Urs Hancox

Feedback Suppression of PI3Kα Signaling in PTEN-Mutated Tumors Is Relieved by Selective Inhibition of PI3Kβ

In PTEN-mutated tumors, we show that PI3Kα activity is suppressed and PI3K signaling is driven by PI3Kβ. A selective inhibitor of PI3Kβ inhibits the Akt/mTOR pathway in these tumors but not in those driven by receptor tyrosine kinases. However, inhibition of PI3Kβ only transiently inhibits Akt/mTOR signaling because it relieves feedback inhibition of IGF1R and other receptors and thus causes activation of PI3Kα and a rebound in downstream signaling. This rebound is suppressed and tumor growth inhibition enhanced with combined inhibition of PI3Kα and PI3Kβ. In PTEN-deficient models of prostate cancer, this effective inhibition of PI3K causes marked activation of androgen receptor activity. Combined inhibition of both PI3K isoforms and androgen receptor results in major tumor regressions.

Small-molecule androgen receptor downregulators as an approach to treatment of advanced prostate cancer

Chemical starting points were investigated for downregulation of the androgen receptor as an approach to treatment of advanced prostate cancer. Although prototypic steroidal downregulators such as 6a designed for intramuscular administration showed insufficient cellular potency, a medicinal chemistry program derived from a novel androgen receptor ligand 8a led to 6-[4-(4-cyanobenzyl)piperazin-1-yl]-3-(trifluoromethyl)[1,2,4]triazolo[4,3-b]pyridazine (10b), for which high plasma levels following oral administration in a preclinical model compensate for moderate cellular potency.

Discovery of (R)-8-(1-(3,5-difluorophenylamino)ethyl)-N,N-dimethyl-2-morpholino-4-oxo-4H-chromene-6-carboxamide (AZD8186): a potent and selective inhibitor of PI3Kβ and PI3Kδ for the treatment of PTEN-deficient cancers.

Several studies have highlighted the dependency of PTEN deficient tumors to PI3Kβ activity and specific inhibition of PI3Kδ has been shown activity against human B-cell cancers. We describe the discovery and optimization of a series of 8-(1-anilino)ethyl)-2-morpholino-4-oxo-4H-chromene-6-carboxamides as PI3Kβ/δ inhibitors, which led to the discovery of the clinical candidate 13, also known as AZD8186. On the basis of the lower lipophilicity of the chromen-4-one core compared to the previously utilized pyrido[1,2-a]pyrimid-4-one core, this series of compounds displayed high metabolic stability and suitable physical properties for oral administration. Compound 13 showed profound pharmacodynamic modulation of p-Akt in PTEN-deficient PC3 prostate tumor bearing mice after oral administration and showed complete inhibition of tumor growth in the mouse PTEN-deficient PC3 prostate tumor xenograft model. 13 was selected as a clinical candidate for treatment of PTEN-deficient cancers and has recently entered phase I clinical trials.

Inhibition of PI3Kβ Signaling with AZD8186 Inhibits Growth of PTEN-Deficient Breast and Prostate Tumors Alone and in Combination with Docetaxel

Loss of PTEN protein results in upregulation of the PI3K/AKT pathway, which appears dependent on the PI3Kβ isoform. Inhibitors of PI3Kβ have potential to reduce growth of tumors in which loss of PTEN drives tumor progression. We have developed a small-molecule inhibitor of PI3Kβ and PI3Kδ (AZD8186) and assessed its antitumor activity across a panel of cell lines. We have then explored the antitumor effects as single agent and in combination with docetaxel in triple-negative breast (TNBC) and prostate cancer models. In vitro, AZD8186 inhibited growth of a range of cell lines. Sensitivity was associated with inhibition of the AKT pathway. Cells sensitive to AZD8186 (GI50 < 1 μmol/L) are enriched for, but not exclusively associated with, PTEN deficiency. In vivo, AZD8186 inhibits PI3K pathway biomarkers in prostate and TNBC tumors. Scheduling treatment with AZD8186 shows antitumor activity required only intermittent exposure, and that increased tumor control is achieved when AZD8186 is used in combination with docetaxel. AZD8186 is a potent inhibitor of PI3Kβ with activity against PI3Kδ signaling, and has potential to reduce growth of tumors dependent on dysregulated PTEN for growth. Moreover, AZD8186 can be combined with docetaxel, a chemotherapy commonly used to treat advanced TBNC and prostate tumors. The ability to schedule AZD8186 and maintain efficacy offers opportunity to combine AZD8186 more effectively with other drugs. Mol Cancer Ther; 14(1); 48–58. ©2014 AACR.

