Belimumab Concentrations in Maternal Serum and Breast Milk During Breastfeeding and the Safety Assessment of the Infant: A Case Study

Abstract

Background: Belimumab is a recombinant human immunoglobulin G1 lambda monoclonal antibody that binds soluble B lymphocyte stimulator protein with high affinity and inhibits its biological activity. Belimumab is not recommended for breastfeeding women due to insufficient data about its excretion into breast milk. In this study, we measured belimumab concentrations in the breast milk of one nursing mother diagnosed with mixed connective tissue disease (MCTD) and evaluated the health of her breastfed infant.

Materials and Methods: Maternal serum and breast milk belimumab concentrations were collected three times (2 weeks after the first dose, the day after the second dose, and 7 weeks after the second dose) after ethical approval and informed consent. An enzyme-linked immunosorbent assay was used to detect belimumab in serum and breast milk samples.

Case Report: A 39-year-old para 4 female was diagnosed with MCTD. The serum concentrations at three times were 29.45, 76.82, and 33.95 mcg/mL. The concentrations in breast milk were 0.12, 0.17, and 0.12 mcg/mL. The milk-to-serum concentration ratios at each sampling point were 0.0041, 0.0022, and 0.0035, respectively. Her infant experienced no health problems. Routine vaccinations were administered without any adverse effects such as infection or immunoreaction.

Discussion and Conclusions: Breast milk levels of belimumab ranged from 1/200 to 1/500 of those in serum, and no harmful effect occurred in her infant. This is the first study reporting belimumab concentrations in human breast milk. Further studies are needed to elucidate the impact of exposure on breastfeeding infants.

Introduction

Belimumab is a recombinant human immunoglobulin G1 lambda monoclonal antibody that binds soluble B lymphocyte stimulator (BLyS) protein with high affinity and inhibits its biological activity.¹ BLyS expression is elevated in patients with systemic lupus erythematosus (SLE) and correlates with disease activity. Approval of belimumab was based on two clinical studies, both of which demonstrated benefit when belimumab was combined with standard of care.^2,3 Because belimumab is a large protein molecule, its amount in milk is likely to be very low and it is probably destroyed in the infant's gastrointestinal tract.⁴ Animal studies have shown that belimumab is excreted in the milk of cynomolgus monkeys.⁵ However, due to species-specific differences, it may not be possible to predict levels in human milk based on animal data. Only one case report has described belimumab use in a female patient with SLE during breastfeeding.⁶ Although there was no harmful effect on her infant, the extent of the infant's breastfeeding was unknown. Thus, more data must be accumulated about the safety of belimumab during breastfeeding. In this study, we measured belimumab concentrations in the breast milk of one nursing mother and evaluated the safety of her breastfed infant.

Materials and Methods

Belimumab detection

An enzyme-linked immunosorbent assay was used to detect belimumab in serum and breast milk samples. Specifically, a microtiter plate was coated with streptavidin, and biotinylated BLyS was added to the wells and bound to the plate. BLyS-reactive belimumab was captured from diluted human serum onto the BLyS-coated plate and detected by the addition of a peroxidase-conjugated secondary rabbit monoclonal antihuman belimumab antibody. Horseradish peroxidase activity was quantified by the color conversion of the substrate tetramethylbenzidine in the presence of hydrogen peroxide. The reaction was stopped using dilute acid and the absorbance was measured at 450/620 nm. Belimumab concentrations in serum and breast milk samples were calculated using a standard curve. Samples and quality controls were analyzed at five dilutions: 1:1, 1:10, 1:100, 1:1,000, and 1:10,000. All results that fell within the range of the standard curve (0.5–200.0 ng/mL) were considered acceptable, and the reported belimumab concentration was the mean of these results multiplied by the dilution factor. Results are reported in mcg/mL. The accuracy (% recovery) and precision (% coefficient of variation) for the set of neat spikes (0.01–20.0 mcg/mL) tested during quantification of the assay were 85.77–106.02% and 2.13–12.14%, respectively. The lower limit of belimumab quantitation in this assay was 0.01 mcg/mL in both serum and breast milk samples.

Ethical approval

This study was approved by the ethics committee of the National Center for Child Health and Development, and the participant provided written informed consent.

