\u003cp\u003eGenomic DNA was extracted from blood samples using the magnetic bead method according to the manufacturer\u0026rsquo;s protocol. Whole-exome sequencing (WES) was performed for the proband. DNA was first fragmented and a library was prepared. The Agilent V6 probe was used to hybridize and capture all coding regions, including flanking intronic sequences, of the FLCN gene. High-throughput sequencing was then used to identify mutations. The target region coverage was 99.6%, with an average sequencing depth of 161.0, and 96.0% of the bases covered at \u0026gt;\u0026thinsp;20\u0026times; depth. Sanger sequencing (details available upon request) was used to validate the variants identified by WES in the proband and other family members.\u003c/p\u003e\n\u003ch3\u003ePathogenicity analysis of the gene mutations\u003c/h3\u003e\n\u003cp\u003eSequencing results were compared to RefSeq NM_144997.7. Variants were filtered using stringent criteria and annotated with ANNOVAR, integrating information from dbSNP, ClinVar, gnomAD, and OMIM. Pathogenicity was assessed according to the standards and guidelines of the American College of Medical Genetics and Genomics (ACMG) for the interpretation of genetic variants[\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Additionally, a three-dimensional (3D) structural model of the folliculin protein was generated and analyzed using SWISS-MODEL and PyMol software to predict the potential impact of the mutation on protein function.\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e\u003ch2\u003eClinical Findings\u003c/h2\u003e\u003cp\u003eThe proband was a 75-year-old female from Cangnan County, Zhejiang Province, China, with a medical history of hypertension and type 2 diabetes mellitus. She had no history of smoking or exposure to pneumotoxic agents. In 2018, a chest computed tomography (CT) scan performed at an outside hospital revealed \"multiple bilateral pulmonary bullae\"; however, no further diagnostic workup or treatment was pursued at that time.\u003c/p\u003e\u003cp\u003eIn October 2023, the patient presented to our hospital with a one-week history of dull, left-sided chest pain. Physical examination revealed no signs of respiratory distress, and her oxygen saturation was 96% on room air. High-resolution computed tomography (HRCT) of the chest revealed numerous thin-walled cysts of varying shapes and sizes distributed bilaterally, predominantly in the lower lobes, along with a small, left-sided spontaneous pneumothorax (Fig.\u0026nbsp;1A, 1B). The largest cyst, measuring 52 \u0026times; 39 \u0026times; 36 mm, was located in the dorsal segment of the right lower lobe (red arrow). The intervening lung parenchyma appeared normal, with no evidence of nodules, ground-glass opacities, septal thickening, or honeycombing.\u003c/p\u003e\u003cp\u003eLaboratory tests, including inflammatory markers, tumor markers, rheumatoid factor, antinuclear antibodies, anti-SSA/SSB antibodies, and anti-neutrophil cytoplasmic antibodies (ANCA), were within normal limits. Pulmonary function testing was deferred due to the presence of pneumothorax.\u003c/p\u003e\u003cp\u003eA detailed medical history revealed that one of the patient\u0026rsquo;s sons had previously been diagnosed with pulmonary cysts and recurrent pneumothorax, raising suspicion of Birt\u0026ndash;Hogg\u0026ndash;Dub\u0026eacute; syndrome (BHDS). Further imaging evaluation revealed a well-circumscribed, oval-shaped mass in the left kidney measuring 30 \u0026times; 23 \u0026times; 28 mm (Fig.\u0026nbsp;1C, red circle) and a low-density lesion in the right paracolic sulcus measuring 41 \u0026times; 26 \u0026times; 28 mm (Fig.\u0026nbsp;1D, red circle). Due to the patient\u0026rsquo;s advanced age and elevated serum creatinine, contrast-enhanced CT or MRI was not performed. Suspected fibrofolliculomas were also observed on the facial skin (Fig.\u0026nbsp;1E, 1F, red arrows), further supporting the clinical diagnosis of BHDS. Although kidney and skin biopsies were recommended, the patient declined both procedures. She was treated conservatively with high-flow oxygen, resulting in resolution of the pneumothorax. No recurrence was observed during subsequent outpatient and telephone follow-up.