A 19-year-old woman with 32+3 weeks of gestation was referred to our hospital due to oligohydramnios. The patient had a history of myomectomy at age 15. At that time of ultrasound examination, there was a mass of 20.0 cm* 8.7 cm in size in the pelvic cavity. Postoperative pathological findings showed cellular uterine leiomyoma.
On admission, both the patient and the fetus were in good condition. Physical examination revealed huge mass in pelvic cavity. Abdominal and pelvic ultrasound confirmed the presence of multiple masses in pelvic, sized 16.9*11.2*10.1cm, 13.1*5.6*6.2cm, 19.2*17.5*12cm respectively next to the gestation without signs of abortion. The masses were connected into large clumps. Abdominal MRI was done to show multiple nodules in the abdominal cavity (Figure 1).
In order to ascertain the diagnosis, an exploratory laparotomy was performed because of aggravated abdominal pain. After delivery the fetus by lower-segment cesarean section, the gynecological oncologist performed further operation. The patient was found to have multiple sporadic leiomyoma in anterior wall of uterus, an 8*6cm leiomyoma in posterior wall of uterus, a 20*15 cm tumor mass in the left pelvis, multiple tumor masses in the right pelvic sized 8*7cm, 7*7cm and 7*5cm separately, up to 10 tumor masses sized 3*2cm in omentum and mesocolon transversum (Figure2 A-D). All macroscopic tumor masses were dissected and removed via an extremely difficult surgery without hysterectomy and bilateral salpingo-oophorectomy because of the patient’s strong objection and the consideration of young age. Post‑operative pathology determined the diagnosis of LPD with red degeneration. The patient recovered well after surgery and was discharged on the ninth day after removal of the abdominal incision suture.
The patient underwent several ultrasound examinations after surgery and no signs of disease recurrence were found without any continuous treatment. The patient was pregnant again 25 months after the surgery. At 7 weeks of gestation, ultrasound examination revealed a fibroid of about 3.7cm*3.7cm in the posterior wall of the uterus, and ultrasound examination during pregnancy indicated that the fibroid was slowly enlarged without any discomfort symptoms. The patient underwent cesarean section again at 39 weeks of gestation. No abnormal lesions were found in the pelvic and abdominal cavity during the operation, and only a uterine fibroid of about 7cm*6cm was found in the posterior wall of the uterus (Figure 2E). Postoperative pathology suggested uterine leiomyoma. The patient was reviewed at 6 months postoperatively and recovered well.
2.2 NGS (next generation sequencing)
We collected 15 of 4μm tissue sections from FFPE samples of LPD and normal tissue adjacent to the lesion for the genetic analyses. QIAamp DNA FFPE Tissue Kit (QIAGEN, Heidelberg, Germany) was used to extract genomic DNA according to the manufacturer’s instructions.
DNA was profiled using a commercial available capture-based targeted sequencing panel (Burning Rock Biotech Ltd, Guangzhou, China), targeting 295 genes which were closely related to the mechanism of cancer and targeted therapy and spanning 1.5MB of Human genomic regions. DNA shearing, end repair and adaptor ligation was performed by the use of Covaris M220 (Covaris, Inc., MA, US). Fragment sizes ranging from 200bp to 400bp were selected using Agencourt AMPure beads (Beckman Coulter, CA, US) followed by hybridization with capture probes baits, hybrid selection with magnetic beads and PCR amplification. Subsequently, Qubit® 3.0 and Agilent 2100 bioanalyzer (Agilent Technologies Inc., CA, US) was performed to assess the quality and size of the fragments. Indexed samples were sequenced on Nextseq500 sequencer (Illumina, Inc., CA, US) with pair-end reads.
Based on the high throughput sequencing, the copy numbers (CNs) of this LPD patient compared with normal population were demonstrated in Figure 3G. There were four somatic cell lines mutations detected in the lesions. The CNs of CDK4, NBN, DAXX and MYC were all amplified for at least 4 times.
2.3 Hematoxylin-eosin (HE) and immunohistochemistry staining
Hematoxylin-eosin (HE) staining slides of this LPD were shown in Figure 3A. Rich blood supply was revealed in LPD in HE staining analysis (Figure 3B).
Immunohistochemistry staining showed that the tumor was strongly positive for smooth muscle markers, SMA and Desmin (Figure 3C, 3D), which suggested that LPD shared partial molecular cytogenetic characteristics with uterine leiomyoma. Immunohistochemistry of hormone receptors, estrogen receptor (ER) and progesterone receptor (PR) were positive (Figure 3E, 3F).
The immunohistochemistry staining analysis of CDK4, MYC, NBN and DAXX in uterine leiomyoma (10 cases), LPD (4 cases) and leiomyosarcoma (10 cases) revealed that the expression profiles of LPD were more similar to leiomyosarcoma. LPD showed CDK4, NBN, DAXX, MYC moderately and strongly positive and uterine leiomyosarcoma displayed strongly positive. However, the four markers in uterine leiomyoma were slightly positive or negative. Therefore, we can infer the conclusion that LPD is an intermediate disease between benign uterine fibroids and malignant leiomyosarcoma.