Discovery of 9-(1-anilinoethyl)-2-morpholino-4-oxo-pyrido[1,2-a]pyrimidine-7-carboxamides as PI3Kβ/δ inhibitors for the treatment of PTEN-deficient tumours

Starting from TGX-221, we designed a series of 9-(1-anilinoethyl)-2-morpholino-4-oxo-pyrido[1,2-a]pyrimidine-7-carboxamides as potent and selective PI3Kβ/δ inhibitors. Structure-activity relationships and structure-property relationships around the aniline and the amide substituents are discussed. We identified compounds 17 and 18, which showed profound pharmacodynamic modulation of phosphorylated Akt in the PC3 prostate tumour xenograft, after a single oral dose. Compound 17 also gave significant inhibition of tumour growth in the PC3 prostate tumour xenograft model after chronic oral dosing.

Intermittent High-Dose Scheduling of AZD8835, a Novel Selective Inhibitor of PI3Kα and PI3Kδ, Demonstrates Treatment Strategies for PIK3CA-Dependent Breast Cancers

The PIK3CA gene, encoding the p110α catalytic unit of PI3Kα, is one of the most frequently mutated oncogenes in human cancer. Hence, PI3Kα is a target subject to intensive efforts in identifying inhibitors and evaluating their therapeutic potential. Here, we report studies with a novel PI3K inhibitor, AZD8835, currently in phase I clinical evaluation. AZD8835 is a potent inhibitor of PI3Kα and PI3Kδ with selectivity versus PI3Kβ, PI3Kγ, and other kinases that preferentially inhibited growth in cells with mutant PIK3CA status, such as in estrogen receptor–positive (ER+) breast cancer cell lines BT474, MCF7, and T47D (sub-μmol/L GI50s). Consistent with this, AZD8835 demonstrated antitumor efficacy in corresponding breast cancer xenograft models when dosed continuously. In addition, an alternative approach of intermittent high-dose scheduling (IHDS) was explored given our observations that higher exposures achieved greater pathway inhibition and induced apoptosis. Indeed, using IHDS, monotherapy AZD8835 was able to induce tumor xenograft regression. Furthermore, AZD8835 IHDS in combination with other targeted therapeutic agents further enhanced antitumor activity (up to 92% regression). Combination partners were prioritized on the basis of our mechanistic insights demonstrating signaling pathway cross-talk, with a focus on targeting interdependent ER and/or CDK4/6 pathways or alternatively a node (mTOR) in the PI3K-pathway, approaches with demonstrated clinical benefit in ER+ breast cancer patients. In summary, AZD8835 IHDS delivers strong antitumor efficacy in a range of combination settings and provides a promising alternative to continuous dosing to optimize the therapeutic index in patients. Such schedules merit clinical evaluation. Mol Cancer Ther; 15(5); 877–89. ©2016 AACR.

Discovery of 1-(4-(5-(5-amino-6-(5-tert-butyl-1,3,4-oxadiazol-2-yl)pyrazin-2-yl)-1-ethyl-1,2,4-triazol-3-yl)piperidin-1-yl)-3-hydroxypropan-1-one (AZD8835): A potent and selective inhibitor of PI3Kα and PI3Kδ for the treatment of cancers

Starting from potent inhibitors of PI3Kα having poor general kinase selectivity (e.g., 1 and 2), optimisation of this series led to the identification of 25, a potent inhibitor of PI3Kα (wild type, E545K and H1047R mutations) and PI3Kδ, selective versus PI3Kβ and PI3Kγ, with excellent general kinase selectivity. Compound 25 displayed low metabolic turnover and suitable physical properties for oral administration. In vivo, compound 25 showed pharmacodynamic modulation of AKT phosphorylation and near complete inhibition of tumour growth (93% tumour growth inhibition) in a murine H1047R PI3Kα mutated SKOV-3 xenograft tumour model after chronic oral administration at 25mg/kg b.i.d. Compound 25, also known as AZD8835, is currently in phase I clinical trials.

Discovery of a series of 8-(2,3-dihydro-1,4-benzoxazin-4-ylmethyl)-2-morpholino-4-oxo-chromene-6-carboxamides as PI3Kβ/δ inhibitors for the treatment of PTEN-deficient tumours

We report the discovery and optimisation of a series of 8-(2,3-dihydro-1,4-benzoxazin-4-ylmethyl)-2-morpholino-4-oxo-chromene-6-carboxamides, leading to compound 16 as a potent and selective PI3Kβ/δ inhibitor: PI3Kβ cell IC50 0.012 μM (in PTEN null MDA-MB-468 cell) and PI3Kδ cell IC50 0.047 μM (in Jeko-1 B-cell), with good pharmacokinetics and physical properties. In vivo, 16 showed profound pharmacodynamic modulation of AKT phosphorylation in a mouse PTEN-deficient PC3 prostate tumour xenograft after a single oral dose and gave excellent tumour growth inhibition in the same model after chronic oral dosing. Compound 16 was selected as a preclinical candidate for the treatment of PTEN-deficient tumours.

