Unusual presentations of sitosterolemia limited to hematological abnormalities: A report of four cases presenting with stomatocytic anemia and thrombocytopenia with macrothrombocytes
Authors: Jie Zheng, Jie Ma, Run‐Hui Wu, Xue Zhang, Yan Su, Rui Zhang, Li‐Qiang Zhang
Journal: American Journal of Hematology (2019)

To the Editor: We present here four cases of sitosterolemia characterized only by hematologic abnormalities without cutaneous xanthomas or atherosclerosis. Diagnosis was delayed by an average of 2.5 years (from 5 months to 9 years) and was preceded by implementation of incorrect therapeutic strategies. Next generation sequencing (NGS) results showed deleterious mutations in ABCG5/8, with six out seven mutations being novel. All our patients achieved hematologic completed response (CR) after ezetimibe therapy. Case 1: A 10-year-old girl was admitted with thrombocytopenia and anemia (Table 1). She had been diagnosed with Evans syndrome at 7 years old. At the time of the first admission, lab examination revealed a high absolute reticulocyte count, a positive Coombs test, and a normal bone marrow smear with increased reactive megakaryocytes and M/E = 1.8:1. She had been treated with IVIG and prednisone for almost 1 year with fluctuating platelet (7 × 109/L to 148 × 109/L) and hemoglobin (Hb) (94 g/L to 113 g/L) levels. Enlargement of cervical lymph nodes and hepatosplenomegaly were present from 8 years of age. The diagnosis of autoimmune lymphoproliferative syndrome (ALPS) was suspected. High-dose intravenous methylprednisolone (starting at 20 mg/kg) had been initiated, followed by oral prednisone (1-2 mg/kg). Platelet count had reached 105 × 109/L temporarily and Hb 115 g/L, but the size of the spleen continued to increase slowly. Peripheral blood smears at admission revealed stomatocytes (about 10%) and giant platelets (GTs); some GTs were as large as the erythrocytes (Supporting Information Figure S1A,B). A novel c.1915delA (p. I639X) and a reported c.1336C>T (p. R446X) compound heterozygous mutations were found in ABCG5 by NGS and were confirmed by the Sanger sequencing. Measurement of plasma phytosterols revealed a high concentration of sitosterol, stigmaterol, and campesterol (Table 1), which supported the diagnosis of sitosterolemia (STSL). Case 2: A 14-year-old boy presented to our clinic for pancytopenia (Table 1). He was diagnosed with ITP at 5 years old. After prednisone therapy for 6 months and supplementary Traditional Chinese Medicine (TCM) therapy for 1.5 years, his platelet counts fluctuated between 30 × 109/L and 50 × 109/L without any substantial improvement and the family decided to stop all treatments. At 14 years of age, appearance of jaundice led to the diagnosis of hepatitis. After TCM therapy for almost 5 months, his liver function recovered, but his WBC level dropped to 3.02 × 109/L, ANC 1.81 × 109/L, Hb 115 g/L, platelet 2 × 109/L, raising the suspicion of hepatitis-associated aplastic anemia (HAAA). At admission in our department, the bone marrow smear showed increased cellularity, with enlarged platelets in the peripheral blood smear, while erythrocytes and WBC looked normal in shape. NGS results showed ABCG5 compound heterozygous mutations: c.1118+3A>C and c.904+1G>A. High concentrations of phytosterols confirmed the diagnosis of STSL. Case 3: An 11-year-old girl came to our department for a second opinion. She had intermittent epistaxis, diagnosed as ITP, and had been treated with IVIG, prednisone, cyclosporine, and Danazol for almost 2 years. The platelet level fluctuated between 8 × 109/L and 87 × 109/L. Some large platelets cells and stomatocytes were observed on the peripheral blood smear. NGS showed ABCG8 compound mutation with c.683T>C; p. L228P and c.1724G>A; p. G575D, and a high concentrations of phytosterols were found in the plasma, supporting the diagnosis of STSL. Case 4: A 15-year-old girl was referred to our department with a history of thrombocytopenia. She was born from healthy, nonconsanguineous parents. She had been diagnosed with ITP, received IVIG, prednisone, and dexamethasone for almost 3 months without any response. Her platelet level reached normal only after platelet transfusion. The CBC showed WBC 6.61 × 109/L, Hb 76 g/L, MCV 68.1 fL, Plt 10 × 109/L, Ret 7.4%. Poikilocytosis and large platelet cells were noticed on the peripheral blood smear. Homogeneous mutations with ABCG5 (c.751G>A; p. Q251X) and high plasma phytosterols were found. She had a history of β-thalassemia, Henoch-Schönlein purpura, and diabetes insipidus. Cranial MRI revealed a uniform soft tissue density mass (10 × 9 mm) with unclear boundaries and no obvious enhancement in her hypothalamus. All four patients were started on ezetimibe therapy (10 mg, daily) and a low plant sterol diet after the diagnosis of STSL. They achieved hematologic CR after a median of 6 months therapy (4 weeks to 11 