A 15-year-old boy was referred to University Children's Hospital of Krakow, Department of Oncology and Hematology, in December 2016, because of progressive pallor, bone pain, and upper respiratory tract infection. Physical examination revealed general lymphadenopathy and no hepatosplenomegaly. At admission, severe anemia (hemoglobin level of 6.0 g/dl), leukopenia (white blood cell count of 3.4 x 109/L), and thrombocytopenia (platelet count 45 x 109/L) were found, prompting a bone marrow examination. Bone marrow biopsy revealed 81% of lymphoblasts presenting expression of CD45, CD19, CD22, and CD10; thus, the diagnosis was pre-B-cell precursor ALL (pre-B-ALL). A normal male karyotype (46, XY) was found in the bone marrow cytogenetic analysis. None of the prognostic genetic abnormalities such as BCR-ABL, MLL-AF4, and ETV6-RUNX1 were found in leukemic cells. Cerebrospinal fluid examination revealed no lymphoblasts, and the patient was classified as CSN1 status.
The treatment according to ALL IC-BFM 2009 protocol was started, beginning with cytoreductive prednisone prophase and methotrexate intrathecally. Good response to glucocorticoids was seen. Induction therapy (Protocol IA) was continued afterwards with prednisolone, vincristine, and daunorubicin once per week with four total doses each and Escherichia coli-derived l-asparaginase with eight total doses and methotrexate intrathecally. Bone marrow aspiration on day 15 revealed no blasts (M1 bone marrow) and minimal residual disease (MRD) of 0.05%, indicating a good response to therapy. According to treatment protocol, the patient was ultimately stratified to intermediate risk group. Further treatment according to Protocol IB was continued: four blocks of cytarabine, cyclophosphamide, oral mercaptopurine, and intrathecal methotrexate. During the induction phase, apart from short-term pancytopenia, the patient did not present major toxicities of induction therapy. Complete remission with negative MRD (<0.01%) was achieved on day 78 (at the end of induction).
At the beginning of consolidation, in March 2017, the patient was admitted to our department because of unexplained fever lasting 3 days. On physical examination at admission, disseminated, single, pearly yellow nodules localized on the face, trunk, and extremities (Figure 1); bruises; and significant hepatosplenomegaly were found. Complete blood count revealed severe pancytopenia with hemoglobin level of 6.0 g/dl, leukocytes count of 1.2 x 109/L, and platelet count of 13 x 109/L. Owing to suspicion of pending relapse, bone marrow biopsy was performed. Bone marrow examination confirmed persistent complete remission with negative MRD of <0.01%; however, an increased number of histiocytes with hemophagocytosis (Figures 2, 3) was seen in bone marrow. Flow cytometric analysis of bone marrow cells revealed abnormal population of cells presenting SSChigh and high expression of CD45, CD64, CD14, and CD33, evaluated as macrophages (16% of all nucleated cells). Bone marrow core biopsy was not performed owing to low platelet count and poor coagulation functions not responding to substitution treatment.
Because remission of ALL was confirmed, we started to explore other causes of pancytopenia. Firstly, the patient underwent infectious disease screening. Blood tests revealed elevated C-reactive protein (CRP) (44 mg/dl), with a low procalcitonin level. Blood and urine cultures, as well as viral panel testing for cytomegalovirus (CMV), Epstein-Barr virus (EBV), human herpes virus 6 (HHV6), HIV, parvovirus B19 (PVB19), hepatitis C virus (HCV), hepatitis B virus (HBV), hepatitis A virus (HAV), and herpes simplex virus (HSV), were negative, as were screening examinations for autoimmune and rheumatoid markers.
Within the next several days, the general status of the patient deteriorated, high-grade fever persisted, and hepatomegaly and splenomegaly progressed. Severe hypoproteinemia (30 g/L), hypoalbuminemia (16 g/L), and hypofibrinogenemia (0.6 g/L) was observed. Additionally, an elevated level of ferritin (1,850 mug/L) and CD25 level [soluble interleukin (IL)-2 receptor--6,541 U/ml) were found. Cytotoxic tests of NK cells revealed their reduced activity (6%, normal range 9.9-26.1%). Finally, based on clinical manifestation (fever and splenomegaly) and laboratory tests (pancytopenia, hypofibrinogenemia, hemophagocytosis in bone marrow, high ferritin, and low activity of NK cells), the diagnosis was HLH according to HLH-2004 trial diagnostic criteria.
Considering the probability of secondary HLH with malignancy in the patient, extensive investigations for pathogens were carried out; however, none of the infectious triggers (bacterial, viral, fungal, or parasitic) were found at that time. In the diagnosis of primary HLH, defect in SH2D1A was excluded.