\u003c/p\u003e\u003cp\u003eA detailed family history was obtained. The proband\u0026rsquo;s parents and one brother had passed away many years ago, and their medical histories were unknown. Her husband (I-2) was in good health. The couple had five children, and the family pedigree is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e2\u003c/span\u003eA. Among them, four children (II-1, II-2, II-3, and II-4) reported a history of pulmonary cysts, with no evidence of renal tumors or cutaneous lesions.Individuals II-1 and II-3 resided in distant provinces and self-reported a history of pulmonary cysts. II-3 had previously undergone lobectomy due to recurrent pneumothorax. However, both individuals had lost their historical medical records and declined further evaluation or imaging.Individual II-2 had a prior chest CT scan that revealed multiple bilateral pulmonary cysts (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e2\u003c/span\u003eB\u0026ndash;C). He had experienced more than four episodes of spontaneous pneumothorax since the age of 40, all of which were treated conservatively with closed thoracic drainage.Individual II-4 had two episodes of spontaneous pneumothorax, the first at age 35 and the second five years later. He subsequently underwent wedge resection of the right upper lobe, which contained multiple emphysematous bullae. A previous chest CT scan showed right-sided pulmonary cysts and pneumothorax, while the left lung appeared normal (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e2\u003c/span\u003eD\u0026ndash;E). Neither II-2 nor II-4 had any documented renal or skin involvement.The proband\u0026rsquo;s husband (I-2) and youngest daughter (II-5) were clinically unaffected and showed no signs of BHDS. None of the proband\u0026rsquo;s grandchildren reported symptoms suggestive of BHDS, and all declined further CT or MRI evaluation.To confirm the genetic diagnosis, peripheral blood samples were collected from the proband (I-1), her husband (I-2), and two children (II-4 and II-5) for free genetic testing after obtaining written informed consent. Unfortunately, individuals II-1, II-2, II-3, and all grandchildren declined genetic testing due to concerns such as potential psychological stress and the lack of curative treatment for BHDS.\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eGenetic test results\u003c/h3\u003e\n\u003cp\u003eWhole-exome sequencing (WES) of the proband identified a novel heterozygous nonsense mutation, c.1222C\u0026thinsp;\u0026gt;\u0026thinsp;T (p.Gln408Ter), in exon 11 of the FLCN gene. This variant has not been previously reported in the literature or in any public population databases. The same heterozygous variant was detected in her fourth child (II-4) (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e3\u003c/span\u003eA). Her husband (I-2) and phenotypically normal daughter (II-5) showed the wild-type sequence at the corresponding locus (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e3\u003c/span\u003eB).\u003c/p\u003e\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\u003ch2\u003ePathogenicity Assessment of the FLCN c.1222C\u0026thinsp;\u0026gt;\u0026thinsp;T Variant\u003c/h2\u003e\u003cp\u003eThe FLCN c.1222C\u0026thinsp;\u0026gt;\u0026thinsp;T (p.Gln408Ter) variant has not been reported in individuals with FLCN-related disorders and is absent from population databases such as gnomAD. This nonsense mutation introduces a premature stop codon at amino acid position 408, which is expected to result in early termination of the folliculin protein or activation of nonsense-mediated mRNA decay, thereby causing loss of protein function. The patient\u0026rsquo;s clinical features are consistent with Birt\u0026ndash;Hogg\u0026ndash;Dub\u0026eacute; syndrome, and the ClinVar database classifies this variant as pathogenic. According to the American College of Medical Genetics and Genomics (ACMG) guidelines, the variant meets the following criteria for pathogenicity: PVS1 (null variant in a gene where loss-of-function is a known mechanism), PM2_Supporting (absent from population databases), and PP4 (patient's phenotype is highly specific for a disease with a single genetic etiology) [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. The folliculin protein contains two major functional domains: the Longin/N-terminal domain (residues Lys105\u0026ndash;Cys265) and the C-terminal domain (residues Pro344\u0026ndash;Met566) [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. The identified nonsense mutation leads to truncation of the protein beyond residue 408, resulting in loss of the majority of the C-terminal domain, which plays a critical role in cellular metabolic signaling pathways. To further assess the functional consequences of the mutation, a three-dimensional (3D) structural model of the wild-type and mutant folliculin proteins was generated using SWISS-MODEL and visualized in PyMol. The model demonstrated that the p.Gln408Ter mutation results in truncation of the C-terminal domain, significantly disrupting the structural integrity of the protein.\u003c/p\u003e\u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe exact prevalence of Birt-Hogg-Dub\u0026eacute; syndrome (BHDS) remains unclear due to its clinical heterogeneity and variable penetrance. M\u0026uuml;ller ME et al. estimated the prevalence of BHDS in the general population to be approximately 2 cases per million, with no gender difference, by applying Bayes\u0026rsquo; theorem to epidemiological data on spontaneous pneumothorax[8]. Based on this estimate, there would be approximately 2,800 BHDS patients in the Chinese population of 1.4 billion. In recent years, more cases and studies from China have been reported, reflecting increased awareness of BHDS[3, 9, 10].