Meis1, which belongs to TALE-type class of homeobox gene family, appeared as one of the key regulators of hematopoietic stem cell (HSC) self-renewal and a potential therapeutical target. However, small molecule inhibitors of MEIS1 remained unknown. This led us to develop inhibitors of MEIS1 that could modulate HSC activity. To this end, we have established a library of relevant homeobox family inhibitors and developed a high-throughput in silico screening strategy against homeodomain of MEIS proteins using the AutoDock Vina and PaDEL-ADV platform. We have screened over a million druggable small molecules in silico and selected putative MEIS inhibitors (MEISi) with no predicted cytotoxicity or cardiotoxicity. This was followed by in vitro validation of putative MEIS inhibitors using MEIS dependent luciferase reporter assays and analysis in the ex vivo HSC assays. We have shown that small molecules named MEISi-1 and MEISi-2 significantly inhibit MEIS-luciferase reporters in vitro and induce murine (LSKCD34 l°w cells) and human (CD34 + , CD133 + , and ALDH hi cells) HSC self-renewal ex vivo. In addition, inhibition of MEIS proteins results in downregulation of Meis1 and MEIS1 target gene expression including Hif-1α, Hif-2α and HSC quiescence modulators. MEIS inhibitors are effective in vivo as evident by induced HSC content in the murine bone marrow and downregulation of expression of MEIS target genes. These studies warrant identification of first-in-class MEIS inhibitors as potential pharmaceuticals to be utilized in modulation of HSC activity and bone marrow transplantation studies. Meis1 is a member of TALE class of transcription factors 1. Through interaction domains in the N terminus, MEIS1 cooperates in transcription factors PBX1 and HOXA9 to transactivate target genes 2,3. MEIS2 and MEIS3 protein sequences demonstrate a high degree of similarity with MEIS1 4. Meis1 was first described in leukemia mouse model and identified as a viral integration site (reviewed in 5). MEIS proteins are characterized by PBX interaction domains and a highly conserved homeodomain (HD). MEIS1 HD shares identical MEIS2 HD amino acid sequence. Studies to understand how MEIS1 HD interacts with DNA led to crystallization of MEIS1 HD with target DNA and identification of DNA sequence preferentially bound by MEIS proteins as "TGACAG" 6-8. Meis1 is highly expressed in the bone marrow 2,9. Lethality occurs in Meis1 knockout mice at mid gestation (E14.5-15.5) with a number of hematopoietic, vascular and cardiac abnormalities 10-12. Conditional and tissue specific deletion of Meis1 in bone marrow led to loss of HSC quiescence associated with expansion of HSC pool in vivo 13. Meis1 has been shown to regulate HSC metabolism through transcriptional regulation of hypoxia factors including Hif1a and Hif2a 13-16. Deletion of Meis1 or Hif-1α in HSCs leads to reduction in the cytoplasmic glycolysis and induction of mitochondrial phosphorylation. Intriguingly, studies showed a fundamental role of Meis1 in neonatal cardiac regeneration. Increased Me...

Aberrations in the centrosome number and structure can readily be detected at all stages of tumor progression and are considered hallmarks of cancer. Centrosome anomalies are closely linked to chromosome instability and, therefore, are proposed to be one of the driving events of tumor formation and progression. This concept, first posited by Boveri over 100 years ago, has been an area of interest to cancer researchers. We have now begun to understand the processes by which these numerical and structural anomalies may lead to cancer, and vice-versa: how key events that occur during carcinogenesis could lead to amplification of centrosomes. Despite the proliferative advantages that having extra centrosomes may confer, their presence can also lead to loss of essential genetic material as a result of segregational errors and cancer cells must deal with these deadly consequences. Here, we review recent advances in the current literature describing the mechanisms by which cancer cells amplify their centrosomes and the methods they employ to tolerate the presence of these anomalies, focusing particularly on centrosomal clustering.

Adropin is a peptide hormone that has been implicated in insulin resistance and as a potential regulator of growth. The aim of this study is to determine the effect of calorie restriction on circulating levels of adropin in the MMTV-TGFα breast cancer mouse model and investigate the effects of adropin peptide on the viability of MCF-7 and MDA-231 breast cancer cells in culture. Ten-week-old mice were assigned to either ad libitum-fed (AL), chronic calorie-restricted, or intermittent calorie-restricted groups. Concentrations of serum adropin were measured using an enzyme-linked immunosorbent assay. Results showed an inverse correlation between serum adropin levels and mouse age that was attenuated by calorie restriction. In the AL group the level of adropin was significantly lower at week 50 compared to levels at week 10. However, among the calorie-restricted groups, serum levels of adropin remained high at week 50. The cell-line-specific effects were observed after treatment of cancer cell lines with a series of adropin concentrations (5, 10, 25, 50 ng/mL). Flow cytometry analysis showed that MCF-7 cells entered the early phase of apoptosis after treatment with 50 ng/mL for 24 h. Adropin may be involved in the protective effects that calorie restriction has on breast cancer risk.

Recent developments in gene editing technology have enabled scientists to modify DNA sequence by using engineered endonucleases. These gene editing tools are promising candidates for clinical applications, especially for treatment of inherited disorders like sickle cell disease (SCD). SCD is caused by a point mutation in human β -globin gene (HBB). Clinical strategies have demonstrated substantial success, however there is not any permanent cure for SCD available. CRISPR/Cas9 platform uses a single endonuclease and a single guide RNA (gRNA) to induce sequence-specific DNA double strand break (DSB). When this accompanies a repair template, it allows repairing the mutated gene. In this study, it was aimed to target HBB gene via CRISPR/Cas9 genome editing tool to introduce nucleotide alterations for efficient genome editing and correction of point mutations causing SCD in 2 human cell line, by Homology Directed Repair (HDR). We have achieved to induce target specific nucleotide changes on HBB gene in the locus of mutation causing SCD. The effect of on-target activity of bone fide standard gRNA and newly developed longer gRNA were examined. It is observed that longer gRNA has higher affinity to target DNA while having the same performance for targeting and Cas9 induced DSBs. HDR mechanism was triggered by co-delivery of donor DNA repair templates in circular plasmid form. In conclusion, we have suggested methodological pipeline for efficient targeting with higher affinity to target DNA and generating desired modifications on HBB gene. Graphical abstract Highlights• HBB gene were targeted by spCas9 in close proximity to the SCD mutation • Long gRNA, which is designed to target SCD mutation, is sickle cell disease specific and exhibits indistinguishable level of cleavage activity on target locus.• Functional HBB HDR repair templates with 1 Kb and 2 Kb size were generated to cover all known mutations in the HBB gene.• Replacement of PAM sequence in HDR template with HindIII recognition sequence allowed a quick assessment of the HDR efficiency.• HDR template: Cas9-GFP vector 2:1 ratio yielded the highest HDR events/GFP+ cells. 5