Compound heterozygous LPIN2 pathogenic variants in a Majeed Syndrome patient with recurrent fever and severe neutropenia: case report CURRENT STATUS:

Background Majeed syndrome is a rare, autosomal recessive autoinflammatory disorder first described in 1989. The syndrome starts during infancy with recurrent relapses of osteomyelitis typically associated with fever, congenital dyserythropoietic anemia (CDA) and often neutrophilic dermatosis. Mutations in the LPIN2 gene located on the short arm of chromosome 18 have been identified as being responsible for the Majeed syndrome. Case presentation We report a 8-month old boy, who presented with recurrent fever, mild to moderate anemia and severe neutropenia. Erythrocyte sedimentation rate and C-reactive protein were elevated. Molecular studies identified a paternal splicing donor site variant c.2327+1G>C and a maternal frameshift variant c.1691_1694delGAGA (Arg564Lysfs*3)in LPIN2. Conclusions only mutation The pallor no lymphadenopathy or hepatosplenomegaly. Blood routine examination showed severe neutropenia (0.38-0.40 ×10 9 /L) with normal white blood cell count, microcytic anemia (hemoglobin 85-95g/L) and slight thrombocytosis. The boy had elevated erythrocyte sedimentation rate (79mm/h) and C-reactive protein (39mg/L, normal<8mg/L). Detection of immunoglobulin and lymphocyte subset were normal. Rheumatoid factors were negative. Antinuclear antibody was positive with a titer of 1:80while the anti-ds DNA antibody was negative. Antineutrophil cytoplasmic antibodies revealed with mild elevated anti-MPO antibody (30.2RU/mland negative anti-PR3 antibody. Measurement of Thyroid Function was normal. Serum iron and transferrin were decrease, which indicated iron-deficiency anemia. Bone marrow hemocytology revealed myeloproliferative, the proportion of myelocyte was decreased in the granulocytosis; Red blood cell is proliferous and active, metarubricyte was dominated with small, hollow and distorted mature erythrocyte. Blood and bone marrow puncture specimens had been cultured for bacteria and fungi, and had showed no growth. Results of viral serologic studies also had been negative. Lymphocyte interferon release assay was negative. Abdominal ultrasound scan gave normal findings. Cardiac ultrasound showed no abnormalities. Chest CT showed no interstitial or parenchymal infiltration.


Background
Majeed syndrome is a rare, autosomal recessive autoinflammatory disorder first described in 1989.
The syndrome starts during infancy with recurrent relapses of osteomyelitis typically associated with fever, congenital dyserythropoietic anemia (CDA) and often neutrophilic dermatosis. Mutations in the LPIN2 gene located on the short arm of chromosome 18 have been identified as being responsible for the Majeed syndrome.

Case presentation
We report a 8-month old boy, who presented with recurrent fever, mild to moderate anemia and severe neutropenia. Erythrocyte sedimentation rate and C-reactive protein were elevated. Molecular studies identified a paternal splicing donor site variant c.2327+1G>C and a maternal frameshift variant c.1691_1694delGAGA (Arg564Lysfs*3)in LPIN2.

Conclusions
Up to now, only a few cases with LPIN2 mutation have been reported, mainly in the Middle East with homozygous variants. Our patient exhibited a mild clinical phenotype and severe neutropenia, distinct from previously reported.

Background
Majeed syndrome is a rare, autosomal recessive autoinflammatory disorder first described in 1989.
The syndrome starts during infancy with recurrent relapses of osteomyelitis typically associated with fever, congenital dyserythropoietic anemia (CDA) and often neutrophilic dermatosis. Mutations in the LPIN2 gene located on the short arm of chromosome 18 have been identified as being responsible for the Majeed syndrome. Here we report, to our knowledge, the first case of Majeed syndrome in individual of Chinese heritage and with variable severity.

Clinical information
This Chinese 8-month old boy presented at the age of 6 month with recurrent fever lasting for 5-7 days and recurrent every 3-7 days. Sometimes he had a slight cough. He had no physical pain or movement problems. He had no rash and other symptoms. The infant was born at full term, normal delivery with a birth weight of 3.0Kg. His parents were not consanguineous marriages. There was a neonatal history of jaundice. The boy had mild pallor when he was admission to our hospital. He had no lymphadenopathy or hepatosplenomegaly. Blood routine examination showed severe neutropenia (0.38-0.40 ×10 9 /L) with normal white blood cell count, microcytic anemia (hemoglobin 85-95g/L) and slight thrombocytosis. The boy had elevated erythrocyte sedimentation rate (79mm/h) and C-reactive protein (39mg/L, normal<8mg/L). Detection of immunoglobulin and lymphocyte subset were normal.
Rheumatoid factors were negative. Antinuclear antibody was positive with a titer of 1:80 while the anti-ds DNA antibody was negative. Antineutrophil cytoplasmic antibodies revealed with mild elevated anti-MPO antibody (30.2RU/ml and negative anti-PR3 antibody. Measurement of Thyroid Function was normal. Serum iron and transferrin were decrease, which indicated iron-deficiency anemia. Bone marrow hemocytology revealed myeloproliferative, the proportion of myelocyte was decreased in the granulocytosis; Red blood cell is proliferous and active, metarubricyte was dominated with small, hollow and distorted mature erythrocyte. Blood and bone marrow puncture specimens had been cultured for bacteria and fungi, and had showed no growth. Results of viral serologic studies also had been negative. Lymphocyte interferon release assay was negative. Abdominal ultrasound scan gave normal findings. Cardiac ultrasound showed no abnormalities. Chest CT showed no interstitial or parenchymal infiltration.