\u003c/p\u003e\n\u003cp\u003eThe typical clinical manifestations of BHDS involve the lungs, skin, and kidneys. Among these, pulmonary involvement is the most common and usually the earliest manifestation. Over 80% of patients develop multiple bilateral thin-walled pulmonary cysts, often distributed subpleurally and parasternally with a basilar predominance. These cysts vary widely in shape and number, while the surrounding lung parenchyma remains normal[11, 12]. Most patients are asymptomatic unless cyst rupture leads to pneumothorax. The risk of pneumothorax in BHDS patients is approximately 50 times higher than in the general population, typically occurring in the third or fourth decade of life[13]. In Chinese patients, 58\u0026ndash;71% have experienced pneumothorax, with a high tendency for recurrence[9, 14]. Skin manifestations in BHDS include fibrofolliculomas, trichodiscomas, and acrochordons, typically developing in the fourth decade of life and increasing with age[15]. Fibrofolliculomas are characteristic of the syndrome, appearing as small, pale, flesh-colored, smooth, dome-shaped papules most commonly located on the face, neck, and upper trunk. The most serious complication of BHDS is renal tumors, which occur in approximately 30% of patients, with a median age at diagnosis of 48 years[16]. These tumors are often bilateral, multifocal, and slow-growing. BHDS patients have a 7-fold increased risk of developing renal tumors, and this risk increases with age[13]. The most commonly reported histopathologic subtype is the hybrid oncocytic tumor, which displays features of both chromophobe renal cell carcinoma and renal oncocytoma[12, 16].\u003c/p\u003e\n\u003cp\u003ePotential racial differences in BHDS clinical manifestations have been observed. Several studies suggest that Asian patients have a higher incidence of pulmonary involvement but a lower prevalence of skin lesions and renal tumors compared to Caucasian patients[4, 17]. BHDS also exhibits considerable phenotypic heterogeneity; patients may present with isolated or combined pulmonary, skin, or renal symptoms, and disease severity can vary significantly even within the same family[18].\u003c/p\u003e\n\u003cp\u003eIn this family, we identified five individuals with varying degrees of lung involvement, four of whom had a history of pneumothorax. Except for the elderly proband, no skin or kidney abnormalities were detected in other affected family members. These findings further confirm that the lungs are the most commonly involved organ in Chinese BHDS patients and suggest that the development of BHDS-related features is age-dependent, underscoring the need for long-term follow-up. In addition to the classic manifestations, colorectal polyps and carcinomas have also been reported in BHDS. Our proband had a neoplasm in the right paracolic sulcus; however, the association between such lesions and BHDS remains controversial and warrants further investigation[19, 20].\u003c/p\u003e\n\u003cp\u003eThe underlying cause of Birt-Hogg-Dub\u0026eacute; syndrome (BHDS) is a germline loss-of-function mutation in the FLCN gene, which encodes the protein folliculin. Folliculin is expressed in various tissues, including the lungs, skin, and kidneys[21]. Although the exact function of folliculin remains incompletely understood, it is believed to regulate cell growth, proliferation, and survival through interactions with the adenosine monophosphate-activated protein kinase (AMPK) and mechanistic target of rapamycin (mTOR) signaling pathways[22, 23]. The FLCN gene is located on chromosome 17p11.2, contains 14 exons, and encodes a protein of 579 amino acids; the translation initiation codon is located in exon 4. Currently, FLCN is the only gene known to be associated with BHDS, and the identification of a pathogenic mutation in this gene remains the gold standard for diagnosis[24]. Due to clinical suspicion of BHDS, FLCN gene testing was performed in this family. The results revealed a heterozygous nonsense mutation, c.1222C\u0026gt;T, in exon 11 of the FLCN gene, confirming the diagnosis of BHDS. As of April 2024, a total of 335 unique public DNA variants in FLCN have been documented in the Leiden Open Variation Database (https://databases.lovd.nl/shared/genes/FLCN). These variants include small deletions, small insertions, small indels, nonsense mutations, splice-site mutations, missense mutations, gross deletions, gross insertions, and regulatory variants. The most frequently reported mutation is c.1285dupC/delC, a cytosine duplication/deletion in a C8 tract within exon 11[25, 26]. Genotypic variation exists among different racial populations, which may explain the atypical phenotypes observed in Asian BHDS patients compared with