Abstract 3264: AZD8186: a potent selective inhibitor of PI3Kβ targeting PTEN-deficient tumours dependent on dysregulated PI3Kβ signalling.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC AZD8186 is a novel potent small molecule that targets the lipid kinase PI3Kβ with selectivity vs PI3Kα. AZD8186 reduces pAKT-S473 in the PTEN deficient MDA-MB-468 cell line with an IC50 <5nM, while in the PI3Kα-dependent PIK3CA mutant line BT474 it is 200 fold less potent. AZD8186 also demonstrates cellular activity versus PI3Kβ, inhibiting IgM-dependent pAKT-S473 with an IC50 of 15nM. In cell line panels, sensitivity to AZD8186 is associated in part with loss of PTEN function suggesting that targeting tumours that have lost normal levels of PTEN function through deletion, mutation or down regulation might enrich for sensitivity to AZD8186. Deficiencies in PTEN are described in a number of tumour types such as prostate, squamous lung, breast, renal and glioblastoma. AZD8186 has single anti-tumour activity in pre-clinical models representing each of these settings, which is associated with dynamic regulation of a number of key pathway biomarkers. Clinically, agents such as AZD8186 are likely to be used in combination. The use of AZD8186 in conjunction with either androgen therapy or docetaxol has been explored in cell lines and pre-clinical disease models. Combining AZD8186 with MDV-3100 resulted in synergistic inhibition of LNCAP prostate tumour cell growth, demonstrating the potential to combine with androgen therapy. Moreover in HCC70 (breast) and PC3 (prostate) tumour xenografts AZD8186 shows increased benefit in combination with docetaxel, demonstrating the ability to combine with cytotoxic chemotherapy. Exploration of dose and schedule revealed that efficacy can be maintained in combination with docetaxol at a lower dose of AZD8186 than required for monotherapy activity. Moreover efficacy was maintained when the frequency of AZD8186 dosing was reduced using a shorter intermittent schedule. Understanding how AZD8186 combines with standards of care for PTEN null disease segments provides important insight into how agents targeting PI3Kβ-dependent tumours may be used most effectively in the clinic. [Current affiliation for S. Cosulich is Novartis, Basel, Switzerland.] Citation Format: Urs Hancox, Sabina Cosulich, Hannah Dry, Lynsey Hanson, Clare Crafter, Bernard Barlaam, Martina Fitzek, Lara Ward, Marie Cumberbatch, Steve Powell, Rebecca Ellston, Mandy Lawson, Steve Wedge, Liz Harrington, Stephen Green, Simon T. Barry. AZD8186: a potent selective inhibitor of PI3Kβ targeting PTEN-deficient tumours dependent on dysregulated PI3Kβ signalling. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3264. doi:10.1158/1538-7445.AM2013-3264

Inhibiting PI3Kβ with AZD8186 Regulates Key Metabolic Pathways in PTEN-Null Tumors

Purpose: PTEN-null tumors become dependent on the PI3Kβ isoform and can be targeted by molecules such as the selective PI3Kβ inhibitor AZD8186. However, beyond the modulation of the canonical PI3K pathway, the consequences of inhibiting PI3Kβ are poorly defined. Experimental Design: To determine the broader impact of AZD8186 in PTEN-null tumors, we performed a genome-wide RNA-seq analysis of PTEN-null triple-negative breast tumor xenografts treated with AZD8186. Mechanistic consequences of AZD8186 treatment were examined across a number of PTEN-null cell lines and tumor models. Results: AZD8186 treatment resulted in modification of transcript and protein biomarkers associated with cell metabolism. We observed downregulation of cholesterol biosynthesis genes and upregulation of markers associated with metabolic stress. Downregulation of cholesterol biosynthesis proteins, such as HMGCS1, occurred in PTEN-null cell lines and tumor xenografts sensitive to AZD8186. Therapeutic inhibition of PI3Kβ also upregulated PDHK4 and increased PDH phosphorylation, indicative of reduced carbon flux into the TCA cycle. Consistent with this, metabolomic analysis revealed a number of changes in key carbon pathways, nucleotide, and amino acid biosynthesis. Conclusions: This study identifies novel mechanistic biomarkers of PI3Kβ inhibition in PTEN-null tumors supporting the concept that targeting PI3Kβ may exploit a metabolic dependency that contributes to therapeutic benefit in inducing cell stress. Considering these additional pathways will guide biomarker and combination strategies for this class of agents. Clin Cancer Res; 23(24); 7584–95. ©2017 AACR.