Gene-modified stem cell therapy for transfusion-dependent beta-thalassemia

Inclusion Criteria:

1. Must be willing to adhere to the protocol as evidenced by written informed consent for adults or parental informed consent and subject assent for adolescents and children.
2. Male and female adults/adolescents/children diagnosed with transfusion-dependent β-thalassemia (homozygous or compound heterozygous). At least 2 out of the 9 patients must have B0/B0 or B0/B0-like genotype. In case the genetic diagnosis available at screening wasn't performed in a certified laboratory (check under PI's or delegated investigator's responsibility), the genetic diagnosis will be repeated at clinical sites during the screening phase.
3. Documented history of at least 100 ml/kg/year or 10 U/year of packed red blood cell transfusions in each of the 2 years prior to signing informed consent.
4. Age ≥ 18 years and ≤ 35 years for Group 1, Age ≥ 3 years and ≤ 35 years for Group 2.
5. Karnofsky Index or Lansky ≥ 80%.
6. Adequate cardiac, renal, hepatic and pulmonary functions resulting in eligibility to undergo autologous HSCT as evidenced by:

   1. Left ventricular ejection fraction (LVEF) greater than 45% by echo and normal ECG or presence of abnormalities not significant for cardiac disease. Absence of severe pulmonary hypertension.
   2. Diffusing capacity of the lung for carbon monoxide (DLCO) \> 50% and forced expiratory volume in 1 sec (FEV1) and forced expiratory vital capacity (FVC) \> 60% predicted (if non cooperative: pulse oximetry \> 95 % in room air).
   3. Serum creatinine \< 2 x upper limit of normal and estimated GFR \> 60 ml/min/1.73m2, calculated using the CKD-EPI formula (Levey 2009) for adults and the modified Schwartz formula (Schwartz 2009) for pediatric patients.
   4. Absent-mild-moderate liver iron overload on T2\*MRI (i.e. LIC \< 15 mg Fe/gr dry weight assessed at screening. T2\*MRI at screening can be avoided if performed less than 6 months before enrolment at treatment centers).
   5. Absent-mild-moderate cardiac iron overload T2\*MRI (i.e. \> 20 msec assessed at screening. T2\*MRI at screening can be avoided if performed less than 6 months before enrolment at treatment centers).
   6. Absence of severe liver fibrosis or cirrhosis on Shear Wave (less than 6 months before enrolment) or of other advanced liver disorder.
7. For all patients in reproductive age, agreement to use highly effective and adequate method of contraception for at least 12 months following DP administration (including both females of childbearing potential and males with partners of childbearing potential).
8. Good adherence to transfusion and chelation programme, as indirect evidence of good adherence to treatment and follow-up evaluations for the current trial.
9. Availability of an adequate and well documented transfusion history (at least previous 24 months) or availability to follow a regular transfusion regimen according to guidelines and provide a detailed transfusion record of the 24 months prior to the DP administration.

Exclusion Criteria:

1. Use of other investigational agents within 4 weeks prior to study enrolment (within 6 weeks if use of long-acting agents).
2. Severe, active viral, bacterial or fungal infection at eligibility evaluation.
3. Current or prior malignant neoplasia (except local skin cancer or cervical intraepithelial neoplasia) or exceptional family history of familial cancer syndromes.
4. Current or prior immunodeficiency disorder.
5. History of uncontrolled seizures.
6. Aspartate transaminase (AST), alanine transaminase (ALT) \>3 × the upper limit of normal (ULN), or direct bilirubin value \>2.5 × ULN.
7. Baseline prothrombin time (International Normalized Ratio; INR) \>1.5 × ULN.
8. Other clinical conditions judged non compatible with the procedure and/or the treatment.
9. Positivity for HIV (serology or RNA), and/or HbsAg and/or HBV DNA and/or HCV RNA (patients who have completed antiviral treatment for HCV may only be enrolled if they have achieved a sustained virologic response, defined as undetectable HCV RNA at least 12 weeks after completion of therapy) and/or evidence of active infection of Treponema Pallidum or Mycoplasma species.
10. Positive history of significant previous thrombotic events. In case of a positive thrombophilic screening, patient's inclusion will be evaluated according to national guidelines.
11. Active alcohol or substance abuse within 6 months of the study.
12. Pregnancy or lactation.
13. Previous allogeneic hematopoietic stem cell transplantation.
14. Previous gene therapy treatment (gene addition or gene editing).
15. For patients until the age of 14 years only: availability of an HLA-matched family donor.
16. Patients with associated α-thalassemia and \>1 alpha deletion, or alpha multiplications.
17. Any other condition that would not allow the potential subject to complete follow-up examinations during the course of the study and, in the opinion of the investigator, makes the potential subject unsuitable for the study.