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Next-generation oral selective estrogen receptor degraders (ngSERDs) are aiming to become the backbone endocrine therapy (ET) for patients with estrogen receptor positive (ER+) breast cancer (BC) by achieving greater ER signaling blockade than current therapies and tackling key mechanisms of resistance. Camizestrant (AZD9833) is an ngSERD for the treatment of ER+ BC which has demonstrated selective ERα degradation, pure ER antagonism and significant anti-tumor activity in ESR1 wild-type (ESR1wt) and mutant (ESR1m) tumors, as well as encouraging clinical activity in early phase trials. ER+ BC is responsive to therapies targeting ER and CDK4/6 signaling in the adjuvant and metastatic settings. To explore camizestrant combination potential as a backbone ET, camizestrant was partnered with either palbociclib or abemaciclib in CDK4/6 inhibitor (CDK4/6i) naïve and resistant in vitro and in vivo models. Combination benefit was observed in vitro in 3 parental ER+ BC cell lines, moreover camizestrant plus abemaciclib showed activity in palbociclib-resistant cell lines including lines harboring CCNE1amp and RB1 loss. In vivo, combination of camizestrant and either CDK4/6i was well tolerated with the combinations promoting improved efficacy in ESR1wt and ESR1m PDX tumor models compared to monotherapy arms. ER+ BC has a high prevalence of alterations in the PI3K/AKT/PTEN pathway which provides opportunities for treatment with PI3K/AKT pathway inhibitors. Camizestrant delivered enhanced efficacy when combined with mTORC1 inhibitor everolimus and AKT inhibitor capivasertib in CTC-174, an ESR1m and PI3KCAm tumor model. Additionally, combination with capivasertib was more efficacious than single treatments, at clinically relevant dose and schedules, in both PI3Kwt and mutated pathway in PDX models. These data demonstrate the interplay between PI3K/AKT/PTEN pathway inhibition and camizestrant mechanism of action. Finally, we explored a potential triple combination of camizestrant, capivasertib and palbociclib in Palbociclib-resistant models representative of PI3KCA/AKT/PTEN wt and altered tumors. This strategy delivered robust efficacy across these models comparing to single treatments and double combinations. The triplet led to durable regressions at clinically achievable doses of all compounds, irrespective of genetic background and aligned to biomarker modulation of the three signaling axis. These preclinical data demonstrate the potential of camizestrant to become the backbone ET, with high combinability in vivo with inhibitors of CDK4/6, mTOR and AKT. These combinations demonstrate the opportunity to impact care of patients with early and metastatic ER+BC, delivering benefit to broad patient populations including those with ESR1wt or ESR1m tumors, independent of PI3K/AKT/PTEN pathway mutation status, and in patients with both CDK4/6i sensitive and resistant tumors. Citation Format: Larissa Carnevalli, Susana Ros, Jelena Urosevic, Natalie Cureton, Sophie Darcy, Mandy Lawson, Stuart Williamson, Pablo Morentin Gutierrez, Azadeh Bashi, Jennifer Moss, Christopher Morrow, Barry Simon, Teresa Klinowska. PD10-04 Combination of the next generation oral SERD camizestrant (AZD9833) with CDK4/6 and mTOR/AKT inhibitors delivers robust efficacy in a broad range of ER+ breast tumors. [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD10-04.

PDF file - 235KB, Activity of AZD9291 and metabolites against rare EGFR mutations using cellular phosphorylation assays.

PDF file - 294KB, Phenotype and phosphorylation assay data for AZD9291 against lung cancer associated HER2 mutation.

Supp Fig 1 Tumour efficacy Supp Fig 2 In vitro gene expression modulation Supp Fig 3 In vivo biomarker analysis Supp Fig 4 HCC70 In vivo FDG uptake imaging Supp Fig 5 U87MG in vivo FDG uptake imaging

PDF file - 107KB, Mass Spectrometry study showing covalent binding of AZD9291 to cysteine 797 in EGFR T790M.

Pharmacokinetics modelling of AZD2014 intermittent dosing schedule.

