2017 EORTC: Characterisation of a novel ERK1/2 inhibitor, which modulates the phosphorylation and catalytic activity of ERK1/2

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Characterisation of a novel ERK1/2 inhibitor, which modulates the phosphorylation and catalytic activity of ERK1/2

Summary

  • The MAPK pathway is commonly hyper-activated in human cancers due to the occurrence of oncogenic mutations in RAF and RAS, and multiple studies have demonstrated that MAPK pathway inhibition suppresses the growth of such cancer cells.
  • MAPK inhibition has been clinically validated by BRAF and MEK inhibitors, which are approved for the treatment of BRAFV600E-mutant melanoma and NSCLC. However, response to such agents is often short-lived due to the onset of resistance mechanisms that result in re-activation of ERK1/2 (ERK) signalling1,2.
  • RAF and MEK inhibitors have also been clinically tested in other cancers, including BRAF-mutant colorectal (CRC) and KRAS-mutantNon-small cell lung cancer (NSCLC), where they had limited clinical activity3,4.
  • As ERK is the primary downstream effector of the MAPK pathway, it is hypothesized that ERK inhibitors may prove to be less susceptible to oncogenic bypass than RAF and MEK inhibitors and therefore have the potential to overcome the limitations of RAF and MEK inhibitors.
  • Using fragment-based drug discovery we have developed a novel, potent and selective ERK inhibitor, which inhibits in vitro ERK catalytic activity with a low nMIC50 value and has strong anti-proliferative effects in a wide range of MAPK-activated cell lines.
  • In addition to inhibiting ERK catalytic activity, the compound also inhibits the phosphorylation of ERK by MEK and confers a decrease in cellular pERKlevels in both BRAF-mutant and KRAS-mutant cell lines (in vitro and in in vivo pharmacodynamic[PD] studies).
  • Once daily oral dosing of the lead compound (50 mg/kg) conferred significant anti-tumoractivity in a range of in vivo models.
  • These data support the further optimisation of this series of compounds for clinical development.

2020 EHA: Characterization of ASTX660, an antagonist of cIAP1/2 and XIAP, in mouse models of T cell lymphoma

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Characterization of ASTX660, an antagonist of cIAP1/2 and XIAP, in mouse models of T cell lymphoma

Abstract:

Background:  ASTX660 is a potent, non-peptidomimetic antagonist of the cellular and X-linked inhibitors of apoptosis proteins (cIAP1/2 and XIAP), which is currently being tested in a first in human phase I-II study in patients with advanced solid tumor and lymphomas (NCT02503423). IAP antagonists have been reported to exhibit broad immuno-modulatory effects on both the innate and adaptive immune systems .

Aims:  We have investigated the profile of ASTX660 in preclinical T cell lymphoma models and evaluated ASTX660’s ability to enhance immune mediated killing of T cell lymphoma cells, both in vitro and in vivo.

Methods:  ASTX660 was tested in a panel of human and mouse tumor cell lines, assessing apoptosis, necroptosis and immunogenic cell death (ICD). ASTX660 was tested in vitro alone or with recombinant death receptor ligands (TNFa, FASL or TRAIL) and with or without caspase-8/RIPK inhibitors to demonstrate mechanism of action. Target engagement along with induction of apoptosis, necroptosis and ICD markers were analysed by Western blotting, and flow cytometry. Murine tumor models in immunocompetent and immunocompromised mice were utilised to evaluate the efficacy of ASTX660 in the presence or absence of an effective immune response. The Nanostring IO360 panel was used to assess immune cell recruitment.

Results:  ASTX660 antagonised IAPs in cell lines, as indicated by a decrease in cIAP1 protein levels and disruption of the XIAP:SMAC protein complex. In murine T cell lymphoma cell lines (BW5147, EL4 and L5178Y), ASTX660 treatment was associated with an increase in apoptosis or necroptosis and ICD biomarkers. In immunocompetent mice, administration of ASTX660 delivered a complete regression of BW5147 tumor growth, which was not seen in mice deficient in T and B cells. These mice remained refractory to subsequent rechallenge after initial complete regression. Biomarker evaluation from this model indicated a potent immunogenic/necroptotic response after ASTX660 dosing and upregulation of immune effector cells.

References:
1. Michie J. et al., The Immuno-Modulatory Effects of Inhibitor of Apoptosis Proteins Antagonists, Cells, 2020, 9(1), 207.
2. A. Hollebecque et al., AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics, 2019.

 

2020 EHA: ASTX295, a novel small molecule MDM2 antagonist, demonstrates potent activity in AML in combination with decitabine

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ASTX295, a novel small molecule MDM2 antagonist, demonstrates potent activity in AML in combination with decitabine

Abstract:

Background: The tumour suppressor p53 is activated in response to various stress signals to induce transcriptional changes leading to cellular responses such as cell cycle arrest and apoptosis. Activity of p53 is tightly regulated by the E3 ubiquitin ligase MDM2, which inhibits p53 function by, for example, targeting it for proteasomal degradation. Targeting the MDM2-p53 interaction to restore p53 function, is therefore, a promising strategy for cancer therapy and a number of these compounds are in clinical development including ASTX295 (NCT03975387). ASTX295 is a novel, orally bioavailable MDM2 antagonist developed through structure-based drug design that has demonstrated potent activity in a range of p53 wild-type pre-clinical models.

