Biology subjects

Jha, B. K.

Publications and source records attributed to Jha, B. K..

4 recordsLinked to original sources

bioRxiv2026

FLT3-ITD signals for CEBPA and p53 proteolysis by the ubiquitin-proteosome pathway

Internal-tandem-duplication of the receptor tyrosine kinase FLT3 (FLT3-ITD) generates ligand-independent signaling and is highly recurrent in acute myeloid leukemias (AMLs). One way signaling pathways can quickly influence cell fates is by phosphorylating key fate-determining proteins to trigger their proteolysis. We investigated the master transcription factor (MTF) driver of granulo-monocytic lineage-fates, CEBPA, for regulation by this mechanism because we found high CEBPA mRNA but little CEBPA protein in FLT3-ITD versus FLT3-wildtype AML cells, and inhibiting FLT3-ITD signaling with tyrosine kinase inhibitors (TKI) rapidly rescued CEBPA protein. Mass spectrometry analyses of CEBPA and its interactome demonstrated prominent interactions with major ubiquitin-proteosome pathway (UPP) components UHRF1 and USP7. TKI treatments decreased CEBPA and USP7 phosphorylations at serine 21 and serine 18 respectively alongside shifts in CEBPA interactions from degradative ubiquitin-ligase UHRF1 toward protective deubiquitinase USP7. The rescued CEBPA activated granulocytic-differentiation. Supporting that the serine-phosphorylations were phospho-degrons, UPP-inhibitors (bortezomib, MG132) increased phosphorylated and total CEBPA and USP7. The MTF regulator of apoptosis p53 is a known USP7 client, therefore, we also evaluated p53 status: TKIs and UPP-inhibitors stabilized USP7 and p53, triggering apoptosis in addition to granulocytic-differentiation specifically in FLT3-ITD but not FLT3-wildtype AML cells. UPP-inhibitors produced these consequences in TKI-resistant FLT3-ITD AML cells also. These data predicted genetic loss-of-function to CEBPA or TP53 is redundant in the FLT3-ITD context, borne out by mutual exclusivity of the mutations in clinical series. In summary, FLT3-ITD signals for CEBPA and p53 proteolysis to block lineage-maturation and apoptosis, positioning UPP-inhibitors as therapeutic candidates acting downstream of TKIs. KEY POINTSO_LIThe oncoprotein kinase FLT3-ITD signals for CEBPA and p53 proteolysis and hence suppresses lineage-differentiation and apoptosis C_LIO_LIProteosome-inhibitors are candidate remedies to restore CEBPA and p53, acting downstream of presently used FLT3-ITD kinase inhibitors C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=171 SRC="FIGDIR/small/738455v1_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@f3a2acorg.highwire.dtl.DTLVardef@13cd700org.highwire.dtl.DTLVardef@1475312org.highwire.dtl.DTLVardef@19efbd1_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

MCL1 may not mediate chemoresistance

The anti-apoptotic BCL2 family member MCL1 is overexpressed in many cancers and has been linked to chemoresistance. Unlike other BCL2 family members, MCL1 displays both well-defined mitochondrial anti-apoptotic activities and also emerging nuclear functions. Prior reports suggest that MCL1 enters the nucleus during chemotherapy and promotes chemoresistance by influencing cell cycle progression and DNA repair. These nuclear roles of MCL1, however, remain poorly characterized. Using a newly validated monoclonal antibody across several cell lines and treatments, we find no evidence that MCL1 enhances chemoresistance or preferentially accumulates in the nucleus after drug exposure. Proximity biotinylation identified novel nuclear MCL1 interactors but did not recover previously reported DNA repair or cell cycle partners. Thus, while MCL1 does reach the nucleus and interact with nuclear proteins, our data do not support a role for MCL1 in chemoresistance. Further work is needed to clarify the functional significance of nuclear MCL1. SIGNFIGANCEPrevious studies have implicated MCL1 in promoting chemoresistance via interactions with DNA repair machinery in the nucleus. Using a validated, monoclonal anti-MCL1 antibody, we were unable to replicate these data. We report that MCL1 neither confers chemoresistance, translocates to the nucleus during chemotherapy treatment, nor interacts with DNA repair proteins in live cancer cells.

cell biology↗

bioRxiv2020

Resistance to decitabine and 5-azacytidine emerges from adaptive responses of the pyrimidine metabolism network

Mechanisms-of-resistance to decitabine and 5-azacytidine, mainstay treatments for myeloid malignancies, require investigation and countermeasures. Both are nucleoside analog pro-drugs processed by pyrimidine metabolism into a nucleotide analog that depletes the key epigenetic regulator DNA methyltranseferase 1 (DNMT1). We report here that DNMT1 protein, although substantially depleted (~50%) in patients bone marrows at response, rebounded at relapse, and explaining this, we found pyrimidine metabolism gene expression shifts averse to the processing of each pro-drug. The same metabolic shifts observed clinically were rapidly recapitulated in leukemia cells exposed to the pro-drugs in vitro. Pyrimidine metabolism is a network that senses and preserves nucleotide balances: Decitabine, a deoxycytidine analog, and 5-azacytidine, a cytidine analog, caused acute and distinct nucleotide imbalances, by off-target inhibition of thymidylate synthase and ribonucleotide reductase respectively. Resulting expression changes in key pyrimidine metabolism enzymes peaked 72-96 hours later. Continuous pro-drug exposure stabilized metabolic shifts generated acutely, preventing DNMT1-depletion and permitting exponential leukemia out-growth as soon as day 40. Although dampening to activity of the pro-drug initially applied, adaptive metabolic responses primed for activity of the other. Hence, in xenotransplant models of chemorefractory AML, alternating decitabine with 5-azacytidine, timed to exploit compensating metabolic shifts, and addition of an inhibitor of a catabolic enzyme induced by decitabine/5-azacytidine, extended DNMT1-depletion and time-to-distress by several months versus either pro-drug alone. In sum, resistance to decitabine and 5-azacytidine emerges from adaptive responses of the pyrimidine metabolism network; these responses can be anticipated and thus exploited. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=85 SRC="FIGDIR/small/958405v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@e3f77aorg.highwire.dtl.DTLVardef@8caaa4org.highwire.dtl.DTLVardef@117236dorg.highwire.dtl.DTLVardef@16ee98_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2018

Development of a Cx46 targeting strategy for cancer stem cells

Gap junction-mediated cell-cell communication enables tumor cells to synchronize the execution of complex processes. Despite the connexin family of gap junction proteins being considered tumor suppressors, we previously found that glioblastoma cancer stem cells (CSCs) express higher levels of Cx46 compared to non-stem tumor cells, and this was necessary and sufficient for CSC maintenance. To develop a Cx46 targeting strategy, we utilized point mutants to disrupt specific functions of Cx46 and found that gap junction coupling was the critical function of Cx46 for CSCs. Based on this finding, we screened a clinically relevant library of small molecules and identified clofazimine as an inhibitor of Cx46-specific cell-cell communication. Clofazimine attenuated proliferation, self-renewal, and tumor growth and synergized with temozolomide to induce apoptosis. These data suggest that combining clofazimine with standard-of-care therapies may target glioblastoma CSCs. Furthermore, these results demonstrate the importance of targeting cell-cell communication as an anti-cancer therapy.

cancer biology↗