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Professor Yonghong ZHANG: Autologous Chimeric Antigen Receptor T Cell (CAR-T) Therapy Successfully Treated One Case of Refractory Burkitt's Lymphoma

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Editor's note: Burkitt's lymphoma (BL) is a highly invasive B-cell lymphoma that is highly prevalent in children, and it is the most common pathological type of non-Hodgkin's lymphoma in children. High-intensity and short-course chemotherapy regimen has achieved good efficacy in the treatment of BL, but some patients still have no response to the treatment. In recent years, autologous chimeric antigen receptor T cell (CAR-T) therapy has successfully treated refractory/relapsed leukemia, and the data of several clinical trials have confirmed that CAR-T cells can treat various subtypes of B-cell lymphomas, including follicular lymphoma, mantle cell lymphoma and marginal zone lymphoma, but no report of CAR-T therapy for Burkitt's lymphoma has been seen before. Beijing Boren Hospital has accumulated rich clinical experience in CAR-T therapy for leukemia. Professor Yonghong ZHANG from Beijing Children's Hospital affiliated to Capital Medical University and Beijing Boren Hospital was invited by our journal to share a case of refractory Burkitt's lymphoma successfully treated with CAR-T therapy.

Overview of Medical History

Case summary: The child, Xiao **, male, 9 years old, was admitted to our hospital due to Burkitt's lymphoma for more than 8 months (stage III, CNS1) and progression for 3 months after chemotherapy.

I. Diagnosis and Treatment

At 9 months before admission to our hospital, the child was admitted to a local hospital due to “pale complexion with fatigue”, and abdominal mass was found. Blood routine test: WBC 8.4×10^9/L, N 57%, RBC 3.65×10^12/L, HB 61 g/L, PLT 672×10^9/L. Abdominal CT: the thickness of the small intestine wall in the right abdomen was significantly increased, up to about 2.6 cm, showing a mass shadow, and the local intestinal lumen was unobstructed; several small nodules were seen in the mesentery and retroperitoneum, about 0.3-0.5 cm. PET-CT: the wall of small intestine and ascending colon in the right abdomen was significantly thickened, with abnormally increased metabolism; the wall of descending colon was slightly thickened, with increased metabolism; the lymph nodes beside the right cardiac border, above the diaphragm and in the mesenteric space were slightly enlarged, with increased metabolism, and the high metabolism at the above sites was considered as lymphoma infiltration; the metabolism of small lymph nodes in the groins was not increased. No obvious abnormalities were found in the whole body by PET/CT imaging. No tumor cells were seen in the bone marrow morphology, iron was reduced internally and externally, considered as iron-deficiency anemia in combination with clinical examination, the flow type, fusion genes and chromosomes were not abnormal, and the cerebrospinal fluid was normal in routine test, biochemistry test, morphology and flow type. The child underwent small intestine tumor resection + intestinal resection + intestinal anastomosis in the local hospital, and postoperative pathology showed: (intestinal tumor + greater omentum) highly malignant lymphoma, tending to Burkitt's lymphoma, with diffuse large B-cell lymphoma (non-special type) GCB type to be excluded. Immunohistochemistry: CD20 (+), CD3 (-), CD43 (partially +), CD79a (+), Bcl-2 (-), CD5 (-), CD10 (+), Bcl-6 (+), Ki-67 (+, >95%), CK-pan (-), MUM-1 (+), Pax-5 (+), CD21 (DC+), c-Myc (+80%), Cyclin D1 (-), CD30 (-), ALK (-), CD 23 (-), P53 (+90%), TdT (-), CD38 (+). EBER (hybridized) (-), C-MYC gene detection indicated that the MYC/IGH fusion genes accounted for 93%. No FISH test was performed.

Eight months before admission, 6 courses of chemotherapy were given using the regimen recommended for the Chinese pediatric B-cell lymphoma (AA+BB+CC+AA+BB+CC). After 6 courses of chemotherapy, evaluation was performed before drug withdrawal. The whole abdomen enhanced CT found nodules in the left front of the abdominal aorta and masses in the left lower abdomen and the pelvic cavity, considered as metastatic tumors, and further PET/CT also found multiple lymphoma infiltrations in the abdominal cavity. After the second-line R-ICE chemotherapy for 2 courses, the PET/CT showed that the treatment was not effective and the abdominal tumors were enlarged. Subsequently, the treatment with ifosfamide + rituximab was continued, and local radiotherapy was given for 10 times, but the dose was unknown. The tumors did not disappear, and the child was transferred to Beijing Boren Hospital after the second tumor biopsy. The biopsy pathology was still diagnosed as Burkitt's lymphoma after consultation with the Department of Pathology in Beijing Children's Hospital and the Peking University Third Hospital. In the local hospital, chemotherapy was given for 9 courses, local radiotherapy for 10 times, and rituximab for 5 times.