Aims: We investigated the therapeutic potential of ASTX295 alone and in combination with decitabine, a DNA- hypomethylating agent, in AML.

Methods: Primary blasts were isolated from AML patient samples using a combination of antibodies against CD34, CD33, CD45 and CD117. A panel of AML cell lines and primary AML blasts were treated with decitabine and ASTX295 at a range of concentrations, alone and in combination. After treatment, viability was assessed by Alamar blue assay or induction of apoptosis by flow cytometry using a fluorescent caspase substrate or Annexin V.

Effects of drug combinations were analysed using the Combenefit software based on different mathematical models (Loewe, bliss & HSA). Target engagement was confirmed by western blotting.

Results: When tested in a panel of p53 wild-type AML cell lines, ASTX295 exerted a strong anti-proliferative effect in which GI50 <30 nM was observed in 9 out of 11 cell lines. Additionally, p53 activation by ASTX295 triggered apoptosis in both AML cell lines, and primary AML blasts isolated from patients.

Activity of ASTX295 was further enhanced by combining with decitabine. Treatment of AML cell lines with ASTX295 and decitabine showed an increase in growth inhibitory effect and apoptosis compared to respective single agent treatments. This combinatory effect, as assessed by Combenefit, was also observed in primary AML blasts in which 7 of 12 samples tested demonstrated increased apoptosis at or above 300 nM ASTX295 and 100 nM decitabine. Target engagement of ASTX295 and decitabine was confirmed by upregulation of p53 transcriptional targets and decreased DNMT-1 expression.

Summary/Conclusion: Our findings demonstrate that the combination of ASTX295 with decitabine exhibits potent activity against p53 wild-type AML cells, and thus merits further investigation.

Erlanson et al., “Fragment-to-Lead Medicinal Chemistry Publications in 2018”; Journal of Medicinal Chemistry, 2020

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Erlanson et al., “Fragment-to-Lead Medicinal Chemistry Publications in 2018”; Journal of Medicinal Chemistry, 2020

https://pubs.acs.org/doi/10.1021/acs.jmedchem.9b01581

Coyle et al., “Applied Biophysical Methods in Fragment-Based Drug Discovery”; SLAS DISCOVERY, 2020

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Coyle et al., “Applied Biophysical Methods in Fragment-Based Drug Discovery”; SLAS DISCOVERY, 2020

https://journals.sagepub.com/doi/full/10.1177/2472555220916168?url_ver=Z39.88-2003&rfr_id=ori:rid:crossref.org&rfr_dat=cr_pub%20%200pubmed

Saur et al. “Fragment-based drug discovery using cryo-EM”, Drug Discovery Today, 2020

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Saur et al. “Fragment-based drug discovery using cryo-EM“, Drug Discovery Today, 2020

https://www.sciencedirect.com/science/article/pii/S1359644619304659?viewFullText=true

Kidger et al., “Dual-mechanism ERK1/2 inhibitors exploit a distinct binding mode to block phosphorylation and nuclear accumulation of ERK1/2”; Mol Cancer Ther, 2020

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Kidger et al., “Dual-mechanism ERK1/2 inhibitors exploit a distinct binding mode to block phosphorylation and nuclear accumulation of ERK1/2”; Mol Cancer Ther, 2020

https://doi.org/10.1158/1535-7163.mct-19-0505

Osbourne et al., “Fragments: where are we now?”; Biochemical Society Transactions, 2020

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Osbourne et al., “Fragments: where are we now?”; Biochemical Society Transactions, 2020

https://portlandpress.com/biochemsoctrans/article-abstract/48/1/271/221949/Fragments-where-are-we-now

Rathi et al., “Practical High-Quality Electrostatic Potential Surfaces for Drug Discovery Using a Graph-Convolutional Deep Neural Network”; Journal of Medicinal Chemistry, 2020

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Rathi et al., “Practical High-Quality Electrostatic Potential Surfaces for Drug Discovery Using a Graph-Convolutional Deep Neural Network”, Journal of Medicinal Chemistry, 2020

https://pubs.acs.org/doi/pdf/10.1021/acs.jmedchem.9b01129

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Mortenson et al., “Fragment-to-Lead Medicinal Chemistry Publications in 2017”, Journal of Medicinal Chemistry, 2019

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Mortenson et al., “Fragment-to-Lead Medicinal Chemistry Publications in 2017“, Journal of Medicinal Chemistry, 2019

https://pubs.acs.org/doi/pdf/10.1021/acs.jmedchem.8b01472