Cyclophosphamide Abrogates Immune Effector Cell-Associated Neurotoxicity Syndrome Associated With CAR-T Cell Therapy

Abstract

The patient was a 50-year-old woman diagnosed with acute lymphoblastic leukemia 6 years before the current presentation. After two cycles of VDCLP induction chemotherapy (vincristine, daunorubicin, cyclophosphamide, L-asparaginase, and prednisone), bone marrow examination indicated complete remission, with flow cytometry showing a minimal residual disease (MRD) level of < 1 × 10⁻⁴. The patient had undergone multiple consolidation treatments over the following 5 years. One year before the current presentation, a follow-up bone marrow flow cytometry revealed that abnormal immature lymphoblasts accounted for 24.4%, indicating leukemia relapse. After the failure of VDLP chemotherapy (vincristine, daunorubicin, L-asparaginase, and prednisone) and MA chemotherapy (methotrexate and cytarabine), and given the lack of a suitable donor for bone marrow transplantation, the patient enrolled in a chimeric antigen receptor T (CAR-T) clinical trial at our hospital (NCT06532630) in March 2024. This CAR-T cell therapy targets CD19 and is manufactured using a nonviral electroporation platform. It incorporates a scFv that directly binds to CD19, linked to a CD8α transmembrane domain, and integrates CD28 and CD3ζ signaling domains to activate and enhance T cell cytotoxicity. Given the patient's high initial tumor burden (49.5% blasts in the bone marrow), bridging chemotherapy with the VIP regimen (vincristine, idarubicin, and prednisone) was administered 20 days prior to CAR-T cell infusion to reduce tumor burden (Figure 1A).

FIGURE 1

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Timeline of the patient (A). Measures of cytokine levels related to cell infusion (B). Measure of CAR-T cell copies related to cell infusion, according to the study protocol, if the CAR-T cell copies fall below the limit of quantification for two consecutive tests, further testing is not required (C). T lymphocyte subset changes over time (D). Brain magnetic resonance imaging and CT scan. The images in the first row show the cranial CT scan results on Day 12 following CAR-T cell therapy, revealing mild cerebral edema. The images in the second row show the cranial MRI results on Day 32, indicating partial resolution of the edema. These images confirm the effectiveness of our treatment (E). Improvement of the patient since the use of cyclophosphamide. The first photo captures the condition on Day 12 after CAR-T cells infusion, where the patient had not yet received cyclophosphamide treatment, had been transferred to the ICU, was in a coma, and relied on mechanical ventilation support. The second photo was taken on Day 24 after the infusion, at which point the patient had received cyclophosphamide treatment, regained clear consciousness, and required only high-flow nasal cannula oxygen therapy. The third photo shows the situation on Day 37 post-infusion, where the patient was fully conscious, had returned to the hematology ward, no longer required any respiratory support, and was able to stand normally (F).

After lymphodepletion (fludarabine at a dose of 25 mg per square meter of body surface area daily, and cyclophosphamide at a dose of 250 mg per square meter of body surface area daily for 3 days) and infusion of 1 × 10⁶ CAR-T cells per kilogram of body weight, the patient developed a fever with a peak temperature of 40.6°C on Day 7 post-infusion. Considering the potential cytokine release syndrome (CRS), we administered acetaminophen and tocilizumab at a dose of 8 mg per kilogram of body weight for three times. Despite this, the patient's condition deteriorated further, with subsequent hypotension and tachycardia. Intravenous norepinephrine was promptly initiated to maintain blood pressure stability, metoprolol was given for heart rate control, while levetiracetam was used to prevent immune effector cell-associated neurotoxicity syndrome (ICANS). Nevertheless, on Day 11 post-infusion, the patient exhibited dysphasia. The condition subsequently progressed to seizures, with an ICE score of 0 (Grade 3 ICANS). The patient had a normal body temperature but low blood oxygen saturation at 85% (Grade 3 CRS). The patient was treated with phenobarbital sodium and diazepam to manage seizure symptoms, and methylprednisolone was utilized to address ICANS and pulmonary inflammation. Even with aggressive steroid treatment, the patient's ICANS symptoms failed to improve and further deteriorated. A CT scan of the head showed cerebral edema and inflammation of the paranasal sinuses. During this period, 10 mg of IV dexamethasone was administered every 6 h for 2 days from Day 9. As the patient's mental status continued to deteriorate, the steroid regimen was adjusted to 1 g of IV methylprednisolone daily for 3 days.

Given the patient's worsening mental status and hypoxemia, the patient was intubated on Day 12 and transferred to the intensive care unit (ICU). In order to abrogate CAR-T cell-associated neurotoxicity, cyclophosphamide was administered on Day 13 at a dosage of 1.5 g per square meter of body surface area. Shortly after the administration of cyclophosphamide, the cytokine levels decreased rapidly (Figure 1B). The copy number of CAR-T cells in the patient also decreased correspondingly (Figure 1C). The monitoring of T cell ratios also suggests that the patient's immune system is undergoing recovery (Figure 1D).

