Bcl-2 inhibition combined with PPARα activation synergistically targets leukemic stem cell-like cells in acute myeloid leukemia.

Persistence of leukemic stem cells (LSCs) is one of the determining factors to acute myeloid leukemia (AML) treatment failure and responsible for the poor prognosis of the disease. Hence, novel therapeutic strategies that target LSCs are crucial for treatment success. We investigated if targeting Bcl-2 and peroxisome proliferator activated receptor α (PPARα), two distinct cell survival regulating mechanisms could eliminate LSCs. This study demonstrate that the Bcl-2 inhibitor venetoclax combined with the PPARα agonist chiglitazar resulted in synergistic killing of LSC-like cell lines and CD34+ primary AML cells while sparing their normal counterparts. Furthermore, the combination regimen significantly suppressed AML progression in patient-derived xenograft (PDX) mouse models. Mechanistically, chiglitazar-mediated PPARα activation inhibited the transcriptional activity of the PIK3AP1 gene promoter and down-regulated the PI3K/Akt signaling pathway and anti-apoptotic Bcl-2 proteins, leading to cell proliferation inhibition and apoptosis induction, which was synergized with venetoclax. These findings suggest that combinatorial Bcl-2 inhibition and PPARα activation selectively eliminates AML cells in vivo and vitro, representing an effective therapy for patients with relapsed and refractory AML.

Ritanserin suppresses acute myeloid leukemia by inhibiting DGKα to downregulate phospholipase D and the Jak-Stat/MAPK pathway.

Refractory or relapsed (R/R) AML is the most challenging form of AML to treat. Due to frequent genetic mutations, therapy alternatives are limited. Here, we identified the role of ritanserin and its target DGKα in AML. Several AML cell lines and primary patient cells were treated with ritanserin and subjected to cell proliferation, apoptosis and gene analyses with CCK-8 assay, Annexin V/PI assay and Western blotting, respectively. We also evaluated the function of the ritanserin target diacylglycerol kinase alpha (DGKα) in AML by bioinformatics. In vitro experiments have revealed that ritanserin inhibits AML progression in a dose- and time-dependent manner, and it shows an anti-AML effect in xenograft mouse models. We further demonstrated that the expression of DGKα was elevated in AML and correlated with poor survival. Mechanistically, ritanserin negatively regulates SphK1 expression through PLD signaling, also inhibiting the Jak-Stat and MAPK signaling pathways via DGKα. These findings suggest that DGKα may be an available therapeutic target and provide effective preclinical evidence of ritanserin as a promising treatment for AML.

Therapeutic inhibition of PPARα-HIF1α-PGK1 signaling targets leukemia stem and progenitor cells in acute myeloid leukemia.

Treatment of acute myeloid leukemia (AML) with chemotherapeutic agents fails to eliminate leukemia stem cells (LSC)，and thus patients remain at high risk for relapse. Therefore, the identification of agents that target LSC is an important consideration for the development of new therapies. Enhanced glycolysis in LSC contributes to the aggressiveness of AML, which is difficult to be targeted. In this study, we showed that targeting peroxisome-proliferator-activated receptor α (PPARα), a ligand-activated transcription factor by chiglitazar provided a promising therapeutic approach. We first identified that chiglitazar reduced cell viability and proliferation of the leukemia stem-like cells population in AML. Treatment with chiglitazar blocked the ubiquitination of PPARα and increased its expression, resulting in the inhibition of glucose metabolism and apoptosis of AML cells. Consistent with its anti-leukemia stem-like cells activity in vitro, chiglitazar treatment in vivo resulted in the significant killing of leukemia stem-like cells as demonstrated in AML patient-derived xenograft (PDX) models. Mechanistically, PPARα overexpression inhibited the expression and promoter activity of PGK1 through blocking HIF1-α interaction on the PGK1 promoter. Thus, we concluded that targeting PPARα may serve as a novel approach for enhancing stem and progenitor cells elimination in AML.

Orelabrutinib and venetoclax synergistically induce cell death in double-hit lymphoma by interfering with the crosstalk between the PI3K/AKT and p38/MAPK signaling.