Suppressive myeloid cells mediate resistance to immune checkpoint blockade. PI3Kγ inhibition can target suppressive macrophages, and enhance efficacy of immune checkpoint inhibitors. However, how PI3Kγ inhibitors function in different tumor microenvironments (TME) to activate specific immune cells is underexplored. The effect of the novel PI3Kγ inhibitor AZD3458 was assessed in preclinical models. AZD3458 enhanced antitumor activity of immune checkpoint inhibitors in 4T1, CT26, and MC38 syngeneic models, increasing CD8+ T-cell activation status. Immune and TME biomarker analysis of MC38 tumors revealed that AZD3458 monotherapy or combination treatment did not repolarize the phenotype of tumor-associated macrophage cells but induced gene signatures associated with LPS and type II INF activation. The activation biomarkers were present across tumor macrophages that appear phenotypically heterogenous. AZD3458 alone or in combination with PD-1–blocking antibodies promoted an increase in antigen-presenting (MHCII+) and cytotoxic (iNOS+)-activated macrophages, as well as dendritic cell activation. AZD3458 reduced IL-10 secretion and signaling in primary human macrophages and murine tumor-associated macrophages, but did not strongly regulate IL-12 as observed in other studies. Therefore, rather than polarizing tumor macrophages, PI3Kγ inhibition with AZD3458 promotes a cytotoxic switch of macrophages into antigen-presenting activated macrophages, resulting in CD8 T-cell–mediated antitumor activity with immune checkpoint inhibitors associated with tumor and peripheral immune activation.

Supplementary Figure 1. Pharmacokinetic (PK) properties of AZD3458 in mouse. Supplementary Figure 2. In vivo pharmacodynamic properties of AZD3458 in mouse neutrophils. Supp Fig 3. PI3K-�/mTOR pathway regulates expression of IL-10/IL-12 via LPS-induced glycolytic burst. Supplementary Figure 4. AZD3458 improves checkpoint activity in CT-26 tumor models. Supplementary Figure 5. AZD3458 improves checkpoint activity in 4T1 tumor models. Supplementary Figure 6. Top canonical IPA pathways across tissues in AZD3458+PD-1 combination group. Supplementary Figure 7. Graphical abstract.

HCC1428-eveR and HCC1428-LTED-eveR cell lines resistant to everolimus remain sensitive to dual mTORC1/2 inhibitor AZD2014.

PDF file - 312KB, Efficacy of AZD9291 in H3255, PC-9VanR and A431 xenograft models. Weight loss data for chronically dosed AZD9291 in PC-9 and H1975 xenograft models.

Supplementary Materials and Methods. Suplementary Table 1. Tumor Immuno-phenotyping. Suplementary Table 2. Flow cytometry panels.

Loss of the tumor suppressor PTEN confers a tumor cell dependency on the PI3Kβ isoform. Achieving maximal inhibition of tumor growth through PI3K pathway inhibition requires sustained inhibition of PI3K signaling; however, efficacy is often limited by suboptimal inhibition or reactivation of the pathway. To select combinations that deliver comprehensive suppression of PI3K signaling in PTEN-null tumors, the PI3Kβ inhibitor AZD8186 was combined with inhibitors of kinases implicated in pathway reactivation in an extended cell proliferation assay. Inhibiting PI3Kβ and mTOR gave the most effective antiproliferative effects across a panel of PTEN-null tumor cell lines. The combination of AZD8186 and the mTOR inhibitor vistusertib was also effective in vivo controlling growth of PTEN-null tumor models of TNBC, prostate, and renal cancers. In vitro, the combination resulted in increased suppression of pNDRG1, p4EBP1, as well as HMGCS1 with reduced pNDRG1 and p4EBP1 more closely associated with effective suppression of proliferation. In vivo biomarker analysis revealed that the monotherapy and combination treatment consistently reduced similar biomarkers, while combination increased nuclear translocation of the transcription factor FOXO3 and reduction in glucose uptake. These data suggest that combining the PI3Kβ inhibitor AZD8186 and vistusertib has potential to be an effective combination treatment for PTEN-null tumors. Mol Cancer Ther; 17(11); 2309–19. ©2018 AACR.

Supplementary Table 1: Cell lines; Supplementary Table 2: Antibodies; Supplementary Table 3: Gene lists

Supplementary data on cell lines

Pharmacokinetics and PK/PD relationships for AZD2014

PDF file - 151KB, Activity of AZD9291 and metabolites against cellular phospho-HER2 assays.

PDF file - 470KB, Effect of AZD9291 and AZ5104 in transgenic models of EGFR-TKI sensitizing (C/L858R and C/ex19del) and T790M resistant (C/L+T) lung cancer.

Supplementary table 1: Biochemical and cellular activity of AZD2014. Supplementary table 2: Biochemical activity of AZD2014. Supplementary table 3: Anti proliferative activity of AZD2014 in a panel of ER+, endocrine resistant and LTED cell lines. Supplementary table 4: Activity of AZD2014 in a number of ER+ breast cancer in vivo models.

PDF file - 182KB, Pharmacokinetic analysis of AZD9291, AZ5104 and AZ7550 total plasma concentrations vs time following single oral dose of 25mg/kg AZD9291 in mice.

PDF file - 89KB, Additional biochemical and cellular profiling data for AZD9291 and metabolites.

