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Macrodactyly refers to a rare congenital deformity characterized by digital gigantism of all structures including subcutaneous fat, tendons, nerves, vessels, skin, and nails [1]. The prevalence of macrodactyly is about 0.08 per 10,000 live births [2]. Macrodactyly can be inherited when it occurs with a syndrome, but isolated macrodactyly is usually sporadic [3, 4]. Macrod-actyly of the hand is more frequently than the foot and usually involved more than one digit. The enlarged digits may grow either in static or progressive condition [2, 5]. Treatments of macrodactyly comprise soft tissue debulking, nerve stripping, epiphysiodesis, osteotomies, and phalangectomy [2].
Mutations in the PIK3CA gene have been previously described as the cause of isolated macrodactyly in Caucasian patients. The mutations were detected only in the affected tissues but not in peripheral blood [6]. Moreover, the mosaic mutations in PIK3CA were found to be correlated with several overgrowth disorders. Therefore, the umbrella term of PIK3CA-related overgrowth spectrum (PROS) was nominated to designate the broad range of clinical symptoms including isolated macrodactyly, fibroadipose overgrowth (FAO), hemi-hyperplasia multiple lipomatosis (HHML), congenital lipomatous overgrowth, vascular malformations, epidermal nevi, scoliosis/skeletal and spinal (CLOVES) syndrome, and related megalencephaly conditions [4, 7, 8].
We identified a Thai patient with diffuse enlarged left middle and left fourth fingers. As pathogenic variants in PIK3CA have been reported as the cause of this symptom [6], we aimed to determine the genetic alteration in our Thai patient with macrodactyly. The mutation could be either a germline or somatic mutation. Causative germline mutations of a genetic disorder can have different frequencies in various populations.
Materials and methods
Patient
The present case report was approved by the Institutional Review Board, Faculty of Medicine, Chulalongkorn University (certificate of approval no. 365/60). The patient’s guardian has provided the written consent for the publication of this case report and any accompanying information and images.
Tissue specimens were obtained during surgical procedures performed for the treatment of macrodactyly. A 5-year-old girl with enlarged left middle and fourth fingers was referred for further treatment at King Chulalongkorn Memorial Hospital (Figure 1).
Figure 1
Clinical and radiologic features of the patient. The radio-graphs show enlarged middle and fourth fingers of the patient’s left hand
Cell culture
Skin fibroblasts from the removed affected finger were cultured in Dulbecco’s modified Eagle medium (DMEM) (Hyclone, GE Healthcare) supplemented with 10% fetal bovine serum (Hyclone, GE Healthcare) and 1% penicillin/streptomycin (Hyclone, GE Healthcare). The morphology of cultured cells was consistent with fibroblasts. The cultured dermal fibroblasts were collected for DNA extraction.
Polymerase chain reaction and Sanger sequencing
DNA was extracted from cultured dermal fibroblasts and affected nerve tissue using the QIAamp according to the manufacturer’s instruction (Qiagen). Genomic DNA was separated from leukocytes in peripheral blood using a Puregene blood kit (Qiagen). PIK3CA exon 21 was amplified using the following primers: the forward primer 5ʹ-GTCTACGAAAGCCTCTCTAA-3ʹ and the reverse primer 5ʹ-GCTGTAAATTCTAAGC TGTTC-3ʹ. The polymerase chain reaction (PCR) products were treated with Exo-Sap-IT (Affymetrix) followed by Sanger sequencing. Chromatograms were analyzed using Chromas.
Results
The patient was evaluated by physical examination and X-ray imaging, which showed diffuse enlarged soft tissue of left middle and left fourth fingers with abnormal enlarged phalanges (Figure 1). The patient did not have obvious asymmetry of other parts of the body. Initially, the patient underwent the surgical correction to reduce the bulk of left middle finger with fusion of distal interphalangeal (DIP) and proximal interphalangeal (PIP) joints. Physical examination and X-ray imaging 1 year after the surgery displayed re-enlargement of the left middle finger (Figure 2). Fatty tissue debulking, corrective and osteotomy of the DIP joint, and fixation by K-wire were performed in the left middle finger. A surgical complication included a volar pulp flap ischemia. Clinical course of the patient was showed in Table 1.
