Rational identification of a Cdc42 inhibitor presents a new regimen for long-term hematopoietic stem cell mobilization

Abstract

Mobilization of hematopoietic stem cells (HSCs) from bone marrow (BM) to peripheral blood (PB) by cytokine granulocyte colony-stimulating factor (G-CSF) or the chemical antagonist of CXCR4, AMD3100, is important in the treatment of blood diseases. Due to clinical conditions of each application, there is a need for continued improvement of HSC mobilization regimens. Previous studies have shown that genetic ablation of the Rho GTPase Cdc42 in HSCs results in their mobilization without affecting survival. Here we rationally identified a Cdc42 activity-specific inhibitor (CASIN) that can bind to Cdc42 with submicromolar affinity and competitively interfere with guanine nucleotide exchange activity. CASIN inhibits intracellular Cdc42 activity specifically and transiently to induce murine hematopoietic stem/progenitor cell egress from the BM by suppressing actin polymerization, adhesion, and directional migration of stem/progenitor cells, conferring Cdc42 knockout phenotypes. We further show that, although, CASIN administration to mice mobilizes similar number of phenotypic HSCs as AMD3100, it produces HSCs with better long-term reconstitution potential than that by AMD3100. Our work validates a specific small molecule inhibitor for Cdc42, and demonstrates that signaling molecules downstream of cytokines and chemokines, such as Cdc42, constitute a useful target for long-term stem cell mobilization.

Acute Myeloid Leukemia: Aging and Epigenetics

“…In addition, the authors reported an altered distribution of a H3K9me2 heterochromatin mark in aged HSCs. Interestingly, treatment with CASIN (a Cdc42-activity inhibitor) [71,92,93] restored Lamin A/C levels, H3K9me2 peripheral localization, and nuclear volume in aged HSCs to the levels observed in young HSCs. Alterations of another heterochromatin mark, H3K9me3, have been reported in aged HSCs by Djeghloul et al [62].…”

Targeting Rac and Cdc42 GEFs in Metastatic Cancer

“…A derivative of Pirl1, CASIN (Cdc42 activity-specific inhibitor), demonstrated in vitro and in vivo efficacy in colorectal cancer models with an IC 50 of 2 µM (Sakamori et al, 2014). CASIN could also prove to be useful in improving hematopoietic stem cell mobilization and benefiting bone marrow transplant recipients (Liu et al, 2019). ZCL278 is another ITSN inhibitor, identified through virtual screening to directly bind Cdc42 by interacting with several residues of its Switch I region, essential for the interaction with ITSN.…”

The Small GTPase Cdc42 Is a Major Regulator of Neutrophil Effector Functions

“…To study the influence of Cdc42 on various neutrophil effector mechanisms, the Cdc42 activity-specific inhibitor (casin) was used. Casin is a Pir1 analog, and confers the most active Cdc42 binding activity without binding to related RhoGTPases (7)(8)(9). Casin was shown to specifically bind to Cdc42, and to competitively interfere with its guanine nucleotide exchange (GEF) activity, therefore, suppressing Cdc42-GTP formation (8).…”