Identification of a Potent Antagonist of Smoothened in Hedgehog Signaling

Abstract Background Hedgehog signaling is essential to the regulation of embryonic development, tissue homeostasis, and stem cell self-renewal, making it a prime target for developing cancer therapeutics. Given the close link between aberrant Hedgehog signaling and cancers, many small molecular compounds have been developed to inhibit this pathway for treating cancer and several of such compounds have been approved by FDA (GDC-0449 and LDE-225). As we known, acquired drug resistance is the main obstacle of first generation of Smoothened inhibitors. Therefore, new Smoothened inhibitors that could inhibit wild-type and mutant receptors are essential for refractory cancers. ResultsWe established the Smoothened β arrestin 2 GFP high throughput screening platform based on the mechanistic discovery of Hedgehog signaling pathway, and discovered several active small molecules targeting Smoothened including 0025A 0025A. Here we show that 0025A can block the translocation of β arrestin 2 GFP to Smoothened , displace Bodipycyclopamine binding to wild type Smoothened or mutant Smoothened D473H and reduce the expression of Gli upon Hedgehog stimulation. In addition, we reveal that 0025A can effectively suppress hair follicle morphogenesis and hair growth in mice. Conclusions Thus, our results demonstrate that 0025A is a potent antagonist targeting Smoothened wild-type and mutant type and mutant receptors in the Hreceptors signaling pathway and may provide a new new therapy for refractory cancers.

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Identification of a potent antagonist of smoothened in hedgehog signaling

Cell & Bioscience ◽

10.1186/s13578-021-00558-9 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Junwan Fan ◽

Haowen Li ◽

Lun Kuang ◽

Zichen Zhao ◽

Wenyan He ◽

...

Keyword(s):

Signaling Pathway ◽

High Throughput Screening ◽

Hedgehog Signaling ◽

Cancer Therapeutics ◽

The United States ◽

Hedgehog Signaling Pathway ◽

Wild Type ◽

Close Link ◽

Acquired Drug Resistance ◽

Potent Antagonist

AbstractBackgroundHedgehog signaling is essential to the regulation of embryonic development, tissue homeostasis, and stem cell self-renewal, making it a prime target for developing cancer therapeutics. Given the close link between aberrant Hedgehog signaling and cancers, many small molecular compounds have been developed to inhibit Smoothened, a key signal transducer of this pathway, for treating cancer and several such compounds have been approved by the United States Food and Drug Administration (GDC-0449 and LDE-225). However, acquired drug resistance has emerged as an important obstacle to the effective use of these first generation Hedgehog pathway blockers. Thus, new Smoothened inhibitors that can overcome such resistance is an urgent need going forward.ResultsWe established the Smoothened/βarrestin2-GFP high-throughput screening platform based on the mechanistic discovery of Hedgehog signaling pathway, and discovered several active small molecules targeting Smoothened including 0025A. Here we show that 0025A can block the translocation of βarrestin2-GFP to Smoothened, displace Bodipy-cyclopamine binding to wild-type Smoothened or mutant Smoothened-D473H, reduce the accumulation of Smo on primary cilia and the expression of Gli upon Hedgehog stimulation. In addition, we show that 0025A can effectively suppress hair follicle morphogenesis and hair growth in mice.ConclusionsOur results demonstrate that 0025A is a potent antagonist targeting Smoothened wild-type and mutant receptors in the Hedgehog signaling pathway and may provide a new therapy for refractory cancers.

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Cooperative Blockade of CK2 and ATM Kinases Drives Apoptosis in VHL-Deficient Renal Carcinoma Cells through ROS Overproduction

Cancers ◽

10.3390/cancers13030576 ◽

2021 ◽

Vol 13 (3) ◽

pp. 576

Author(s):

Sofia Giacosa ◽

Catherine Pillet ◽

Irinka Séraudie ◽

Laurent Guyon ◽

Yann Wallez ◽

...

