Treatment of Erythroid Precursor Cells from β-Thalassemia Patients with Cinchona Alkaloids: Induction of Fetal Hemoglobin Production

Zuccato, Cristina; Cosenza, Lucia Carmela; Zurlo, Matteo; Lampronti, Ilaria; Borgatti, Monica; Scapoli, Chiara; Gambari, Roberto

doi:10.3390/ijms222413433

Cited by 20 publications

(24 citation statements)

References 63 publications

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“…Data of RT-qPCR experiments were analyzed using CFX Manager™ software (Bio-Rad). The relative expression of globins’ mRNAs was calculated using the comparative cycle threshold method (ΔΔCt method) using as reference genes human GAPDH sequences [ 24 , 58 , 61 ].…”

Section: Methodsmentioning

confidence: 99%

“…Analysis of HbA, HbF and free α-globin chains was performed with HPLC as elsewhere reported [ 24 , 58 , 61 , 63 ]. Lysates have been loaded into a PolyCAT-A cation exchange column and then eluted in a sodium-chloride-BisTris-KCN aqueous mobile phase using HPLC Beckman Coulter instrument System Gold 126 Solvent Module-166 detector, which allows to obtain for the quantification of the hemoglobins present in the sample.…”

Section: Methodsmentioning

confidence: 99%

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Co-Treatment of Erythroid Cells from β-Thalassemia Patients with CRISPR-Cas9-Based β039-Globin Gene Editing and Induction of Fetal Hemoglobin

Cosenza

Zuccato

Zurlo

et al. 2022

Genes

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Gene editing (GE) is an efficient strategy for correcting genetic mutations in monogenic hereditary diseases, including β-thalassemia. We have elsewhere reported that CRISPR-Cas9-based gene editing can be employed for the efficient correction of the β039-thalassemia mutation. On the other hand, robust evidence demonstrates that the increased production of fetal hemoglobin (HbF) can be beneficial for patients with β-thalassemia. The aim of our study was to verify whether the de novo production of adult hemoglobin (HbA) using CRISPR-Cas9 gene editing can be combined with HbF induction protocols. The gene editing of the β039-globin mutation was obtained using a CRISPR-Cas9-based experimental strategy; the correction of the gene sequence and the transcription of the corrected gene were analyzed by allele-specific droplet digital PCR and RT-qPCR, respectively; the relative content of HbA and HbF was studied by high-performance liquid chromatography (HPLC) and Western blotting. For HbF induction, the repurposed drug rapamycin was used. The data obtained conclusively demonstrate that the maximal production of HbA and HbF is obtained in GE-corrected, rapamycin-induced erythroid progenitors isolated from β039-thalassemia patients. In conclusion, GE and HbF induction might be used in combination in order to achieve the de novo production of HbA together with an increase in induced HbF.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Co-Treatment of Erythroid Cells from β-Thalassemia Patients with CRISPR-Cas9-Based β039-Globin Gene Editing and Induction of Fetal Hemoglobin

Cosenza

Zuccato

Zurlo

et al. 2022

Genes

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“…Interestingly, our group is further exploring the possibility of repositioning old drugs as new drugs for thalassemia, as shown by the recent study who identified Cinchona alkaloids (cinchonidine, quinidine and cinchonine) as natural HbF-inducing agents in human erythroid cells 60 . Two highly active compounds, cinchonidine and quinidine, were able to induce γ-globin mRNA and HbF in erythroid progenitor cells isolated from β-thalassemia patients.…”

Section: Drug Repositioning For Rare Diseases: β-Thalassemiamentioning

confidence: 99%

A Rational Approach to Drug Repositioning in β-thalassemia: Induction of Fetal Hemoglobin by Established Drugs