Additionally, owing to the appearance of new skin lesions on the face, trunk, and extremities of the patient, we decided to biopsy the nodules. Skin biopsy revealed diffuse infiltration by histiocytes expressing CD45 and CD68 but negative for CD1a, langerin, and S100 (Figure 4); thus, JXG was recognized. Furthermore, the DNA was extracted from the initial ALL bone marrow smears, with the bone marrow specimen showing histiocytic proliferation and current histiocytic lesions in the skin. The BIOMED-2-primer was used to amplify the immunoglobulin heavy chain gene. The PCR products were analyzed by capillary electrophoresis. There were predominant monoclonal amplifications of 305, 239, and 104 bp found in the bone marrow cells. Identical products were detectable in the histiocytic lesion of the skin, which finally proved clonal relationship between both diseases. No lymphoblasts were found in the skin biopsy.
Despite intensive supportive therapy, patient's status deteriorated gradually. Serious enteral enteropathy with extremely low protein (23 g/L) and albumin (11 g/L) levels resulted in considerable peripheral edemas and hydrothorax. Because the patient stayed in complete remission of ALL with negative MRD status, further therapy according to ALL IC-BFM 2009 was ceased, and treatment, based on HLH 2004 protocol, was introduced. At the beginning, methylprednisolone and etoposide were given; however, no improvement was seen. Thus, cyclosporine and vinblastine were added after 2 weeks. Owing to remarkable fluid retention, cyclosporine was stopped eventually. The patient still needed everyday transfusion of red blood cells, platelet, and plasma concentrates, as well as intravenous immunoglobulin supply. He was fed by gastric feeding tube and then by jejunal tube to fulfill caloric requirements.
In the beginning of April, the patient started to complain about bone pain of left scapula and upper extremities. MRI scan showed bone structure alterations of left clavicula, scapula, and humerus. However, biopsy of the lesion was not done owing to the poor general status of the patient, low platelet count, and persistent coagulation abnormalities.
Despite multidrug, intensive therapy, there was no improvement, neither in patient status nor in bone marrow function or skin lesions. Progression of hepatosplenomegaly was observed. Moreover, the patient developed additional side effects of steroids, such as diabetes, hypertension, and adrenal insufficiency.
Additionally, laboratory findings showed prominent pancytopenia, significantly elevated serum ferritin level, reduced fibrinogen level, and persistent hemophagocytosis in bone marrow. Looking for another treatment modalities of HLH, we decided to apply tocilizumab (IgG anti-IL-6 receptor). Once tocilizumab was given, a slow although unequivocal improvement of skin manifestation was seen. Increase of protein and fibrinogen levels, along with decrease of ferritin concentration, were achieved. Unfortunately, no change in blood parameters and features of hemophagocytosis in bone marrow were observed.
Finally, a total of five tocilizumab doses were given. Because there was no further improvement, the patient was planned to have HSCT. In August 2017, the allogeneic HSCT from matched sibling donor was performed. Patient received treosulfan, fludarabine, thiotepa, and alemtuzumab as a part of his conditioning. Since day 15, reconstitution of leukocyte count > 1 x 109/L has been observed; neutrophils count > 0.5 x 109/L since day 17; and platelet count > 50 x 109/L since day 65 after HSCT. General and nutritional status of the patient gradually improved; however, some nodules on the skin of the face, trunk, and extremities were still present. One month after HSCT, the patient was discharged home. Ten days later, he was admitted to the hospital again owing to severe herpetic mucositis (WHO grade III). Because he started complaining of chest wall pain on the left side, X-ray was performed, and left side pleuritis was found. The pleura was punctured, and fluid examinations revealed high protein concentration and leukocytosis with presence of lymphocytes and macrophages. HHV6 PCR tests were positive, whereas aerobic and anaerobic bacterial, mycologic, and mycobacteria cultures, from both fluid and blood, were negative. Mycobacterium tuberculosis DNA by PCR test was not detected in pleural fluid.
Despite negativity of tests, still, atypical mycobacterial infection was suspected in our patient, as he had been treated with immunosuppressive therapy. Therefore, we decided to introduce antitubercular agents on an empirical basis. Once isoniazid and rifampicin were given, remarkable regression of all skin lesions was observed. Gradual increase of platelet number was also seen within the next weeks. The patient continued with antitubercular drug therapy for further 6 months. Currently, 2 years after HSCT, the patient's general health status is good, in continuous remission of ALL, HLH, and JXG.