Caucasians. By the end of 2021, a total of 63 distinct FLCN mutations had been identified in Chinese patients[3]. The previously reported mutational hotspot, c.1285dupC/delC, remains the most common variant in Chinese patients, with a similar frequency (25%)[3]. Despite this shared hotspot, the FLCN mutation spectrum in Chinese BHDS patients shows notable differences from that in other populations. More than half of the mutations identified in Chinese individuals have not been previously reported in other races[14]. The novel heterozygous nonsense mutation c.1222C\u0026gt;T identified in this study has not been reported in the literature. Some genotypes have been suggested to be associated with specific clinical manifestations[27, 28], but the exact genotype-phenotype correlation in BHDS remains to be fully established.\u003c/p\u003e"},{"header":"Conclusion ","content":"\u003cp\u003ethis study described the clinical and genetic characteristics of a Chinese family with BHDS. A novel nonsense mutation, c.1222C\u0026gt;T (p.Gln408Ter), was identified in exon 11 of the FLCN gene, providing a molecular basis for clinical diagnosis and genetic counseling in this family and expanding the known FLCN mutation spectrum in BHDS. Based on these findings, BHDS should be considered in patients with multiple lung cysts and a family history of pneumothorax, and FLCN genetic testing is recommended.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eBHDS \u0026nbsp; Birt\u0026ndash;Hogg\u0026ndash;Dub\u0026eacute; syndrome\u003c/p\u003e\n\u003cp\u003eWES \u0026nbsp; \u0026nbsp;Whole-exome sequencing\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eHRCT \u0026nbsp; High-resolution computed tomography\u003c/p\u003e\n\u003cp\u003eAMPK \u0026nbsp; \u0026nbsp;Adenosine monophosphate-activated protein kinase\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was approved by the Medical Ethics Committee of Cangnan Hospital of Wenzhou Medical University (No. 2024025). Written informed consent was obtained from the proband and family members.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declares no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was supported by Hainan Provincial Natural Science Foundation of China, No.823RC571 and Hainan Provincial Health Industry Research Projects of China, No.22A200023.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u0026nbsp;\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data analyzed in this study are not subject to any license restrictions. Requests for access should be directed to [email protected]. The datasets generated during the study are available in public repositories.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank the patients and their families for their cooperation in this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026rsquo; contributions \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMKJ and DXX designed the study and performed the research. HSJ and CZFanalyzed the data and wrote the manuscript. HSJ and ZL collected the data. All authors discussed the results and revised the manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eMenko FH, van Steensel MA, Giraud S, Friis-Hansen L, Richard S, Ungari S, Nordenskjold M, Hansen TV, Solly J, Maher ER\u003cem\u003e et al\u003c/em\u003e: \u003cstrong\u003eBirt-Hogg-Dube syndrome: diagnosis and management\u003c/strong\u003e. \u003cem\u003eLancet Oncol \u003c/em\u003e2009, \u003cstrong\u003e10\u003c/strong\u003e(12):1199-1206.\u003c/li\u003e\n\u003cli\u003eWoodford MR, Andreou A, Baba M, van de Beek I, Di Malta C, Glykofridis I, Grimes H, Henske EP, Iliopoulos O, Kurihara M\u003cem\u003e et al\u003c/em\u003e: \u003cstrong\u003eSeventh BHD international symposium: recent scientific and clinical advancement\u003c/strong\u003e. \u003cem\u003eOncotarget \u003c/em\u003e2022, \u003cstrong\u003e13\u003c/strong\u003e:173-181.\u003c/li\u003e\n\u003cli\u003eZhou W, Liu K, Xu KF, Liu Y, Tian X: \u003cstrong\u003eClinical and Genetic 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Birt-Hogg-Dube syndrome\u003c/strong\u003e. \u003cem\u003eClin Genet \u003c/em\u003e2016, \u003cstrong\u003e90\u003c/strong\u003e(5):403-412.\u003c/li\u003e\n\u003cli\u003eSattler EC, Syunyaeva Z, Mansmann U, Steinlein OK: \u003cstrong\u003eGenetic Risk Factors for Spontaneous Pneumothorax in Birt-Hogg-Dube Syndrome\u003c/strong\u003e. \u003cem\u003eChest \u003c/em\u003e2020, \u003cstrong\u003e157\u003c/strong\u003e(5):1199-1206.\u003c/li\u003e\n\u003cli\u003eWang Y, Cai M, Jiang X, Lv G, Hu D, Zhang G, Liu J, Wei W, Xiao J, Shen B\u003cem\u003e et al\u003c/em\u003e: \u003cstrong\u003eExons 1-3 deletion in FLCN is associated with increased risk of pneumothorax in Chinese patients with Birt-Hogg-Dube syndrome\u003c/strong\u003e. \u003cem\u003eOrphanet J Rare Dis \u003c/em\u003e2023, \u003cstrong\u003e18\u003c/strong\u003e(1):115.