Her eldest daughter individual had a normal birth and health
Her eldest daughter (individual 4) had a normal birth and health history in early childhood. She developed contractures of her fourth fingers at 4–5 years of age; these contractures were associated with painless, non-erythematous swelling of the involved joints. Her PIP contractures were ∼90 degrees on the right fourth finger, but less severe on the left third finger. On her palms, she had associated scar-like thickening overlying the flexor tendons of the fourth digits (Figures 2K and 2L). She had a right cholesteatoma removed at age 4; this recurred and was removed again at age 8. On evaluation at age 8.5, she had a faint papular erythematous rash on her arms, face, neck, and chest; a 3 mm bluish papule with a punctate eschar on her right upper arm; and a soft, slightly bluish 1.5–2.5 cm mass on the sole of her left foot. Although her vision was normal, she had a superior corneal vascular pannus and reduced central corneal thickness (right eye 491 μm, left eye 484 μm), but normal endothelial cell morphology on specular microscopy. She has a long face with midface retrusion, a narrow nose, short palpebral fissures, epicanthal folds, and thin ear cartilage. Her monensin is thin and somewhat lax, similar to her mother’s, although her feet appear relatively normal. The sixth affected individual (individual 6) is a 35-year-old female who presented with a complex combination of congenital and acquired symptoms. She was born to healthy, unrelated parents with an unremarkable family history, apart from the presence of polycystic kidney disease in her father and paternal uncle. She presented with pyloric stenosis and facial dysmorphism in infancy. Psychomotor development was normal, but she suffered from recurrent ear infections that led to conductive hearing loss during childhood. In addition, she also has polycystic kidney disease. Her arms had angiodermatofibromas (benign and superficial fibrous histiocytoma), and her scalp had lichenoid skin lesions. Furthermore, she has hypothyroidism and mild mitral valve insufficiency, and during adolescence a susceptibility to bruising was noted. She has had surgical corrections of dental crowding and malpositioned teeth. At age 18 years, spontaneous pneumothorax was diagnosed. At age 22 years, she underwent sigmoid resection as a result of chronic diverticulitis and developed stenotic scarring as a later complication. At age 31years she had peritonitis after surgery associated with an ovarian abscess. Wound healing was delayed, and she developed a thick, protruding scar that in some areas was keloid-like. During treatment of her abdominal illness, she sustained an ischemic stroke, and she developed cysts in the liver. She has hypermobile joints, flat feet, contractures, and generalized joint enlargements of the fingers. Chronic ulcerations on her toes eventually led to the loss of all toes and a significant portion of forefoot tissue. She has surgical wound management on a regular basis. At her latest clinical examination (at age 35), she presented with apparently normal cognition, tall stature (∼95th percentile), reduced subcutaneous body fat, enlarged and low-set-ears, widely spaced eyes, bilateral epicanthus, bilateral blepharophimosis, a thin nose with small alae nasi, and a high palate. She is able to walk unsupported but suffers from limitations due to chronic ulcerations on her feet. Clinical testing for genomic copy-number aberrations and a connective-tissue gene panel of individuals 1 and 2 were normal (data not shown). Research exome sequencing, using both dominant and recessive models, was undertaken as described in the Supplemental Methods. We made a list of all variants shared by these unrelated individuals, then removed all variants previously classified as benign in our diagnostic pipeline, as well as all variants present in dbSNP build 137. Only four heterozygous missense variants in two genes remained. Two variants were in MUC4 [MIM: 158372] in non-conserved nucleotides and amino acids, and both were predicted to be benign by the in silico tools provided by Alamut (Interactive Biosoftware). Two variants were in DDR2 [MIM: 191311]; these variants affected conserved nucleotides and amino acids, and both were predicted to be damaging by the following in silico prediction programs: PolyPhen2, MutationTester, SIFT, and LRT. Individual 1 was heterozygous for DDR2 (GenBank: NM_001014796.1) c.1829T>C (p.Leu610Pro), and individual 2 was heterozygous for DDR2 c.2219A>G (p.Tyr740Cys). In programs provided through Alamut, neither of the variants were predicted to affect splicing. In addition, the variants had CADD scores of 30 for p.Leu610Pro and 31 for p.Tyr740Cys, also decreasing the likelihood that this was a chance finding (see Supplemental Methods for details). DDR2 is relatively intolerant to loss-of-function variants (Exac database pLI score of 0.99) and more tolerant to missense variation (Exac database Z score of 2.09). Neither of the DDR2 variants found were present in the gnomAD database, nor were any other missense changes to either the Leu610 or Tyr740 codons. Both missense changes affect conserved amino acids located in the DDR2 tyrosine kinase domain, as can be seen from the cross-species comparison made in Figure 3. The same amino acids are also conserved in DDR1 [MIM: 600408]. Subsequently, we found that individual 3 had the same p.Tyr740Cys variant previously found in individual 2 and that individual 6 had the same p.Leu610Pro variant previously found in individual 1. The variant found in individual 3 was proven to be de novo after parental testing (Figure S3). In the other individuals, samples from both parents were not available (Figures S1, S2, and S4). The two affected children of individual 3 (individuals 4 and 5) had inherited the p.Tyr740Cys variant (Figure S3). The variants were verified by Sanger sequencing (Figures S1–S3). We conclude that the identification of two unique and recurrent missense variants, p.Leu610Pro and p.Tyr740Cys, affecting conserved DDR2 amino acids, in four families with a similar clinical phenotype (one proven de novo occurrence) confirms the pathogenicity of these variants.