Supplementary Figure from A Phase I Study Investigating AZD8186, a Potent and Selective Inhibitor of PI3Kβ/δ, in Patients with Advanced Solid Tumors

Supplementary Figures: Supplementary Figure 1. Summary of duration on AZD9496 therapy, previous therapies, and circulating biomarkers status at baseline for each patient ranked by ascending dose; Supplementary Figure 2. Tumor pharmacodynamics. Supplementary Tables: Supplementary Table 1. Study Plan showing time-points of collection of biosamples for exploratory research; Supplementary Table 2. ddPCR primers and probes;Supplementary Table 3. Antibodies and conditions for IHC analysis Antibodies for IHC analysis; Supplementary Table 4. Patients with {greater than or equal to}1 CTC/7.5 mL WB within 120 days of fulvestrant wash-out period prior C1D1.

Purpose:To characterize safety and tolerability of the selective PI3Kβ inhibitor AZD8186, identify a recommended phase II dose (RP2D), and assess preliminary efficacy in combination with abiraterone acetate or vistusertib.Patients and Methods:This phase I open-label study included patients with advanced solid tumors, particularly prostate cancer, triple-negative breast cancer, and squamous non–small cell lung cancer. The study comprised four arms: (i) AZD8186 monotherapy dose finding; (ii) monotherapy dose expansion; (iii) AZD8186/abiraterone acetate (with prednisone); and (iv) AZD8186/vistusertib. The primary endpoints were safety, tolerability, and identification of the RP2D of AZD8186 monotherapy and in combination. Secondary endpoints included pharmacokinetics (PK), pharmacodynamics, and tumor and prostate-specific antigen (PSA) responses.Results:In total, 161 patients were enrolled. AZD8186 was well tolerated across all study arms, the most common adverse events being gastrointestinal symptoms. In the monotherapy dose-finding arm, four patients experienced dose-limiting toxicities (mainly rash). AZD8186 doses of 60-mg twice daily [BID; 5 days on, 2 days off (5:2)] and 120-mg BID (continuous and 5:2 dosing) were taken into subsequent arms. The PKs of AZD8186 were dose proportional, without interactions with abiraterone acetate or vistusertib, and target inhibition was observed in plasma and tumor tissue. Monotherapy and combination therapy showed preliminary evidence of limited antitumor activity by imaging and, in prostate cancer, PSA reduction.Conclusions:AZD8186 monotherapy had an acceptable safety and tolerability profile, and combination with abiraterone acetate/prednisone or vistusertib was also tolerated. There was preliminary evidence of antitumor activity, meriting further exploration of AZD8186 in subsequent studies in PI3Kβ pathway–dependent cancers.

Purpose:Common resistance mechanisms to endocrine therapy (ET) in estrogen receptor (ER)–positive metastatic breast cancers include, among others, ER loss and acquired activating mutations in the ligand-binding domain of the ER gene (ESR1LBDm). ESR1 mutational mediated resistance may be overcome by selective ER degraders (SERD). During the first-in-human study of oral SERD AZD9496, early changes in circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) were explored as potential noninvasive tools, alongside paired tumor biopsies, to assess pharmacodynamics and early efficacy.Experimental Design:CTC were enumerated/phenotyped for ER and Ki67 using CellSearch in serial blood draws. ctDNA was assessed for the most common ESR1LBDm by droplet digital PCR (BioRad).Results:Before starting AZD9496, 11 of 43 (25%) patients had ≥5 CTC/7.5 mL whole blood (WB), none of whom underwent reduction to +, two of whom had CTC-ER+ reduction. CTC-Ki67 status did not change appreciably. Patients with ≥5 CTC/7.5 mL WB before treatment had worse progression-free survival (PFS) than patients with P = 0.0003). Fourteen of 45 (31%) patients had ESR1LBDm+ ctDNA at baseline, five of whom had ≥2 unique mutations. Baseline ESR1LBDm status was not prognostic. Patients with persistently elevated CTC and/or ESR1LBDm+ ctDNA at C1D15 had worse PFS than patients who did not (P = 0.0007).Conclusions:Elevated CTC at baseline was a strong prognostic factor in this cohort. Early on-treatment changes were observed in CTC-ER+ and ESR1LBDm+ ctDNA, but not in overall CTC number. Integrating multiple biomarkers in prospective trials may improve outcome prediction and ET resistance mechanisms' identification over a single biomarker.

