A 42-year-old woman with a history of B-LBL was admitted to the Institute of Hematology and Blood Diseases Hospital, Tianjin, China, because of persistent dyspnea. In May 2019, the patient was evaluated at another hospital for recurrent pain in her right hip, which had begun 6 months earlier and gradually worsened with no obvious predisposing causes. PET-CT scans revealed multiple high-density nodules in both the kidneys, left acetabulum, left ilium, bilateral tibia, and right fibula as well as intense FDG uptake by these nodules. Immunohistochemical (IHC) analysis of a left kidney biopsy specimen indicated positivity for CD10, TdT, CD79a, Pax-5, CD43, c-myc (20%), and Ki-67 (90%) and negativity for CD3, CD20, CD5, CD23, Bcl-2, Bcl-6, MUM1, and CyclinD1. Neither bone marrow smear nor flow cytometry revealed any abnormal lymphocytes, and bone marrow biopsy revealed active proliferation of the hematopoietic tissue. Cytogenetic analysis revealed an abnormal karyotype: 46,XX,t(1,19)(q23;p13),-4,+mar[18]/46,XX(2). Gene fusion analysis showed positivity for E2A/PBX1. These findings led to the diagnosis of B-LBL with E2A/PBX1.
The patient had received two cycles of induction chemotherapy with a combination of cyclophosphamide, vincristine, doxorubicin, dexamethasone (hyper-CVAD A), methotrexate, and cytarabine (hyper-CVAD B). Bone marrow examination revealed 10% blasts, while PET-CT revealed no intense FDG uptake. Subsequently, the patient received vindesine, daunorubicin, cyclophosphamide, pegaspargase, prednisone (VDCLP), cyclophosphamide, cytarabine, 6-mercaptopurine (CAM), vindesine, cyclophosphamide, idarubicin, dexamethasone (VICD). Every bone marrow smear after each chemotherapy showed CR, while E2A/PBX1 was undetectable, 3.68% and 0.22% as determined via polymerase chain reaction, respectively.
The patient remained well for 3 months. In May 2020, bone marrow smear revealed 47% lymphoblasts, while the rate of E2A/PBX1 rearrangement was 45.55%. Lymphoblasts accounted for 4.96% of the marrow cellularity, as determined using flow cytometry. These results suggested a diagnosis of B-ALL with E2A/PBX1 expression. Treatment with orally administered 6-mercaptopurine and prednisone was initiated to relieve the tumor burden.
In June 2020, the patient underwent allo-HSCT from a matched, sibling donor after a conditioning regimen with cyclophosphamide, fludarabine, idarubicin, and antithymocyte globulin and total body irradiation. Early post-transplantation complications included mild diarrhea and mucositis. The bone marrow smear indicated CR, no E2A/PBX1 amplification, and full donor chimerism. PET-CT revealed that the treatment was effective and that the CR of the disease was achieved, with a Deauville score of 1 point.
In September 2020, the patient complained of left breast pain that had begun 1 week earlier. Breast ultrasound showed multiple nodules in the left breast, while IHC of a left breast biopsy specimen indicated extramedullary recurrence of B-ALL after transplantation. Neither bone marrow aspirate nor flow cytometry showed any abnormal blasts, while E2A/PBX1 was undetectable. PET-CT revealed progressive disease, with a maximum standard uptake value (SUVmax) of 23.0 and a Deauville score of 5 points ( Figure 1A ). Then, the administration of decitabine, fludarabine, cytarabine, and granulocyte colony-stimulating factor was initiated. Acute graft-versus-host-disease of the skin occurred after donor lymphocyte infusion. Eventually, the breast nodules decreased in size, and PET-CT revealed PR of the disease, with a Deauville score of 4 points.
In November 2020, the patient underwent donor-derived anti-CD19 CAR T-cell infusion after a conditioning regimen with fludarabine and cyclophosphamide. The sequence of CD19 CAR consists of an anti-CD19 single-chain antibody fragment (FMC63), CD28 transmembrane domain, 4-1BB, CD3zeta, T2A autocleavage sequences, and endodomain-deleted EGFR (tEGFR). Four days later, the patient had a recurrent high fever with a temperature of 40 C, along with lethargy and vomiting. Based on a previous study, a grade-2 cytokine release syndrome (CRS) was identified. These clinical issues improved after providing supportive care with antibiotics (meropenem and caspofungin) and dexamethasone. Meanwhile, no clear etiological evidence with next-generation sequencing (NGS) or blood culture was obtained. The patient achieved CR and was negative for minimal residual disease on the 14th day after CAR T-cell therapy and remained well for 5 months.
