Biology subjects

Haileselassie, B.

Publications and source records attributed to Haileselassie, B..

3 recordsLinked to original sources

bioRxiv2026

Dual targeting of PDPK1 and BRAF V600E is synthetically lethal

PDPK1 functions downstream of PI3K and is essential for activating AKT and other AGC kinases. Although PDPK1 has a central role in the PI3K/AKT/mTOR signaling pathway, there has been limited evaluation of it as a target for cancer therapy. Anaplastic thyroid cancer (ATC) has one of the highest mortality rates of all human malignancies. Although combined BRAF and MEK inhibition in BRAF V600E mutant ATC (present in 45% of cases) results in response, resistance is common, and there is no curative treatment for ATC. The majority (up to 95.8%) of ATC cases have activation in the PI3K/AKT/mTOR and RAS/RAF/MEK/MAPK pathways due to genetic alterations involved in these pathways. In this study, we investigated PDPK1 as a therapeutic target for ATC. We used in vitro, ex vivo, and in vivo ATC models to evaluate the effect of targeting PDPK1 (BX795) alone and in combination with BRAF V600E inhibition (dabrafenib), and the mechanism of action that resulted in ATC cell death. BX795 monotherapy significantly reduced ATC cell proliferation, invasion, colony formation, and spheroid size. Combination BX795 and dabrafenib treatment had strong synergistic anticancer activity in BRAF V600E-mutant ATC models and led to simultaneous and sustained suppression of PDPK1/AKT and MAPK signaling, preventing the compensatory pathway reactivation observed with single-agent treatment. Mechanistically, combined inhibition induced pronounced oxidative stress, DNA damage, and G2-phase cell-cycle arrest, accompanied by mitochondrial dysfunction and robust activation of apoptosis in ATC cells. These effects resulted in marked tumor regression in in vitro, ex vivo, and in vivo ATC models. Our findings identify PDPK1 as a critical therapeutic vulnerability in ATC. Co-targeting PDPK1 and BRAF V600E produces potent synergistic anticancer activity by shutting down convergent oncogenic signaling pathways and amplifying apoptotic stress responses. These data support PDPK1 inhibition alone and in combination with BRAF blockade as a promising therapeutic strategy in BRAF V600E-mutant cancers.

cancer biology↗

bioRxiv2023

Extracellular release of damaged mitochondria induced by cytotoxic conditioning exacerbates GVHD

Despite therapeutic advancements, GVHD is a major complication of HSCT. In current models of GVHD, tissue injury induced by cytotoxic conditioning regimens, along with translocation of microbes expressing Pathogen Associated Molecular Patterns (PAMPs), result in activation of host antigen-presenting cells (APC) to stimulate alloreactive donor T lymphocytes. Recent studies have demonstrated that in many pathologic states, tissue injury results in the release of mitochondria from the cytoplasm to the extracellular space. We hypothesized that extracellular mitochondria, which are related to archaebacteria, could also trigger GVHD by stimulation of host APC. We found that clinically relevant doses of radiation or busulfan induced extracellular release of mitochondria by various cell types, including cultured intestinal epithelial cells. Conditioning-mediated mitochondrial release was associated with mitochondrial damage and impaired quality control but did not affect the viability of the cells. Extracellular mitochondria directly stimulated host APCs to express higher levels of MHC-II, co-stimulatory CD86, and pro-inflammatory cytokines, resulting in increased donor T cell activation, and proliferation in mixed lymphocyte reactions. Analyses of plasma from both experimental mice and a cohort of children undergoing HSCT demonstrated that conditioning induced extracellular mitochondrial release in vivo. In mice undergoing MHC mismatched HSCT, administration of purified syngeneic extracellular mitochondria increased host APC activation and exacerbated GVHD. Our data suggests that pre-HSCT conditioning results in extracellular release of damaged mitochondria which increase alloreactivity and exacerbate GVHD. Therefore, decreasing the extracellular release of damaged mitochondria following conditioning could serve as a novel strategy for GVHD prevention.

immunology↗

bioRxiv2023

Cardiac Troponin I Directly Binds and Inhibits Mitochondrial ATP Synthase: a Noncanonical Role in the Post-Ischemic Heart

Cardiac troponin I (cTnI) is a sarcomeric protein critical to myocyte contraction. Unexpectedly, we found that some cTnI localized to the mitochondrial matrix in the heart, inhibited mitochondrial functions when stably expressed in non-cardiac cells and increased opening of the mitochondrial permeability transition pore under oxidative stress. Direct, specific, and saturable binding of cTnI to ATP synthase was demonstrated in vitro, using immune-captured ATP synthase, and in cells using proximity ligation assay. cTnI binding doubled F1F0 ATPase activity, whereas skeletal troponin I and several human mutant cTnI variants associated with familial hypertrophic cardiomyopathy did not. A rationally-designed ten amino acid peptide, P888, inhibited cTnI binding to ATP synthase, inhibited cTnI-induced increase in ATPase activity in vitro, and reduced cardiac injury following transient ischemia in vivo. We therefore suggest that mitochondria-associated cTnI may inhibit cardiac ATP synthase under basal conditions; pharmacological agents that release this inactivating effect of cTnI and thus preventing ATP hydrolysis during cardiac ischemia may increase the reservoir of functional mitochondria to reduce cardiac injury. Significance StatementCardiac troponin I (cTnI) is a key sarcomeric protein involved in the regulation of myocardial contractility. We found that some cTnI is present in the mitochondrial matrix where it binds to ATP synthase, disrupting mitochondrial function; inhibition of the cTnI-ATP synthase interaction with a selective peptide inhibitor reduces cardiac dysfunction following ischemia and reperfusion injury. Several pathogenic cTnI mutations associated with hypertrophic cardiomyopathy do not affect ATP synthase activity, suggesting a potential mechanism that contributes to the diverse pathologies associated with these mutations.

cell biology↗