II. Diagnosis of Relapsed Foci after Admission

Pathological diagnosis in Beijing Children's Hospital (biopsy of relapsed foci): (abdominal wall) non-Hodgkin's lymphoma changed after chemotherapy, consistent with Burkitt's lymphoma in WHO classification; most of the tumors were necrotic, most of the remaining tumor cells were squeezed and deformed, only a small number of round and elliptical tumor cells infiltrated among connective tissues and fat; immunohistochemical results: CD20 (+), CD19 (locally weak +), CD22 (+), Ki-69 (90%), Bcl-2 (-), Bcl-6 (+), P53 (+), TdT (-), CD3 (-), PD-1 (-), CD10 (+), MUN1 (-), PD-L1 (-); in-situ hybridization result: EBER (-).

Pathological consultation on relapsed foci in Peking University Third Hospital: in combination with morphology and immunohistochemistry, (abdominal wall) punctured tissue: Burkitt's lymphoma. Obvious tissue necrosis was seen in morphology, tumor cells were degenerated, but not typical BL change, and this diagnosis was supported in combination with medical history and protein level markers. Tumor cells were highly proliferated, diffusely and strongly expressing CD20. CD19 and CD22 also well expressed well, but were slightly weaker than CD20, and the cells were slightly reduced. FISH test suggested that C-myc was ruptured and positive, supporting the diagnosis of Burkitt's lymphoma. The results of pathology and FISH test of the relapsed foci are shown in Figure 1.

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Figure 1: Pathology And FISH Test of The Relapsed Foci

III. Tumor Evaluation After Admission

Abdominal CT: a mass was seen at the anterior wall of the lower abdomen, about 8.5X8.3X4.0 cm in size. Bone marrow morphology and flow type were normal. The cerebrospinal fluid was normal in routine test, biochemistry test, morphology and flow type. Head MRI did not suggest obvious abnormalities. In combination with pathology, FISH and tumor evaluation results, it was diagnosed as Burkitt's lymphoma (stage III, CNS-1).

Process of CAR-T Therapy

The child still had tumor progression after high-intensity chemotherapy and radiotherapy, suggesting drug resistance and poor prognosis. We communicated with the family members on the conditions, patients who progressed in similar treatments were unlikely to achieve complete remission of tumors after chemotherapy alone, and the remission rate was less than 5%, as reported in foreign literatures. The family members voluntarily entered the “Phase I Clinical Study on Sequential CART Cell Therapy for Children and Young Adults with Refractory/Relapsed Mature B-cell Lymphoma (Registration No. ChiCTR18000144)” and signed the informed consent form.

The peripheral blood lymphocytes of the child were collected to culture second-generation murine CD19-CART cells that provide signals for costimulatory molecule 4-1BB. On September 25, 2018, cyclophosphamide and fludarabine were used for pretreatment for 3 days. Murine CD19 CAR-T cells 7.43x10^5/Kg were then back infused (d0). The patient was then monitored for CART cell count, cytokines and vital signs.

I. CRS

At the night of October 1, 2018 (d1), the child developed low-grade fever (37.7°C), with no fear of cold and chills, mild dry cough; with mild fatigue; the peak temperature gradually increased (the trend of change is shown in Figure 2). On October 1, 2018 (d11 ), the temperature peaked at 40°C, with low levels of three indicators, white blood cell count (0.09-1.55)×10^9/L, neutrophil count (0.02-1.23)×10^9/L, lowest on d0, highest on d5, hemoglobin (62-98) g/L, lowest on d3, platelet count (13-107)×10^9/L, lowest on d3, and C-reactive protein (0-182.16) mg/L, highest on d9. With no shortness of breath, wheezing, cyanosis and decreased blood oxygen; no vomiting, diarrhea, abdominal distension and abdominal pain; no bleeding tendency, no tremor, convulsions, etc. (The change in hemogram within 2 weeks of CAR-T cell therapy is shown in Figure 8; CRS and use of hormones are shown in Table 1).

Highest body temperature within 14 days after back infusion

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Daily highest Body Temperature

Figure 2: Trend in Body Temperature Peak After Back Infusion of CAR-T Cells

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Table 1: CRS, grading and use of hormones after back infusion of CAR-T cells

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The levels of cytokines IL-2, IL-6, IFN-γ, TNF-α, soluble CD25 and CRP peaked at least once in 15 days, and the level of CAR-T cells peaked on d15. With clinical observation of gradual decrease of peak temperature to normal, the cytokines gradually decreased to normal ranges, and CAR-T cells fell below the detectable level. (Trend of changes in cytokines and CRP after back infusion of CAR-T cells is shown in Figure 3, and the trend of change in the level of CAR-T cells is shown in Figure 4).