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-medical-genomics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mgnm","sideBox":"Learn more about [BMC Medical Genomics](http://bmcmedgenomics.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/mgnm/default.aspx","title":"BMC Medical Genomics","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Birt–Hogg–Dubé syndrome, FLCN, Genetic mutation, Pedigree study, Pulmonary cysts, Renal tumor","lastPublishedDoi":"10.21203/rs.3.rs-7841541/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7841541/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e\u003cp\u003eBirt\u0026ndash;Hogg\u0026ndash;Dub\u0026eacute; syndrome (BHDS) is a rare hereditary disorder characterized by skin lesions, pulmonary cysts, spontaneous pneumothorax, and renal neoplasia. Mutations in the \u003cem\u003eFLCN\u003c/em\u003e gene are known causes, yet novel variants continue to be identified, expanding genotype\u0026ndash;phenotype correlations.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e\u003cp\u003eWe investigated a Chinese family with suspected BHDS. The proband was admitted to the Affiliated Cangnan Hospital of Wenzhou Medical University in October 2023. Comprehensive clinical evaluations and imaging studies were performed. Peripheral blood samples were collected from the proband and available family members after obtaining informed consent. Whole-exome sequencing (WES) was conducted to identify potential variants in the FLCN gene. Candidate variants were subsequently validated by Sanger sequencing and analyzed for co-segregation within the family. Pathogenicity was assessed using multiple bioinformatic prediction tools in accordance with the American College of Medical Genetics and Genomics (ACMG) guidelines. To evaluate the structural impact of the variant, a three-dimensional model of the folliculin protein was generated using SWISS-MODEL and visualized with PyMol.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e\u003cp\u003eThe proband presented with bilateral pulmonary cysts, a small left-sided pneumothorax, a left renal tumor, and suspected cutaneous lesions. Family screening revealed that several children had pulmonary cysts, some accompanied by pneumothorax, and two had previously undergone lobectomy. Whole-exome sequencing identified a novel heterozygous nonsense mutation in exon 11 of FLCN, c.1222C\u0026thinsp;\u0026gt;\u0026thinsp;T (p.Gln408Ter), which has not been previously reported. This mutation was also detected in one of the proband\u0026rsquo;s sons, while the proband\u0026rsquo;s husband and an asymptomatic daughter were wild type at this locus. Functional prediction indicates a loss of normal folliculin activity. Based on ACMG guidelines, the variant is classified as pathogenic.\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e\u003cp\u003eThe novel \u003cem\u003eFLCN\u003c/em\u003e nonsense variant \u003cem\u003ec\u003c/em\u003e.1222C\u0026thinsp;\u0026gt;\u0026thinsp;T (p.Gln408Ter) is the likely genetic basis of BHDS in this family. These findings expand the mutational spectrum of \u003cem\u003eFLCN\u003c/em\u003e, inform genotype\u0026ndash;phenotype relationships, and support improved diagnosis and genetic counseling of BHDS.\u003c/p\u003e","manuscriptTitle":"Clinical Characteristics and Genetic Analysis of a Chinese Family with Birt–Hogg–Dubé Syndrome Harboring a Novel FLCN Gene Mutation","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-15 09:02:42","doi":"10.21203/rs.3.rs-7841541/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-10-14T12:43:37+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-10-14T01:48:52+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-10-14T01:47:43+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Medical Genomics","date":"2025-10-12T15:04:37+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-medical-genomics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mgnm","sideBox":"Learn more about [BMC Medical Genomics](http://bmcmedgenomics.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/mgnm/default.aspx","title":"BMC Medical Genomics","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"4d875bba-f010-4409-a575-2a98c2d6d078","owner":[],"postedDate":"October 15th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-01-26T16:13:16+00:00","versionOfRecord":{"articleIdentity":"rs-7841541","link":"https://doi.org/10.1186/s12920-026-02311-y","journal":{"identity":"bmc-medical-genomics","isVorOnly":false,"title":"BMC Medical Genomics"},"publishedOn":"2026-01-19 15:59:15","publishedOnDateReadable":"January 19th, 2026"},"versionCreatedAt":"2025-10-15 09:02:42","video":"","vorDoi":"10.1186/s12920-026-02311-y","vorDoiUrl":"https://doi.org/10.1186/s12920-026-02311-y","workflowStages":[]},"version":"v1","identity":"rs-7841541","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7841541","identity":"rs-7841541","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}