A somatic activating NRAS variant associated with kaposiform lymphangiomatosis

Abstract

Purpose: Kaposiform lymphangiomatosis (KLA) is a rare, frequently aggressive, systemic disorder of the lymphatic vasculature, occurring primarily in children. Even with multimodal treatments, KLA has a poor prognosis and high mortality rate secondary to coagulopathy, effusions, and systemic involvement. We hypothesized that, as has recently been found for other vascular anomalies, KLA may be caused by somatic mosaic variants affecting vascular development.

Methods: We performed exome sequencing of tumor samples from five individuals with KLA, along with samples from uninvolved control tissue in three of the five. We used digital polymerase chain reaction (dPCR) to validate the exome findings and to screen KLA samples from six other individuals.

Results: We identified a somatic activating NRAS variant (c.182 A>G, p.Q61R) in lesional tissue from 10/11 individuals, at levels ranging from 1% to 28%, that was absent from the tested control tissues.

Conclusion: The activating NRAS p.Q61R variant is a known "hotspot" variant, frequently identified in several types of human cancer, especially melanoma. KLA, therefore, joins a growing group of vascular malformations and tumors caused by somatic activating variants in the RAS/PI3K/mTOR signaling pathways. This discovery will expand treatment options for these high-risk patients as there is potential for use of targeted RAS pathway inhibitors.

Keywords: exome sequencing; high-throughput sequencing; lymphatic malformation; mosaic; vascular anomaly.

Figure 1.. Clinical-Pathologic Features of KLA

(A-D) Participant KLA2. (A) Extensive cutaneous and subcutaneous involvement of trunk and thigh accompanied by hemorrhage and ascites. (B-D) Magnetic resonance imaging. (B) Coronal T2 fat-saturated image. Hyperintense infiltrative tissue in retroperitoneum and posterior mediastinum (arrow) with heterogeneous high signal of adjacent vertebral bodies. Ascites (asterisk) and multiple cystic lesions in the spleen (curved arrow). (C) Axial T2 fat-saturated image. High signal soft tissue in mediastinum (arrow), extending along bronchovascular bundles. Similar soft tissue abnormalities axillae. (D) Axial T1 contrast enhanced image. Intense enhancement of the abnormal soft tissue. (E-F) Participant KLA3. (E) Thoracoscopic image with extensive visceral pleural involvement. Parietal pleural adhesions also present (not shown). (F) Photomicrograph of pleural surface lesion with thin-walled anastomotic lesional channels with adjacent spindled endothelial cell component and erythrocyte extravasation. (G-J) Participant KLA6. (G) Swollen thigh and perineum with cutaneous telangiectasias and brown discoloration. (H-I) Photomicrographs of skin biopsy with (H) dilated dermal lymphatic channels (asterisk) and small cellular clusters (circle). Higher-magnification of circled area (I) shows complex abnormal lymphatic vessel adjacent to cluster of spindled hemosiderotic cells. (J) D2–40 immunopositivity in spindled cells similar to those in adjacent lymphatic channel. Focal separation of spindled cells suggestive of nascent lumens. (K-N) Participant KLA5. (K) Intraoperative image of anterior mediastinal fibrofatty mass with dilated blood-filled lymphatic channels. (L-N) Photomicrographs of lesional tissue demonstrating (L) several large, often blood-filled lymphatic vessels and anastomosing cellular cords containing interspersed red cells (circle). Higher-magnification of circled area (M) with clusters/ribbons of lymphatic endothelial cells, seemingly canalized and with luminal red cells. (N) Ribbons composed of D2–40 immunopositive lymphatic endothelial cells adjacent to abnormal, dilated lymphatic channels. Although no discernible histopathologic differences were found between participants, this type of rudimentary canalization was more prevalent in tissue sampled at autopsy.

Figure 2.. Exome Sequencing and digital PCR (dPCR) of NRAS c.182A>G, p.Q61R Allele in Participant KLA4

(A) Integrative Genomics Viewer (IGV) screenshot of sequencing reads from lesional tissue exome of Participant KLA4 supporting the variant allele. Variant is on 26/188 reads (14%). (B) dPCR results from lesional tissue of Participant KLA4, demonstrating amplification of variant allele (14%). (C) dPCR results from an unrelated unaffected control, demonstrating no amplification of variant allele.