hox-loci-focused-crisprsgrna-library-screening-identifying-critical

<em>HOX</em> Loci Focused CRISPR/sgRNA Library Screening Identifying Critical CTCF Boundaries

CCCTC-binding factor (CTCF)-mediated stable topologically associating domains (TADs) play a critical role in constraining interactions of DNA elements ...

A Xenograft Mouse Model to Assess Efficacy of Therapeutic Agents for Human Acute Leukemia

Preclinical evaluation of therapeutic agents using an appropriate animal model is a critical step and a requirement for selecting drugs worth testing in ...

Pennsylvania State University College of Medicine

Bruton's tyrosine kinase (BTK) regulates many vital signaling pathways and plays a critical role in cell proliferation, survival, migration, and resistance. Previously, we reported that a small molecule, KS99, is an inhibitor of tubulin polymerization. In the present study, we explored whether KS99 is a dual inhibitor of BTK and tubulin polymerization. Although it is known that BTK is required for clonogenic growth and resistance, and microtubules are essential for cancer cell growth, dual targeting of these two components has not been explored previously. Through docking studies, we predicted that KS99 interacts directly with the catalytic domain of BTK and inhibits phosphorylation at the Y223 residue and kinase activities. Treatment of KS99 reduces the cell viability of multiple myeloma (MM) and CD138 cells, with an IC of between 0.5 and 1.0 μmol/L. We found that KS99 is able to induce apoptosis in MM cells in a caspase-dependent manner. KS99 suppressed the receptor activator of NF-κB ligand (RANKL)-induced differentiation of macrophages to osteoclasts in a dose-dependent manner and, importantly, inhibited the expression of cytokines associated with bone loss. Finally, we found that KS99 inhibits the in vivo tumor growth of MM cells through the inhibition of BTK and tubulin. Overall, our results show that dual inhibition of BTK and tubulin polymerization by KS99 is a viable option in MM treatment, particularly in the inhibition of refraction and relapse.

A novel dual inhibitor of microtubule and Bruton's tyrosine kinase inhibits survival of multiple myeloma and osteoclastogenesis.

Fast identification of BCR-ABL fusion genes is critical for precise diagnosis, risk stratification and therapy scheme selection in leukemia. More convenient methods are needed for quickly detection of the BCR-ABL fusion genes. Multiplex fluorescent reverse transcription quantitative real-time PCR (Multiplex RT-qPCR) methods are developed for detection of the at least 14 subtypes of BCR-ABL fusion genes in one tube at a time by using patients' bone marrow samples. The new Multiplex RT-qPCR method could quickly detect BCR-ABL fusion genes with sensitivity up to 10-10 copies. It can detect the fusion genes in patients' bone marrow samples containing any subtypes of the major bcr (M-bcr) e13a2, e14a2, e13a3 and e14a3, the minor bcr (m-bcr) e1a2 and e1a3, the micro bcr (μ-bcr) e19a2 and e19a3, and the nano bcr (n-bcr) e6a2 and e6a3. The specificity is comparable to the FISH methods. The cutoff for clinical diagnosis of BCR-ABL(+) is also determined by testing in clinical chronic myeloid leukemia samples. This is a new fast method with high sensitivity and specificity for clinical detection of BCR-ABL fusion genes. It will benefit the precise diagnosis, targeted therapy and minimal residual disease (MRD) monitoring in leukemia.

New rapid method to detect BCR-ABL fusion genes with multiplex RT-qPCR in one-tube at a time.

Mutations and single nucleotide polymorphisms of AT-rich interactive domain-containing protein 5B (ARID5B) are involved in the oncogenesis of acute lymphoblastic leukemia (ALL) and treatment outcomes. However, ARID5B expression and clinical significance in ALL remain unclear. We found ARID5B is significantly down-regulated in ALL compared to healthy bone marrow controls. ARID5B also interacts with PHD finger protein 2 (PHF2). Low expression of ARID5B (ARID5B) or ARID5B and PHF2 (ARID5BPHF2) is correlated with the markers of cell proliferation and poor prognosis in ALL patients. Ikaros directly regulates ARID5B expression in ALL. Restoring Ikaros function by Casein Kinase II inhibition also promotes ARID5B expression through recruitment of trimethylation of lysine 4 on histone H3 (H3K4me3) at its promoter region. In summary, our data show that aberrant expression of ARID5B and PHF2 is related to leukemic cell proliferation and several poor prognostic markers. Our data indicate ARID5B expression, particularly ARID5BPHF2 expression, is linked to Ikaros dysfunction and involved in the oncogenic effect of high-risk ALL, which may represent a high-risk subgroup of ALL.

Sinisa Dovat

Department of Pediatrics

Pennsylvania State University College of Medicine

HOX Loci Focused CRISPR/sgRNA Library Screening Identifying Critical CTCF Boundaries

A Xenograft Mouse Model to Assess Efficacy of Therapeutic Agents for Human Acute Leukemia