DJ4 Targets the Rho-Associated Protein Kinase Pathway and Attenuates Disease Progression in Preclinical Murine Models of Acute Myeloid Leukemia

Abstract

The poor prognosis of acute myeloid leukemia (AML) and the highly heterogenous nature of the disease motivates targeted gene therapeutic investigations. Rho-associated protein kinases (ROCKs) are crucial for various actin cytoskeletal changes, which have established malignant consequences in various cancers, yet are still not being successfully utilized clinically towards cancer treatment. This work establishes the therapeutic activity of ROCK inhibitor (5Z)-2–5-(1H-pyrrolo[2,3-b]pyridine-3-ylmethylene)-1,3-thiazol-4(5H)-one (DJ4) in both in vitro and in vivo preclinical models of AML to highlight the potential of this class of inhibitors. Herein, DJ4 induced cytotoxic and proapoptotic effects in a dose-dependent manner in human AML cell lines (IC50: 0.05–1.68 μM) and primary patient cells (IC50: 0.264–13.43 μM); however, normal hematopoietic cells were largely spared. ROCK inhibition by DJ4 disrupts the phosphorylation of downstream targets, myosin light chain (MLC2) and myosin-binding subunit of MLC phosphatase (MYPT), yielding a potent yet selective treatment response at micromolar concentrations, from 0.02 to 1 μM. Murine models injected with luciferase-expressing leukemia cell lines subcutaneously or intravenously and treated with DJ4 exhibited an increase in overall survival and reduction in disease progression relative to the vehicle-treated control mice. Overall, DJ4 is a promising candidate to utilize in future investigations to advance the current AML therapy.

Characterization of Anticancer Effects of the Analogs of DJ4, a Novel Selective Inhibitor of ROCK and MRCK Kinases

“…In our previously reported studies, we observed that DJ4 treatment exhibited minor effects on proliferation of A549 and MDA-MB-231 cells after 24 h of exposure [ 11 ]. Interestingly, we observed much more profound effects on cell viability in acute myeloid leukemia (AML) cell lines at 24 h [ 12 ]. To determine whether the effects on cell viability in AML cells were related to the specific cancer subtypes, or to differences in temporal response to DJ4, we examined whether DJ4 affected the viability of various NSCLC cells over longer periods of drug exposure.…”

Preclinical evaluation of the ROCK1 inhibitor, GSK269962A, in acute myeloid leukemia

“…Recently, ROCK1 was identified as a crucial gene for leukemic bulk and stem/progenitor cells using large-scale RNAi screens in primary human AML cells ( Wermke et al, 2015 ). DJ4, a novel ROCK inhibitor, was demonstrated to suppress ROCK/MYPT1/MLC2 pathway and result in cell death of AML cells ( Golla et al, 2021 ). As these findings suggested that ROCK1 could be a potential therapeutic target for AML, evaluating the anti-leukemia effect of ROCK1 inhibitor in preclinical models might be of potential value.…”

Rho-associated kinase1 promotes Laryngeal squamous cell carcinoma tumorigenesis and progression via the FAK pathway

“…Rho-associated kinase1 (ROCK1) is a classic serine/threonine protein kinase. ROCK1 can regulate downstream substrates such as muscle Globulin, adducin and protein kinases [3]. ROCK1 also can phosphorylate various substrates such as myosin light chain (MLC) phosphatase, LIM kinase, PTEN, insulin receptor substrate (IRS), ezrin/radixin/moesin (ERM) proteins, and JNK interacting protein (JIP-3) [4][5][6].…”