Case Report

A 39-year-old para 4 female was diagnosed with mixed connective tissue disease (MCTD) at age 22 years based on signs and symptoms of MCTD and positivity of anti-U1-ribonucleoprotein antibodies. During her latest pregnancy, she was continuously treated with 6 mg/day of prednisolone. After delivery, levels of antidouble-stranded DNA (anti-dsDNA) antibody rose and serum complement levels fell, and the patient exhibited mild proteinuria (30 mg/dL). Therefore, 520 mg (10 mg/kg) of intravenous belimumab and 200–400 mg of daily oral hydroxychloroquine were initiated. She received belimumab injection every 2 weeks for the first two doses followed by the monthly injection. This additional treatment successfully suppressed disease activity and ameliorated the changes in anti-dsDNA antibody and complement levels.

The patient's prednisolone dosage was decreased to 3 mg/day during breastfeeding. Her infant was exclusively breastfed until 6 months of age. Since then, 50–60% of complementary feeding was continued until 15 months of age (457 days). Except for poor weight gain (10 kg at age 2.5 years), the child experienced no health problems. The reason of poor weight gain was unknown. Routine vaccinations, including those for rotavirus, hepatitis B, Haemophilus influenzae type b, and pneumococcal conjugate (PCV13), were administered without any adverse effects such as infection or immunoreaction.

Maternal serum and breast milk belimumab concentrations were measured three times (2 weeks after the first belimumab dose, the day after the second dose, and 7 weeks after the second dose). As given in Table 1, the maternal serum concentrations before belimumab dose, 2 weeks after the first dose, and 7 weeks after second dose were 29.45, 76.82, and 33.95 mcg/mL, respectively. Breast milk samples were collected at the same time as blood samples. The belimumab concentrations in breast milk were 0.12 mcg/mL 2 weeks after the first dose (before the second dose), 0.17 mcg/mL on the following day, and 0.12 mcg/mL at 7 weeks after the second dose. The milk-to-serum concentration ratios at each sampling point were 0.0041 (0.12/29.45), 0.0022 (0.17/76.82), and 0.0035 (0.12/33.95), respectively. The mean relative infant dose (RID) of belimumab was calculated from the therapeutic dose for adult and children aged 5 years and older (10 mg/kg),⁵ average belimumab concentration (0.14 mcg/mL) in breast milk, and the standard milk intake [150 mL/(kg·d)].⁷ The standard therapeutic dose was divided by the dosing interval of belimumab (14 or 28 days) to convert the weekly dose into the daily dose. As the result of the calculation, mean RID through breast milk was 3.67%.

Table 1.

Belimumab Concentrations in Serum and Breast Milk

Time course
Days postpartum	+395	+409	+410	+411	+451
Belimumab i.v.	520 mg		520 mg
Days after belimumab	0	14	15	1	28
Maternal (mcg/mL)
Serum	—	29.45	—	76.82	33.95
Breast milk	—	0.12	—	0.17	0.12
Milk-to-serum ratio	—	0.41%	—	0.22%	0.35%

i.v., intravenous.

Discussion and Conclusions

Breast milk levels of belimumab ranged from 1/200 to 1/500 of those in maternal serum, which was similar to the other biologic agents previously reported.^8–10 The mean RID of belimumab was 3.67%. Although the dosing interval of belimumab for pediatric patients is 2–4 weeks, and the administration route is different, the RID did not exceed 10% of the maternal dose. These results suggest that exposure of belimumab through breast milk is usually not considered to be clinically relevant for the infant. In our case, belimumab concentration in breast milk at the day after the second dose was same level at 14 and 28 days after a dose. Transfer of other monoclonal antibodies into breast milk takes a certain amount of time, and they have peak concentrations a few days after the dose.^9–11 The measured concentration in breast milk at day 1 appears to be equivalent to the trough concentration. Our data from only one nursing mother with MCTD are limited, and further studies are needed to elucidate the impact of exposure on breastfeeding infants.

Disclosure Statement

The authors haveno conflicts of interest.

Funding Information

This study was supported by the Research Program from the Japanese Agency for Medical Research and Development (JP19mk0101086h0003) awarded to Atsuko Murashima and by a domestic research and development grant from National Center for Child Health and Development (2019C-6) awarded to Jumpei Saito.