et al. 2022

Self Cite

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Drug repositioning and the relevance of orphan drug designation for β-thalassemia is reviewed. Drug repositioning and similar terms ('drug repurposing', 'drug reprofiling', 'drug redirecting', ‘drug rescue’, ‘drug re-tasking’ and/or 'drug rediscovery') have gained great attention, especially in the field or rare diseases (RDs), and represent relevant novel drug development strategies to be considered together with the “off-label” use of pharmaceutical products under clinical trial regimen. The most significant advantage of drug repositioning over traditional drug development is that the repositioned drug has already passed a significant number of short- and long-term toxicity tests, as well as it has already undergone pharmacokinetic and pharmacodynamic (PK/PD) studies. The established safety of repositioned drugs is known to significantly reduce the probability of project failure. Furthermore, development of repurposed drugs can shorten much of the time needed to bring a drug to market. Finally, patent filing of repurposed drugs is expected to catch the attention of pharmaceutical industries interested in the development of therapeutic protocols for RDs. Repurposed molecules that could be proposed as potential drugs for β-thalassemia, will be reported, with some of the most solid examples, including sirolimus (rapamycin) that recently has been tested in a pilot clinical trial.

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“…Importantly, the utilization of natural HbF inducers in countries with a high prevalence rate of disease is more affordable as compared to other ways of treatment. Some reported HbF inducers from natural sources are resveratrol, mithramycin, rapamycin, curcuminoid, cucurbitacin D, cinchonidine, and quinidine ( Liu et al, 2010 ; Ng and Ko, 2014 ; Iftikhar et al, 2019 ; Iftikhar et al, 2020 ; Zuccato et al, 2021 ). Yet, many of these are recently identified HbF inducing agents, and their therapeutic applications are restricted due to their carcinogenic and cytotoxic properties and require further clinical trials ( de Dreuzy et al, 2016 ).…”

Section: Introductionmentioning

confidence: 99%

In Vitro and In Vivo Studies for the Investigation of γ-Globin Gene Induction by Adhatoda vasica: A Pre-Clinical Study of HbF Inducers for β-Thalassemia

Iftikhar¹,

Rahman

Khan³

et al. 2022

Front. Pharmacol.

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Fetal hemoglobin (HbF) is a potent genetic modifier, and the γ-globin gene induction has proven to be a sustainable therapeutic approach for the management of β-thalassemia. In this study, we have evaluated the HbF induction ability of A. vasica in vitro and in vivo, and the identification of potential therapeutic compounds through a bioassay-guided approach. In vitro benzidine-Hb assay demonstrated strong erythroid differentiation of K562 cells by A. vasica extracts. Subsequently, an in vivo study with an aqueous extract of A. vasica (100 mg/kg) showed significant induction of the γ-globin gene and HbF production. While in the acute study, the hematological and biochemical indices were found to be unaltered at the lower dose of A. vasica. Following the bioassay-guided approach, two isolated compounds, vasicinol (1) and vasicine (2) strongly enhanced HbF levels and showed prominent cellular growth kinetics with ample accumulation of total hemoglobin in K562 cultures. High HbF levels were examined by immunofluorescence and flow cytometry analysis, concomitant with the overexpression in the γ-globin gene level. Compound 1 (0.1 µM) and compound 2 (1 µM) resulted in a greater increase in F-cells (90 and 83%) with marked up (8-fold and 5.1-fold) expression of the γ-globin gene, respectively. Molecular docking studies indicated strong binding affinities of (1) and (2) with HDAC2 and KDM1 protein that predict the possible mechanism of compounds in inhibition of these epigenetic regulators in the γ-globin gene reactivation. Altogether, these observations demonstrated the therapeutic usefulness of A. vasica for fostering HbF production in clinical implications for blood disorders.

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Treatment of Erythroid Precursor Cells from β-Thalassemia Patients with Cinchona Alkaloids: Induction of Fetal Hemoglobin Production

Cited by 20 publications

References 63 publications

Co-Treatment of Erythroid Cells from β-Thalassemia Patients with CRISPR-Cas9-Based β039-Globin Gene Editing and Induction of Fetal Hemoglobin

Co-Treatment of Erythroid Cells from β-Thalassemia Patients with CRISPR-Cas9-Based β039-Globin Gene Editing and Induction of Fetal Hemoglobin

A Rational Approach to Drug Repositioning in β-thalassemia: Induction of Fetal Hemoglobin by Established Drugs

In Vitro and In Vivo Studies for the Investigation of γ-Globin Gene Induction by Adhatoda vasica: A Pre-Clinical Study of HbF Inducers for β-Thalassemia

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