After cyclophosphamide administration and support treatments, the patient's mental status gradually improved. The patient was then weaned off the ventilator and extubated on Day 21. On Day 24, the patient was retransferred to the hematological ward. MRI indicated that cerebral edema had improved, with no significant intracranial abnormalities except for multiple areas of high signal intensity in the white matter (Figure 1E). Bone marrow examination indicated remission of the leukemia. The patient was then discharged home on Day 37 (Figure 1F).

Over the subsequent 8 months, the patient underwent monthly MRD assessments, all of which remained negative. The patient is still awaiting a suitable donor for bone marrow transplantation. At the last follow-up, the patient exhibited no late-onset neurotoxicity, and her leukemia remained in complete remission.

CAR-T cell therapy is a revolutionary approach for treating relapsed/refractory hematologic malignancies. CRS and ICANS are the two most common adverse events of CAR-T cell therapy. ICANS presents with neurotoxic symptoms that include confusion, delirium, seizures, headache, and aphasia. It is hypothesized that the activation of endothelial cells may lead to blood–brain barrier dysfunction, which in turn could initiate inflammation in the central nervous system and result in neurotoxicity [1]. Mild ICANS is typically managed with supportive care and antiseizure therapy; severe cases require corticosteroids to reduce inflammation and neurological symptoms. However, corticosteroids may not be effective in some severe ICANS cases. Some patients with severe ICANS deteriorate rapidly, experience malignant cerebral edema, and even succumb to this adverse event.

There is a lack of standardized treatment approaches and clear guidelines for managing steroid-resistant ICANS (Table S1). Research has identified a correlation between elevated serum IL-1 levels following CAR-T cell therapy and the development of severe ICANS [2]. Anakinra, an IL-1 receptor antagonist, has been employed in clinical studies to treat and prevent severe ICANS. In a study of 14 ICANS patients treated with anakinra, nine patients experienced symptom relief within 24 h after the last anakinra administration [3]. Nevertheless, anakinra exhibits a relatively low response rate and a longer onset time in the treatment of steroid-refractory ICANS, and may not be suitable for the emergency treatment of ICANS [4]. Siltuximab is a monoclonal antibody targeting IL-6, and it may alleviate the inflammatory response and neurotoxicity in ICANS patients by inhibiting the IL-6 signaling pathway. Current research on the efficacy of siltuximab in the treatment of ICANS remains limited, and further extensive studies are needed to confirm its therapeutic effects. The occurrence of ICANS has been associated with CAR-T cell doses that exceed the patient's maximum tolerated dose, particularly in relation to the tumor burden [1]. Foster et al. [5] have reported successful management of high-grade steroid-resistant ICANS using rimiducid as a molecular safety switch to deactivate CAR-T cells. Notwithstanding, the effectiveness of rimiducid depends on the presence of specific engineered receptors in CAR-T cells, requiring specific genetic modifications for rimiducid responsiveness. Introducing rimiducid into clinical practice may increase treatment complexity and costs. Besides, recent studies advocate the use of intrathecal corticosteroids along with cytotoxic drugs for the treatment of steroid-resistant ICANS. Even so, some ICANS patients simultaneously developed severe thrombocytopenia and coagulation disorders, rendering them unable to tolerate intrathecal chemotherapy. Graham et al. [6] reported a case of successful treatment using cyclophosphamide for steroid-refractory ICANS following BCMA CAR-T cell therapy. In their case, the patient exhibited a slow progression of symptoms. The most severe grade of ICANS that was observed reached only Grade 2. This indicates that the patient experienced relatively mild neurotoxic effects, as Grade 2 is associated with moderate symptoms that are manageable and less severe compared to higher grades. In contrast, among patients receiving CD19-targeted CAR-T cell therapy, ICANS develops and progresses more rapidly, with more severe symptoms that can even be fatal. After all, there are currently no universally effective treatments for fatal ICANS in clinical practice.

In this case, glucocorticoids failed to effectively halt the progression of ICANS. The patient also experienced severe CRS, leading us to promptly administer cyclophosphamide to terminate CAR-T cell therapy. The rationale for choosing cyclophosphamide lies in its extensive use in clinical practice, its easy accessibility for physicians, and their familiarity with its application. Additionally, cyclophosphamide can rapidly eliminate CAR-T cells and reduce cytokine levels, effectively controlling neurotoxicity and preventing fatal cerebral edema. Once severe cerebral edema develops, the damage is often irreversible. This timely intervention with comprehensive supportive care in the ICU successfully saved the patient's life. We also observed that although cyclophosphamide eliminated most CAR-T cells, it did not completely suppress their antileukemia activity. Furthermore, over time, the number of CAR-T cells in the patient's body increased once more. Under supportive care in the ICU, the patient quickly overcame the increased risk of infection and myelosuppression caused by chemotherapy, and no new complications were observed.

To our knowledge, this is the first case of successfully treating fatal ICANS induced by CD19 CAR-T cell therapy using cyclophosphamide. It is straightforward to administer, economical, and highly effective. Further studies are needed to evaluate the impact of recurrent lymphocyte depletion on CAR-T cell therapy efficacy and the cytotoxic effects of cyclophosphamide on patients.