Figure 2
Postoperative 1-year follow-up radiologic features of the patient. The radiographs show enlarged middle and fourth fingers of the patient’s left hand (A and B)
Medical history timeline
Time of the investigation
Medical record
07/04/2015
PE* and X-ray imaging: Large soft tissue of the middle and ring fingers of the left hand (Figure 1)
20/09/2015
First surgery: Reduction of the left middle finger with fusion of distal interphalangeal and proximal interphalangeal joints
05/04/2016
PE* and X-ray imaging: Enlargement of subcutaneous, digital nerve, bone, and tendon of the left middle finger (Figure 2)
07/2016
Second surgery: Debulking and correction osteotomy of the left middle finger
PE, physical examination.
PCR sequencing of DNA extracted from the affected nerve tissue and cultured dermal fibroblasts isolated from the affected finger revealed the c.3140A>T in PIK3CA (NM_006218.3 and rs121913279), expected to result in an amino acid substitution at position 1047 changing from histidine to leucine (NP_006209.2: p.(H1047L)). This variant was not detected in her peripheral blood leukocytes (Figure 3).
Figure 3
Mutation analysis of the PIK3CA gene. The sequence electropherograms of the PIK3CA from (A) the cultured dermal fibroblasts, (B) the nerve tissues, and (C) peripheral leukocytes. The c.3140A>T mutation (arrows) is found in the dermal fibroblasts and nerve tissue but not peripheral leukocytes
Discussion
Genetic investigation to identify causative mutations in patients with genetic disorders is important. It will provide more accurate recurrence risk to the patients and families and may lead to more specific treatment. This patient’s parents would like to know the recurrence risk for their next child. The Caucasian patients with isolated macrodactyly have previously been found to have somatic mutations in PIK3CA [6, 7, 8]. Moreover, other mosaic PIK3CA mutations including p.E542K and p.H1047R have been reported as hotspot mutations in macrodactyly patients of other populations [7]. We therefore aimed to determine whether the proband had a variant in the PIK3CA gene. As the mutation in PIK3CA is somatic, DNA from the affected tissues in addition to leukocytes is required. Fortunately, we received tissues surgically removed from the enlarged finger. PCR sequencing identified p.(H1047L) of the PIK3CA in the proband’s nerve and fibroblasts cultured from the affected skin but not in leukocytes. As consistent with previous reports, our study identified the mutation in PIK3CA from both the affected nerve tissue and cultured dermal fibroblasts. The finding suggests that a skin biopsy may be adequate to identify mutations in PIK3CA [9]. This finding confirms that she has the somatic mutation; therefore, the recurrence risk for other family members including future pregnancies is the same as for all other children, which is approximately 0.08/10,000.
The p.H1047L variant in PIK3CA is a gain-of-function mutation [10]. PIK3CA is the upstream regulator of the AktmTOR pathway. The p.H1047 affects the kinase domain, enhances lipid binding, and results in increased kinase activity [11]. Furthermore, p.H1047L of PIK3CA has been found in colon cancer, breast cancer, and ovarian cancer [12, 13, 14]. This leads to a hypothesis that small molecule inhibitors targeting the PI3K-AKT-mTOR pathway may be beneficial to patients with gain-of-function mutations in PIK3CA [13]. According to the previous report, treatment with PI3K/ mTOR inhibitor in transgenic mice harboring a hot spot mutation in PIK3CA restored proliferation rate of endothelial cells to normal levels and reduced numbers of senescent cells [15]. Therefore, genetic determination of macrodactyly in our patient may be beneficial not only for more accurate counselling but also for specific treatment in the future.
Conclusion
As previously identified in other populations, we found the somatic c.3140A>T p.(H1047L) variant of PIK3CA in a Thai patient with isolated macrodactyly in the affected nerve tissue and cultured dermal fibroblasts, suggesting that a skin biopsy may be adequate to identify variants in PIK3CA.