months). Sitosterol and campesterol levels were reduced significantly, while hepatosplenomegaly gradually improved. At the last visit, all patients were in hematologic CR; low stomatocytes with GTs were still be observed in some cases (Supporting Information Figure S1C). The hypothalamic mass in case 4 almost disappeared and a very low dosage of desmopressin achieved control of her diabetes insipidus after 2 years of ezetimibe therapy. The primary clinical manifestations of STSL (OMIM #210250) are cutaneous xanthoma, premature atherosclerosis, and its associated complications.1 All four patients reported here were diagnosed with STSL due to severely elevated levels of plasma phytosterols, ascertained by HPLC, which are the biochemical hallmark of this disease.1 Pathological examination of STSL patients had shown that xanthomas seemed to be formed by phytosterols, even at lower plasma levels (30-40 mg/dL).2 An in vitro experiment seems to show that the hematologic abnormalities would be due to the toxic effect of plant sterols on blood cells.3 All of our four patients had high plasma phytosterols but without cutaneous xanthomas or coronary atherosclerosis. To date, but for our four patients, only eight unrelated STSL patients have previously been reported to present with hematologic abnormalities without xanthomas at the time of diagnosis (Supporting Information Table 1). The underlined mechanisms in these special STSL patients are unclear so far. All of the four patients here were misdiagnosed as ITP. The median time from symptom onset to the STSL diagnosis was 2.5 years (from 5 months to 9 years). In fact, all of the patients showed mild to moderate anemia with elevated reticulocytes. That meant they might be afflicted with hemolytic anemia. Most pediatric doctors would suspect Evans' syndrome or autoimmune disease when there is a coexistence of hemolytic anemia and thrombocytopenia. The diagnosis of these four patients and other similar cases4, 5 showed that unexplained hemolytic anemia and thrombocytopenia might also be the presentation of STSL. The most significant difference between Evans' syndrome (or other autoimmune disease) and STSL appeared on the blood films. Almost all patients' blood films here showed stomatocytes and macrothrombocytopenia. In general, the coexistence of stomatocytes and macrothrombocytopenia was very rare, reported only in STSL and MDS. It should be emphasized that careful examination of the peripheral blood smear is the most important aspect of the diagnostic approach to STSL. STSL is caused by mutations in either of ABCG5 or ABCG8.1 Seven variants were found in our patients, with only the R446X being previously reported. Although 28 variants have previously been reported in the ABCG5 gene, only 11 variants are associated with macrothrombocytopenia (supplemental Figure 2, asterisks), among which only six variants, including R446X, are presented in more than one patient (supplemental Figure 2, green highlights). It seems that exon 9 is the relevant hot region for ABCG5 associated STSL. No mutations had been reported in exon 5 until now. Ezetimibe, an NPC1L1 inhibitor that effectively inhibits the intestinal absorption of both cholesterol and phytosterols, is currently considered the treatment of choice for STSL.6 Long-term treatment with ezetimibe was safe, tolerable, and effective in reducing phytosterol concentrations and shrinking the cutaneous xanthomas. Our patients all accepted ezetimibe with no obvious side effects during the median follow-up period of 9 months (8 month to 24 month). It seems like ezetimibe would improve stomatocytic anemia and thrombocytopenia with macrothrombocytes: increased RBC and platelet number, reduced platelet size, and improved the shape of RBCs. Whether normalization of phytosterols is essential to protect against cardiovascular disease, or whether the total cholesterol normalization is sufficient, has not yet been clearly evaluated. Our patients showed no cutaneous xanthomas or any sign of cardiovascular disease until the median age of 13 years old, but they are still at risk of cardiovascular diseases considering that they have higher plasma phytosterols. Therefore, long-term and cautious follow-up of these patients is recommended. We are thankful to all of the patients who participated in this study and to all of our colleagues at our center. This work was supported in part by funds from Beijing Children's Hospital Young Investigator Program BCHYIPA-2016-12. All authors declare no competing financial interests. Fig S1 peripheral blood smear of sitosterolemia patients before and after Ezetimibe Figure S2. ABCG5 gene structure. Table S1. Gene mutations, clinical features, therapy and the outcomes of sitosterolemia patients with only hematology abnormalities Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. 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