RXRA DT448/9PP generates a dominant active variant capable of inducing maturation in acute myeloid leukemia cells

Abstract

The retinoid receptors RARA and RXRA contribute to myeloid maturation in both mice and humans, and deletion of Rxra and Rxrb augments leukemic growth in mice. While defining the domains of RXRA that are required for anti-leukemic effects in mouse KMT2A-MLLT3 leukemia cells, we unexpectedly identified RXRA DT448/9PP as a constitutively active variant capable of inducing maturation and loss of their proliferative phenotype. RXRA DT448/9PP was associated with ligand-independent activity in reporter assays, with enhanced co-activator interactions, reduced engraftment in vivo, and activation of myeloid maturation transcriptional signatures that overlapped with cells treated with the potent RXRA agonist bexarotene, suggestive of constitutive activity that leads to leukemic maturation. Phenotypes of RXRA DT448/9PP appear to differ from two other RXRA mutations with forms of constitutive activity (F318A and S427F), in that DT448/9PP activity was resistant to mutations at critical ligand-interacting amino acids (R316A/L326A) and was resistant to pharmacologic antagonists, suggesting it may be ligand independent. These data provide further evidence that activated RXRs can regulate myeloid maturation and provide a novel constitutively active variant that may be germane for broader studies of RXR in other settings.

Modulating retinoid-X-receptor alpha (RXRA) expression sensitizes chronic myeloid leukemia cells to imatinib in vitro and reduces disease burden in vivo

“… Di Martino et al (2021a) showed that deleting endogenous RXRA in an MLL-AF9 AML mouse model increased leukemogenic potential and reduced survival. The same authors subsequently reported that a constitutively overexpressing mutant variant of RXRA resulted in myeloid maturation and prolonged survival in the murine model ( Di Martino et al, 2021a ; Di Martino et al, 2021b ). In line with this, in our RXRA overexpressing CDX CML mice model, we observed decreased leukemic burden and improved survival compared to EV-transduced CDX.…”

“…The dual activation of RXRA-RARA heterodimers by the specific ligands, ATRA and bexarotene effectively targeted MLL-AF9 myelomonocytic leukemic cells in-vitro and improved the survival in-vivo in mice models ( Di Martino et al, 2021a ). DT448/9PP mutated RXRA receptor overexpression enhanced transcriptional activity leading to differentiation and loss of colony-forming ability in KMT2A-MLLT3 transformed AML cells ( Di Martino et al, 2021b ). Bexarotene treatment increased the protein expression of RXR isoforms ( RXRA , RXRB & RXRG ) in SH-SY5Y cells ( Dheer et al, 2018 ).…”

An in vivo CRISPR screen in chick embryos reveals a role for MLLT3 in specification of neural cells from the caudal epiblast

“…Direct binding of MLLT3 to Retinoic Acid Receptors recruits the super elongation complex to rapidly induce neural programmes ( 80 ). Moreover, a mutant form of RXRα disrupts proliferation in murine KMT2A-MLLT3 leukemia cells ( 92 ). Consistent with a relationship between MLLT3 and RA signalling, our results from complementary approaches, indicate that disrupting the ability of RARα to interact with MLLT3 phenocopied the neural tube defects observed with perturbing MLLT3 .…”

An Isochroman Analog of CD3254 and Allyl-, Isochroman-Analogs of NEt-TMN Prove to Be More Potent Retinoid-X-Receptor (RXR) Selective Agonists Than Bexarotene

“…Luciferase detection. HEK293T cells were transfected using Lipofectamine 2000 (Invitrogen) [80]. Six hours after transfection, the cells were collected and plated into a 48-well plate in 1% BSA media in biological triplicates and treated with compounds.…”