Molecular genetic studies
After obtaining informed consent, we isolated DNA from peripheral blood samples obtained from the patient and parents by using the Gentra Puregene Blood Kit (QIAGEN, Hilden, Germany). Whole exome library was captured by SureSelect Human All Exon Kit (Agilent Technologies, Santa Clara, America) according to the manufacturer's instructions. Target regions were sequenced and aligned to the GRCh37/hg19 human reference sequence. Variants were annotated and filtered by TGex (tgexapp.genecards.cn). Variants were classified following the ACMG/AMP standards and guidelines 1 .
Putative pathogenic variants were confirmed by Sanger sequencing.
c.2327+1G>C is not reported in dbSNP, 1000 genome, ESP, ExAC or gnomAD databases, indicating it is very rare in normal population. This variant was first reported in a Arabic family, the proband was a 3-year-old girl with Majeed Syndrome [2] . This variant was predicted to disarrange the donor site and caused exon 17 deletion or intron 17 insertion either entirely or partly. c.1691_1694delGAGA located in exon 12, leading to premature termination codon at position 3 amino acids after mutation. It is expected to produce a truncated protein or lead to early degradation of mRNA through the mechanism of nonsense-mediated decay. This variant was not reported in dbSNP , ESP or 1000 genome databases. The frequency in ExAC database was 0.000008236, suggesting that the frequency was extremely low. Both variants were classified as pathogenic variants according to ACMG/AMP guidelines.
Majeed syndrome is a rare autosomal recessive disorder characterized by chronic recurrent multifocal osteomyelitis (CRMO). This is an early-onset disorder with a lifelong course and congenital dyserythropoietic anemia (CDA) that presents as hypochromic, microcytic anemia during the first year of life and ranges from mild to transfusion dependent. Some individuals also develop a transient inflammatory dermatosis, often manifesting as Sweet syndrome (neutrophilic skin infiltration). It is often accompanied by recurrent fever. The diagnosis is based on clinical findings and molecular genetic testing of LPIN2, the only gene in which pathogenic variants are known to cause Majeed syndrome. Up to now, only a few cases with LPIN2 mutationhave been reported, mainly in the Middle East with homozygous variants 2-9 .
LPIN2 encodes a phosphatidate phosphatase which plays important roles in controlling the metabolism of fatty acids at different levels. The function of LPIN2 is not well known. According to pervious study, it acts as a magnesium-dependent phosphatase converting phosphatidic acid to diacylglycerol in the biosynthesis of triglycerides, phosphatidylcholine and phosphatidylethanolamine.
It can also act as a nuclear transcriptional coactivator of PPARGC1A and therefore, regulate lipid metabolism 10,11 . Homozygous knock our Lpin2 in mice displayed increase in mean platelet volume, red blood cell distribution width and lymph nodes, decrease in mean corpuscular hemoglobin, bone mineral density and content. They also had abnormal circulating phosphate level, hydrometra, and preweaning lethality with incomplete penetrance. LPIN2 has 19 exons and three lipin domains located in N-terminal, middle and C-terminal, respectively (Fig. 2). Lipin domains are highly conserved in lipin proteins, in which when mutated in mice lead to fatty liver dystrophy.
Here we report, to our knowledge, the first case of Majeed syndrome in individual of Chinese heritage and with variable severity. Our patient exhibited a mild clinical phenotype, distinct from previously reported cases (Table1). He had recurrent fever, mild to moderate hypochromic and microcytic anemia without severe CRMO. He had no physical pain, swelling or movement disorders. Majeed reported a Palestinian Arab boy whose presented at the age of 2 months with recurrent episodes of high fever and irritability 12 . At the age of 9 months, these episodes started to be associated with periarticular swellings with hotness, tenderness and limitation of movement. Therefore, the symptoms and signs of our patient's need to be observed continuously. Our patient had severe neutropenia from 6 months and his absolute neutrophil count was 0.38-0.40×10 9 /L . This phenotype has been reported in few cases. Mosawi reported an Arabic female with Majeed syndrome who had mild neutropenia (1.08×10 9 /L ) in the neonatal period 3 . RAO reported a 15-year-old boy with Majeed syndrome complicated by mild neutropenia 5 . Those cases suggest that neutropenia may be part of the phenotype. More cases are needed to be observed to confirm this phenotype.

Conclusions
Majeed syndrome is an autosomal recessive, autoinflammatory disorder. It characterized by CRMO and CDA. Our patient exhibited a mild clinical phenotype, distinct from previously reported cases.