The beta-thalassemias are serious blood diseases, caused by diverse molecular mutations affecting the beta-chain of adult hemoglobin A, (a2b2), and are designated as a global health burden. The conditions have early mortality and no approved definitive therapy. Fetal globin (HbF, a2g2) is the major modulator of disease severity, and increasing its synthesis is established to reduce globin imbalance and ameliorate these conditions. A challenge to successful application of HbF inducers in the beta-thalassemias has been the genetic heterogeneity, which produces variable baseline fetal globin levels (and as yet unpredictable) responsiveness to HbF-inducing therapeutics between individuals. This proposal combines two distinct investigative approaches: 1) a long-term investigation to identify high potency, molecularly-based therapeutics to induce HbF, and 2) expertise in identifying thalassemia genotypes, modifiers, and mechanisms which modulate severity. First, two drug discovery screening campaigns, using molecular modeling or high throughput screening (HTS), have identified a select panel of novel, structurally-unrelated, high-potency HbF-inducing therapeutics, including some drugs already FDA-approved for other indications. Second, a 5-year genome wide screening (GWAS) study in international patient populations identified 3 influential polymorphisms, or quantitative trait loci, (QTL), associated with elevated HbF, less clinical severity, and mechanism of action. We will apply these 2 experiences to determine 1) which novel therapeutics most effectively augment HbF in erythroid cells cultured from beta thalassemia patients, genotyped for 2 major subtypes of thalassemia, or patients with sickle cell disease, and the 3 influential genetic modifier (QTL) profiles; 2) molecular mechanisms of action of the new therapeutic agents, and 3) combinations of the targeted therapeutics, which are most potent in erythroid cells cultured from different subsets +/- epigenetic inducing agents. This work will provide a basis for evaluating the optimal therapeutics in patients most likely to respond favorably in clinical trials. Our Aims include: Aim I: To determine which novel therapeutics enhance gamma globin expression most effectively in erythroid cells cultured from patients genotyped for beta thalassemia mutations and the 3 influential QTLs. Aim II: To determine molecular effects of the novel therapeutics on select known components of repressors of the fetal globin gene promoter and if the three HbF QTLs correlate with more or less robust responses to select inducing agents. Determine if new therapeutics suppress components of established repressor complexes (BCL-11A, Sox 6, HDAC-3). Determine whether differences in QTL profiles correlate with responses or lack thereof to different gamma globin inducing therapeutics. Aim III: To determine if a combination of an epigenetic-acting HbF inducer and a promoter-targeted inducer will produce more potent HbF induction than single agents alone, in the beta thalassemia subsets and in HbSS erythroid cells with low baseline HbF.