Repeated failed non‐invasive prenatal testing owing to low cell‐free fetal DNA fraction and increased variance in a woman with severe autoimmune disease

Abstract

A 41-year-old woman (gravida 2 para 1) with autoimmune disease was referred for genetic counseling following an increased risk of trisomy 21 (1:164) on combined first-trimester screening. She had a history of severe autoimmune thrombocytopenia, with past treatments including splenectomy, steroids, intravenous immunoglobulin (IVIG) and cyclosporin. She also had an autoimmune neutropenia, a past history of myasthenia gravis treated with thymectomy and Hashimoto's thyroiditis. Her weight was 67 kg. Given her low platelet count of 36 × 109/L, the patient elected to have non-invasive prenatal testing (NIPT) at 15 weeks rather than an amniocentesis for the risk of trisomy 21. The NIPT did not return a result owing to a low fetal fraction (< 4%). A repeat NIPT test was discussed, with the advice that > 50% of women with a failed assay have a successful result on redraw1. A repeat NIPT sample was therefore sent at 17 weeks to the same provider. This sample also failed because of a fetal fraction of < 4%. Owing to the turnaround time for offshore NIPT processing, the patient was 19 weeks at the time of the second failed NIPT. After counseling, the patient decided to proceed with amniocentesis. The autoimmune disease in this patient was considered a likely cause for the persistently low fetal fraction at 15 and 17 weeks. We hypothesized that the steroids and IVIG used to treat her thrombocytopenia and neutropenia before the planned amniocentesis might also improve the fetal fraction by suppressing maternal cell destruction. With Research Ethics Committee approval, a third NIPT sample was taken, after 7 days of oral steroids, at 21 + 0 weeks. Her platelet count improved, from 36 × 109/L to 98 × 109/L. A fourth NIPT sample was taken, 2 days later, after an IVIG infusion (1 mg/kg). The third NIPT sample taken after the steroid course successfully returned a result of ‘low risk’ for trisomies 21, 18 and 13 (fetal fraction, 4.6%). The fourth NIPT sample, taken after the IVIG infusion (21 + 2 weeks) did not return a result owing to ‘unusually high variance in cell-free DNA counts’. The fetal fraction was not reported. The final karyotype from the amniocentesis was normal. The mean fetal fraction at 11–14 weeks is 10%1; approximately 2% of women will not receive an NIPT result because of a low fetal fraction2, 3. The successful NIPT result after the course of steroids in this patient suggests that the fetal fraction was increased by immunosuppression. However, this increase cannot be confidently distinguished from a background gestational age-related increase in fetal fraction, estimated at 0.1% per week3. The laboratory would not reveal the fetal fractions from the tests at 15 and 17 weeks. The high variance in cell-free DNA counts in the fourth sample, post-IVIG, suggests specific interference from this therapy. The biological basis of ‘increased variance’ is still unknown. The multiple NIPT failures in this patient suggest that caution is advised when counseling women with severe autoimmune disease about NIPT. Unfortunately, these patients frequently have conditions that make avoidance of invasive procedures particularly desirable. L. Hui*†‡, M. Bethune§¶, A. Weeks§, J. Kelley† and L. Hayes†** †Department of Perinatal Medicine, Mercy Hospital for Women, Heidelberg, VIC, Australia; ‡Department of Obstetrics & Gynaecology, University of Melbourne, Parkville, VIC, Australia; §Specialist Women's Ultrasound, Box Hill, VIC, Australia; ¶Department of Medical Imaging, Mercy Hospital for Women, Heidelberg, VIC, Australia; **Clinical Haematology Service, Northern Health, Epping, VIC, Australia *Correspondence. (e-mail: lisahui77@gmail.com)