Relapsed/refractory acute myeloid leukemia (R/R AML) currently lacks effective CAR-T therapeutic agents due to the absence of tumor-specific target antigens. Most AML-associated antigens are expressed on normal hematopoietic stem/progenitor cells (HSPCs) and healthy tissues, increasing the risk of on-target off-tumor toxicity and non-neoplastic toxicity. CD33 is present on leukemic cells in over 80% of AML patients. Compared with CLL-1, CD123 and other targets, CD33 exhibits higher expression across diverse AML subtypes, reducing the risk of treatment failure and relapse caused by antigen escape and thus serving as an ideal therapeutic target for AML. However, conventional CD33-targeted CAR-T cells demonstrate suboptimal efficacy in clinical trials, accompanied by significant toxicity and inadequate in vivo expansion. To further investigate the safety and efficacy of CAR-T therapy for AML, our center has initiated a clinical trial of functionally optimized CD33 CAR-T (FO33 CAR-T) cells for R/R AML. We constructed a lentiviral CAR vector containing the CD33-targeting scFv, 4-1BB, and CD3ζ, followed by insertion of adjuvant molecule X. FO33 CAR-T cells showed superior cytotoxicity against AML cell lines and enhanced biological activity compared with conventional CD33 CAR-T cells, and exerted safe and effective antitumor effects in preclinical models. This single-center, open-label, prospective clinical trial aims to evaluate the safety and efficacy of FO33 CAR-T cells in patients with R/R AML, as well as to characterize the pharmacokinetic and pharmacodynamic (PK/PD) profiles of this therapy.

Inclusion Criteria:

• Subjects diagnosed with refractory/recurrent acute myeloid leukemia (excluding M3) who meet any of the following criteria:

  1. Relapse: Recurrence of leukemia cells in peripheral blood or ≥5% blast cells in bone marrow after complete remission (CR) of AML (excluding other causes such as bone marrow regeneration following consolidation chemotherapy), or extramedullary leukemia infiltration.
  2. Refractory: First-time cases unresponsive to two cycles of standard therapy; relapse within 12 months after consolidation therapy following CR; relapse after 12 months without response to conventional chemotherapy; two or more relapses; persistent extramedullary leukemia.

• During enrollment screening, bone marrow flow cytometry must demonstrate a CD33+ expression rate of ≥80% in leukemia cells and/or pathological immunohistochemical confirmation of CD33+ extramedullary lesions.

• Estimated survival duration exceeding 3 months as of the date of informed consent signing.

• Participants with Eastern Cooperative Oncology Group (ECOG) physical status scores ranging from 0 to 2.

• Age range of 14 years ≤ ≤ 75 years, inclusive, with no gender restriction.

• Hemoglobin (HGB) level ≥70 g/L with transfusion capability.

• Liver/kidney function and cardiopulmonary function meeting the following criteria:

  1. Creatinine ≤1.5×ULN;
  2. Left ventricular ejection fraction ≥50%;
  3. Blood oxygen saturation>90%;
  4. Total bilirubin ≤1.5×ULN; ALT and AST ≤2.5×ULN.

• Acceptance of autologous CART cells with peripheral blood tumor burden ≤ 30%;

• The subject or guardian understands and signs the informed consent form.

Exclusion Criteria:

• Presence of one of the following cardiac criteria:

  1. Atrial fibrillation;
  2. Myocardial infarction (MI) within the past 12 months;
  3. Prolonged QT syndrome or secondary QT prolongation as determined by the investigator;
  4. Echocardiographic left ventricular systolic fraction (LVSF) <30% or left ventricular ejection fraction (LVEF) <50%;
  5. Clinically significant pericardial effusion; New York Heart Association (NYHA) class III or IV heart failure (confirmed by echocardiography within 12 months after treatment).

• Active graft-versus-host disease (GVHD).

• History of severe pulmonary dysfunction.

• Concurrent other progressive malignancies.

• Concurrent severe or persistent infections that cannot be effectively controlled.

• Concurrent severe autoimmune diseases or congenital immunodeficiency.

• Active hepatitis (HBV-DNA ≥ 500 IU/ml with abnormal liver function or HCV antibody [HCV-Ab] positivity, HCV-RNA exceeding the detection limit of analytical methods with abnormal liver function).

• Human immunodeficiency virus (HIV) infection or syphilis infection.

• History of severe allergic reactions to biological products (including antibiotics).

• Presence of central nervous system disorders such as uncontrolled epilepsy, cerebrovascular ischemia/hemorrhage, dementia, or cerebellar diseases.

• Female patients in pregnancy or lactation, or planning pregnancy within 12 months.

• Situations where investigators consider may increase subject risk or interfere with trial outcomes.