PURPOSE: Double-hit lymphoma (DHL) is a rare and aggressive mature B-cell malignancy with concurrent MYC and BCL2 rearrangements. When DHL becomes relapsed or refractory, it becomes resistant to the majority of therapeutic approaches and has subpar clinical results. Therefore, innovative therapeutics for this particular patient population are urgently needed. METHODS: Orelabrutinib, a new oral BTK inhibitor, combined with the Bcl-2 inhibitor venetoclax, was used to confirm the antitumor effect of DHL. Cell counting kit-8 and Annexin V-FITC/PI assays were used to examine the interaction of this combined regimen on DHL cell lines and primary lymphoma cells. RNA sequencing, EdU incorporation assay, mitochondrial membrane potential assay, and western blotting were employed to explore the molecule mechanism for the cytotoxicity of orelabrutinib with or without venetoclax against DHL cell lines. RESULTS: In this study, orelabrutinib combined with venetoclax synergistically induced DHL cell death, as evidenced by the inhibition of cell proliferation, the induct of cell cycle arrest, and the promotion of cell apoptosis via the mitochondrial pathway. Orelabrutinib treatment alters genome-wide gene expression in DHL cells. The combined regimen decreases the expression of BTK and Mcl-1, potentially interfering with the activity and crosstalk of PI3K/AKT signaling and p38/MAPK signaling. In addition, the combination of orelabrutinib and venetoclax shows cytotoxic activity in primary B-lymphoma cells. CONCLUSION: In summary, these findings reveal a novel therapy targeting BCR signaling and the Bcl-2 family for DHL patients with a poor prognosis.

Preclinical Studies of Chiauranib Show It Inhibits Transformed Follicular Lymphoma through the VEGFR2/ERK/STAT3 Signaling Pathway.

Transformed follicular lymphoma (t-FL), for which there is no efficient treatment strategy, has a rapid progression, treatment resistance, and poor prognosis, which are the main reasons for FL treatment failure. In this study, we identified a promising therapeutic approach with chiauranib, a novel orally developed multitarget inhibitor targeting VEGFR/Aurora B/CSF-1R. We first determined the cytotoxicity of chiauranib in t-FL cell lines through CCK-8, EdU staining, flow cytometry, and transwell assays. We also determined the killing effect of chiauranib in a xenograft model. More importantly, we identified the underlying mechanism of chiauranib in t-FL tumorigenesis by immunofluorescence and Western blotting. Treatment with chiauranib significantly inhibited cell growth and migration, promoted apoptosis, induced cell cycle arrest in G2/M phase, and resulted in significant killing in vivo. Mechanistically, chiauranib suppresses the phosphorylation level of VEGFR2, which has an anti-t-FL effect by inhibiting the downstream MEK/ERK/STAT3 signaling cascade. In conclusion, chiauranib may be a potential therapy to treat t-FL, since it inhibits tumor growth and migration and induces apoptosis by altering the VEGFR2/ERK/STAT3 signaling pathway.

PEGylated gold nanorods are not cytotoxic to human endothelial cells but affect kruppel-like factor signaling pathway.

Gold (Au) nanomaterials (NMs), particularly those with PEG surface functionalization, are generally considered to be biocompatible for biomedical applications due to relatively low cytotoxicity. Herein, we investigated the toxicity of PEGylated Au nanorods (NRs) to human umbilical vein endothelial cells (HUVECs), a commonly used in vitro model for human endothelium. We found a previously unknown effect that up to 10 µg/mL Au NRs, albeit not cytotoxic, decreased the mRNA and protein levels of kruppel-like factor 2 (KLF2), a transcription factor with well-documented vasoprotective effects. The results from PCR array showed that a number of genes associated with risk of cardiovascular diseases were altered by Au NRs, and several genes are downstream genes of KLF2 according to ingenuity pathway analysis (IPA). These effects could be observed with or without the presence of inflammatory stimuli lipopolysaccharide (LPS), which suggests a pre-existing inflammatory state is not required for Au NRs to alter KLF2 signaling pathway. We further identified that Au NRs significantly decreased eNOS mRNA/p-eNOS proteins as well as increased MCP-1 mRNA/sMCP-1 release, which are targets of KLF2. Combined, our data revealed a novel pathway that PEGylated Au NPs at non-cytotoxic concentrations might alter KLF leading to the increase of risk of cardiovascular diseases in human endothelial cells. Given the importance of KLF in vascular homeostasis, our data indicate that it is necessary to evaluate the influence of engineered NPs to KLF signaling pathways, especially for NPs with biomedical uses.