In May 2021, the patient complained of chest pain that worsened when the patient breathed in or lay flat. Electrocardiogram (ECG) and myocardial enzyme levels were normal. Echocardiography (Echo) revealed massive pericardial effusion with rapid progression to cardiac tamponade. Ultrasound-guided pericardiocentesis, which was performed to alleviate the clinical symptoms and drain the excess fluid around the heart, revealed the presence of a hemorrhagic and exudative liquid. Multiple diagnostic tests for pericardial effusion, including bacterial gram staining, bacterial culture, fungal culture, acid fast bacteria staining, tuberculosis (TB) antibody examination, biopsy, and NGS, were performed. The results of all the above mentioned tests were negative. Flow cytometry of pericardial effusion and bone marrow aspirate did not reveal any tumor cells or CAR T cells. However, quantitative real-time polymerase chain reaction (qPCR) revealed that the copy number of CAR DNA per microgram of genome was 1.92 x 101. The results of additional laboratory tests, including those of cytokines, are shown in  Table 1 . Given that some cytokines of the pericardial fluid were notably increased, CRS was considered as the possible cause of pericardial effusion. The pericardial effusion was gradually reduced and removed after immunosuppressive therapy with methylprednisolone and rucotinib. Bone marrow aspirate and PET-CT showed CR, with a Deauville score of 1 point.
On presentation to our hospital (November 2021), the patient complained of dyspnea and fatigue that had begun 2 weeks earlier. The patient's medical history included hypertension. However, the patient did not need to take medications on a regular basis to control it under the guidance of a cardiologist. Immunosuppressive therapy with rucotinib and dexamethasone and preventive fungal treatment with orally administered posaconazole were continued. There were no known drug allergies. The patient lived with her husband and one child in rural China. The patient had previously worked as a farmer and did not smoke or drink.
On examination, the patient's temperature was 36.6 C, her heart rate was 104 beats per minute, blood pressure was 94/68 mmHg, respiratory rate was 25 breaths per minute, and oxygen saturation was 93% while the patient breathed ambient air. Severe edema of the limbs was identified, while the results of the remaining examinations were normal. The copy number of cytomegalovirus (CMV) was 8,953 per milliliter (reference range: <1,000). Other laboratory test results are shown in  Table 2 . ECG revealed changes in the ST segment, and Echo revealed that the left ventricular ejection fraction (LVEF) had decreased to 49%. The level of serum cytokines was significantly increased ( Figure 2A ). Chest CT revealed interstitial inflammation of both lungs.
The patient received diuretic, albumin infusion, antiviral with ganciclovir, and supportive treatments. As part of CAR T-cell therapy, tocilizumab was administered to control elevated serum cytokine levels. Clinical symptoms were slightly improved within 1 week of admission. Notably, the level of cardiac troponin I (cTNI) was elevated prior to the administration of tocilizumab.
On the 12th day after presentation, PET-CT of the body and head was performed, which showed cardiac involvement and abnormal uptake in the liver ( Figures 1B, C ). Bone marrow aspirate and cytogenetic analysis were normal, while flow cytometry revealed lymphoblasts that accounted for 0.15% of the marrow cellularity. E2A/PBX1 rearrangement was also positive. The results of CAR T cells are shown in  Figure 2B . A cardiac mass biopsy was needed to achieve the pathological diagnosis but was not performed due to the patient's poor physical condition.
Owing to her poor physical condition, the patient was finally administered a reduced intensity chemotherapy with cyclophosphamide, vindesine, and prednisolone (VCP). Nevertheless, on the third day of chemotherapy, her blood pressure and oxygen saturation suddenly dropped to 61/29 mmHg and 76%, respectively. Along with a heart rate of 170 beats per minute, moist rales could be heard at the base of the left lung. ECG revealed tachycardia, ST-segment depression, and inverted T waves in the V2-6 leads. Although timely first-aid measures were provided, the patient could not be rescued and was eventually discharged from the hospital. The clinical course of the patient is shown in  Figure 3 .