Beta-thalassemias are hereditary blood disorders caused by reduced or absent synthesis of hemoglobin beta chains, with variable outcomes ranging from severe anemia to clinically asymptomatic individuals. Treatment is symptomatic and thalassemia is a major unmet medical need. Survival is increased, even in patients needing transfusions, in comparison with a few years ago, but the quality of life is poor for many patients. In some patients, an anomalous expression of gamma-globin genes has been observed, with a consequent rise in Fetal Hemoglobin levels. The patients displaying a clinical phenotype known as Hereditary Persistence of Fetal Hemoglobin (HPFH) exhibit a positive clinical status. To mimick HPFH, several compounds able to induce expression of fetal hemoglobins (HbF) have been evaluated. Within this framework, sirolimus is particularly interesting as an inducer of HbF. It has been used for many years for different indications and the available preclinical evidence warrant the start of a clinical development plan in thalassemia. The investigators propose a clinical trial in beta-thalassemia patients, designed to evaluate the effect of sirolimus on several parameters related to red blood cell status and to the level of HbF in particular, as a first step for the full clinical development in this new indication.

The general aim of the protocol is to demonstrate the applicability of a personalised and precision medicine approach in beta-thalassemia in a clinical trial setting for a repurposed drug, namely sirolimus. The presence of high level of Fetal Hemoglobin (HbF) is considered a condition predictive of a favourable outcome in thalassemia and its increase induced by pharmacological agents is considered a potential way to improve clinical status of the patients. In the present trial, in terms of efficacy analysis, the investigators will focus their attention on HbF levels.

Primary objective:

• To evaluate the suitability of sirolimus for the treatment of beta-thalassemia patients within the frame of a comprehensive project aimed to the reduction of their transfusions need (consequently ameliorating their quality of life). This goal can be obtained through a pharmacologically mediated increased level of HbF, with a prerequisite to be verified, namely the correlation between induction of HbF in vitro and in vivo in single patients.

Secondary objectives:

• To assess safety of sirolimus and correlation between administered dose and blood levels in beta-thalassemia patients,
• To assess the influence of sirolimus on transfusion regimen
• To assess the effect of sirolimus on hematopoietic and immune system of thalassemia patients.

• Patients over 18 years of age;
• Patient able to understand the informed consent and to sign it before any study procedure;
• With β+/β+ and β+/β0 thalassemia genotype;
• Documented diagnosis of major or intermediate thalassemia transfusion-dependent (nr of transfusion not less than 8 over the past 12 months before selection);
• On regular transfusion since at least 6 years;
• With splenectomy performed at least 60 days before selection or spleen dimensions < 20 cm in the largest part as detected by abdominal echography;
• Female participants who are surgically sterilised / hysterectomised or post-menopausal for longer than 2 years or female participants of childbearing potential using and/or willing to continue using a medically reliable method of contraception for the entire study duration, such as oral, injectable, or implantable contraceptives, or intrauterine contraceptive devices, or using any other method considered sufficiently reliable by the investigator in individual cases. Patients must be counseled concerning measures to be used to prevent pregnancy and potential toxicities prior to the first dose of sirolimus;
• Patient willing to follow all the study requirements and perform all the study visits and to cooperate with the investigator;
• Patient followed by the same clinical site since at least 6 months.

Note that patients will be treated with oral sirolimus only in the case their Erythroid Precursor Cells (ErPCs) are responsive to the in vitro treatment with sirolimus according to laboratory specific definition (≥ 20% increase of HbF in comparison with samples not treated with sirolimus);

Exclusion Criteria:

• Patient treated with hydroxyurea at selection visit or in the last 6 months;
• Ongoing treatment with drugs possibly affecting sirolimus actions;
• Documented aspartate aminotransferase (AST) or alanine aminotransferase (ALT) > 3x Upper Limit of Normal (ULN) at selection;
• Documented Platelet count <150.000/microliter and >1.000.000/microliter at selection;
• Heart failure as classified by the New York Heart Association (NYHA) classification 3 or higher;
• Uncontrolled hypertension defined as systolic blood pressure (BP) ≥ 140 mm Hg or diastolic BP ≥ 90 mm Hg;
• Significant arrhythmia requiring treatment,
• Corrected QT interval> 450 msec on selection ECG;
• Ejection fraction <50% by echocardiogram, multiple gated acquisition scan or cardiac magnetic resonance;
• Myocardial infarction within 6 months prior of selection;
• Positivity for human immunodeficiency virus (HIV) antibody, active hepatitis B (HBV) or hepatitis C (HCV) as demonstrated by the presence of hepatitis B surface antigen (HBsAg) and a positive HCV-RNA test, HBcAb and HBV-DNA positivity
• White blood cell [WBC] count <3000 cells per μL and/or Granulocytes <1500/mm3;
• Total cholesterol > 240 mg/dl;
• Triglycerides > 200 mg/dl;
• Proteinuria with urinary protein >1g/24 hrs;
• Current participation in another trial with investigational drug or experimental device, or inclusion in another trial with investigational drug or experimental device within the preceding month;
• Major surgery (including splenectomy) within 60 days before selection (patients must have fully recovered from any previous surgery);
• Iron chelation therapy changed in the last 3 months prior to selection (note that Deferiprone is not accepted as a chelation therapy drug in this study while Desferioxamine and Deferasirox are tolerated at stable dose);
• Current treatment with macrolide antibiotics (clarithromycin);
• Pregnant or lactating women;
• History of severe allergic or anaphylactic reactions or hypersensitivity to excipients in the experimental drug;
• Treatment with live vaccines within 90 days preceding the selection;
• Subject with history or current malignancies (solid tumors and haematological malignancies) or presence of masses/tumor detected by ultrasound at selection;
• Subject with any significant medical condition and/or laboratory abnormality considered by the investigator as not adequately controlled at the time of selection.

At the end of the study the study protocol and the clinical trial report will be available to other researchers. Publication of the data is planned

After completion of the Clinical Study Report preparation

Free availability of the publication. Free availability of the study protocol upon request