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Cytokine

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Figure 3: Trend of Changes in Cytokines And CRP After Back Infusion of CAR-T cells

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Figure 4: Trend of Change in The Level of CAR-T Cells

II. Tumor changes

Abdominal B-ultrasound found that the volume of tumor was gradually reduced. B-ultrasound showed that the size of tumor was 7.2*2.3*6.3 cm3 before pretreatment, 5.3*1.2*3.8 cm3 1 day before back infusion and respectively 1.1*1.3*1.2 cm3 and 1.1*1.1*1.0 cm3 on d30 and d44 after back infusion, with no significant change (Trend of change in tumor size by abdominal B-ultrasound monitoring is shown in Figure 5). On d37 after back infusion of CAR-T cells, PET-CT showed complete remission of tumor and no obvious hypermetabolic lesions in the whole body (Comparison of PET-CT before and after CAR-T therapy is shown in Figure 6).

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Left: Before CAR-T therapy (August 2, 2018)

Right: After CAR-T therapy (November 6, 2018, d37 after CAR-T therapy)

Figure 6: Comparison of PET-CT Before And After CAR-T Therapy

III. Follow-up After Discharge

The child was discharged on d50 after back infusion of CAR-T cells and followed up at clinic. By January 6, 2019 (d98 after back infusion of CAR-T cells), CAR-T cell count was maintained at a low level or below the detectable level, the general condition was good, the vital signs were stable, abdominal B-ultrasound found no significant change in the mass, hemogram showed that white blood cell count and neutrophil count did not return to normal levels, and no infection and special comorbidities were found in 3 months of follow-up.

Changes in Immune Cells After CAR-T Therapy

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Table 2: Comparison of Humoral Immunity Before And After CAR-T Therapy

Humoral immunity:

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Discussion

Burkitt's lymphoma (BL) is a highly invasive B-cell lymphoma that is highly prevalent in children, and it is the most common pathological type of non-Hodgkin's lymphoma in children. High-intensity and short-course chemotherapy was given to treat BL and achieved good efficacy, especially use of rituximab achieved a 5-year event-free survival rate up to 80% in high-risk patients. However, 10-15% of patients with Burkitt's lymphoma still had no response to chemotherapy. Once the intensity of chemotherapy was weak or the chemotherapy was delayed, the tumor cells would rapidly develop drug resistance, and the patients would have no response to chemotherapy again, be difficult to achieve remission again and lose the possibility of cure. It has been internationally reported that the survival rate of patients who progressed during treatment and recently relapsed is less than 5%.

This patient was diagnosed with Burkitt's lymphoma and did not achieve remission after treatment with conventional high-intensity chemotherapy combined with rituximab, so it was a refractory tumor. The international treatment strategy for refractory/relapsed Burkitt's lymphoma is recommended as the second-line chemotherapy regimen with R-ICE as the first choice and hematopoietic stem cell transplantation after remission, but this patient also did not respond to the second-line R-ICE regimen and even die not respond to large-dose radiotherapy that is rarely used in children, suggesting that it was difficult to achieve remission, and the survival rate of tumor-bearing transplantation was still zero, so the treatment could only be abandoned in similar cases.

In recent years, autologous chimeric antigen receptor T cell (CAR-T) therapy has successfully treated refractory/relapsed leukemia, then it was reported that CAR-T therapy successfully treated adult diffuse large B-cell lymphoma in 2017, and the data of several clinical trials have confirmed that CAR-T cells can treat various subtypes of B-cell lymphomas, including follicular lymphoma, mantle cell lymphoma and marginal zone lymphoma. However, no report of CAR-T therapy for Burkitt's lymphoma has been seen before. In this case, autologous chimeric antigen receptor CD19 CAR-T therapy had significant efficacy on refractory Burkitt's lymphoma and achieved complete remission after one back infusion of cells, indicating that CAR-T therapy has good efficacy on refractory Burkitt's lymphoma. The child well tolerated CAR-T cell therapy in the treatment. Although he developed grade I CRS such as fever, he could get through smoothly after symptomatic treatment (see Figure 8 for change in hemogram within 2 weeks after back infusion of CAR-T cells), indicating that after close observation and symptomatic treatment, the side effects were controllable and the therapy was safe and effective.

Trend of Change in White Blood Cell Count

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Trend of Change in Neutrophil Count

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Trend of Change in Hemoglobin

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Trend of Change in Hemoglobin

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Figure 8: Change in Hemogram Within 2 Weeks After Back Infusion of CAR-T Cells

Although this patient has achieved remission, does he need bridging transplantation? There is no conclusion whether sustained complete remission can be achieved without transplantation. In theory, mature B-cell lymphoma may be cured if it achieves CR, and whether relapse may occur if no transplantation is given needs further observation and follow-up. In addition, there is no report on whether children may develop long-term side effects such as immunodeficiency, and it is still necessary to closely observe the changes in tumors and various indicators and to follow up for a longer period of time.

CAR-T cell therapy for lymphoma is a new and rapidly developing treatment regimen with very good prospects, and it is becoming a new and salvage treatment for patients with refractory/relapsed B-cell lymphoma and will bring good news to patients who cannot be cured by conventional chemotherapy.