Keyword(s):

High Throughput Screening ◽

Renal Carcinoma ◽

Kinase Inhibitors ◽

Ex Vivo ◽

Cancer Therapeutics ◽

Carcinoma Cells ◽

Wild Type ◽

Cell Renal Cell Carcinoma ◽

Von Hippel Lindau ◽

Renal Carcinoma Cells

Kinase-targeted agents demonstrate antitumor activity in advanced metastatic clear cell renal cell carcinoma (ccRCC), which remains largely incurable. Integration of genomic approaches through small-molecules and genetically based high-throughput screening holds the promise of improved discovery of candidate targets for cancer therapy. The 786-O cell line represents a model for most ccRCC that have a loss of functional pVHL (von Hippel-Lindau). A multiplexed assay was used to study the cellular fitness of a panel of engineered ccRCC isogenic 786-O VHL− cell lines in response to a collection of targeted cancer therapeutics including kinase inhibitors, allowing the interrogation of over 2880 drug–gene pairs. Among diverse patterns of drug sensitivities, investigation of the mechanistic effect of one selected drug combination on tumor spheroids and ex vivo renal tumor slice cultures showed that VHL-defective ccRCC cells were more vulnerable to the combined inhibition of the CK2 and ATM kinases than wild-type VHL cells. Importantly, we found that HIF-2α acts as a key mediator that potentiates the response to combined CK2/ATM inhibition by triggering ROS-dependent apoptosis. Importantly, our findings reveal a selective killing of VHL-deficient renal carcinoma cells and provide a rationale for a mechanism-based use of combined CK2/ATM inhibitors for improved patient care in metastatic VHL-ccRCC.

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Discovery of novel drug candidates based on herbaric acid derivates as potential inhibitors of the hedgehog signaling pathway in cervical cancer therapeutics

Journal of Physics Conference Series ◽

10.1088/1742-6596/1442/1/012052 ◽

2020 ◽

Vol 1442 ◽

pp. 012052

Author(s):

M A F Nasution ◽

A A Parikesit ◽

U S F Tambunan

Keyword(s):

Cervical Cancer ◽

Signaling Pathway ◽

Hedgehog Signaling ◽

Cancer Therapeutics ◽

Hedgehog Signaling Pathway ◽

Drug Candidates ◽

Potential Inhibitors ◽

Novel Drug

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Investigation of mTOR, JAK/STAT, and Hedgehog pathways inhibitor effect on the proliferation of haematological cancer cell lines

Biologija ◽

10.6001/biologija.v67i2.4456 ◽

2021 ◽

Vol 67 (2) ◽

Author(s):

Roberta Vadeikienė ◽

Aistė Savukaitytė ◽

Danguolė Laukaitienė ◽

Rasa Ugenskienė ◽

Elona Juozaitytė

Keyword(s):

Signaling Pathway ◽

Cell Lines ◽

Hedgehog Signaling ◽

Myeloproliferative Neoplasms ◽

Wild Type ◽

Trypan Blue Exclusion ◽

Signaling Systems ◽

Ic50 Values ◽

Signaling Inhibitors

Constitutively activated JAK/STAT signaling pathway is a common feature of the BCR/ABL-negative classic myeloproliferative neoplasms (MPN). JAK2 small-molecule inhibitors have been proven to be clinically efficacious; however, they are not mutation-specific and competent enough to suppress neoplastic clonal haematopoiesis. There is a need for exploring new therapeutic strategies for MPN. Additional signaling systems, such as PI3K/Akt/mTOR and Hedgehog, are a potential treatment target. The aim of this study was to characterise and compare the effects of specific JAK/STAT, PI3K/Akt/mTOR, and Hedgehog signaling inhibitors in haematological cell cultures. JAK2 p.V617F mutated SET-2 and JAK2 wild-type UT-7 human cell lines were employed in our study. The effect of specific signaling pathway inhibitors was studied as time- and dose-response experiments. Viability was measured by trypan blue exclusion and alamarBlue assays. IC50 values were used to compare the effectiveness of inhibitors in decreasing cell viability. Independent sample t-test was used for statistical comparisons between experimental groups. p < 0.05 was considered significant. Our results indicate that all specific inhibitors progressively reduced the number of viable cells as the concentration and exposure duration increased. Inhibitors impaired the proliferation of JAK2 mutated cells at significantly lower doses compared to wild-type JAK2 cell line. These in vitro data indicate that JAK/STAT and alternative PI3K/Akt/mTOR and Hedgehog inhibitors have a potential anti-proliferative efficacy. Future studies, involving direct screening of PI3K/Akt/ mTOR, JAK/STAT, and Hedgehog signaling molecules activity, at gene and protein level in cell-based MPN model, are required.

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