Supplementary Data from AZD8931, an Equipotent, Reversible Inhibitor of Signaling by Epidermal Growth Factor Receptor, ERBB2 (HER2), and ERBB3: A Unique Agent for Simultaneous ERBB Receptor Blockade in Cancer

PK parameters for M5 following a single dose (Day 1) and multiple doses (Day 15)

GAZD9496 metabolite, M3 (Panel A) and M5 (Panel C), geometric mean plasma concentration following a single dose of AZD9496 on Day 1 (semi-log scale). AZD9496 metabolite, M3 (Panel B) and M5 (Panel D), geometric mean plasma concentration following multiple doses of AZD9496 on Day 15 (semi-log scale). On Day 15 at all doses except 20 mg, the 24 h time point is the 12 h trough concentration following the evening dose of AZD9496 and therefore not shown. aPooled data from dose escalation and expansion groups. BID=twice daily; QD=once daily; EXP=expansion cohort.

Supplementary Data from A Phase I Study Investigating AZD8186, a Potent and Selective Inhibitor of PI3Kβ/δ, in Patients with Advanced Solid Tumors

Purpose: AZD9496 is an oral nonsteroidal, small-molecule inhibitor of estrogen receptor alpha (ERα) and a potent and selective antagonist and degrader of ERα. This first-in-human phase I study determined the safety and tolerability of ascending doses of oral AZD9496 in women with estrogen receptor (ER)+/HER2− advanced breast cancer, characterized its pharmacokinetic (PK) profile, and made preliminary assessment of antitumor activity.Patients and Methods: Forty-five patients received AZD9496 [20 mg once daily (QD) to 600 mg twice daily (BID)] in a dose-escalation, dose-expansion “rolling 6” design. Safety, tolerability, and PK activity in each cohort were reviewed before escalating to the next dose. PK was determined by mass spectrometry. Adverse events (AEs) were graded according to the Common Terminology Criteria for Adverse Events (CTCAE) v4.0. Objective tumor response was evaluated by Response Evaluation Criteria in Solid Tumors (RECIST) v1.1.Results: Most common causally related AEs were diarrhea (35.6%), fatigue (31.1%), and nausea (22.2%), and seven patients had grade ≥3 AEs. Three patients experienced a dose-limiting toxicity: one each at 150 mg BID (abnormal hepatic function), 400 mg BID (diarrhea and elevated liver function tests), and 600 mg BID (diarrhea), and all were reversible. The maximum tolerated dose was not reached. Partial response was confirmed in one patient, who also had decreased tumor marker Ca15.3. Four patients had stable disease at 12 months' follow-up.Conclusions: AZD9496 is well tolerated with an acceptable safety profile, showing evidence of prolonged disease stabilization in heavily pretreated patients with ER+/HER2− advanced breast cancer. Clin Cancer Res; 24(15); 3510–8. ©2018 AACR.See related commentary by Jordan, p. 3480

A total of 45 patients were recruited and four to six patients were allocated to cohorts to receive escalating doses of AZD9496. In the BID cohorts, patients received QD dosing on Day 1 and BID dosing thereafter. Six patients entered the expansion cohort, receiving AZD9496 250 mg BID. BID=twice daily, DLT=dose-limiting toxicity; n=number of patients included; LFT=liver function tests; QD=once daily.

Purpose: To test the hypothesis that simultaneous, equipotent inhibition of epidermal growth factor receptor (EGFR; erbB1), erbB2 (human epidermal growth factor receptor 2), and erbB3 receptor signaling, using the novel small-molecule inhibitor AZD8931, will deliver broad antitumor activity in vitro and in vivo.Experimental Design: A range of assays was used to model erbB family receptor signaling in homodimers and heterodimers, including in vitro evaluation of erbB kinase activity, erbB receptor phosphorylation, proliferation in cells, and in vivo testing in a human tumor xenograft panel, with ex vivo evaluation of erbB phosphorylation and downstream biomarkers. Gefitinib and lapatinib were used to compare the pharmacological profile of AZD8931 with other erbB family inhibitors.Results: In vitro, AZD8931 showed equipotent, reversible inhibition of EGFR (IC50, 4 nmol/L), erbB2 (IC50, 3 nmol/L), and erbB3 (IC50, 4 nmol/L) phosphorylation in cells. In proliferation assays, AZD8931 was significantly more potent than gefitinib or lapatinib in specific squamous cell carcinoma of the head and neck and non–small cell lung carcinoma cell lines. In vivo, AZD8931 inhibited xenograft growth in a range of models while significantly affecting EGFR, erbB2, and erbB3 phosphorylation and downstream signaling pathways, apoptosis, and proliferation.Conclusions: AZD8931 has a unique pharmacologic profile providing equipotent inhibition of EGFR, erbB2, and erbB3 signaling and showing greater antitumor activity than agents with a narrower spectrum of erbB receptor inhibition in specific preclinical models. AZD8931 provides the opportunity to investigate whether simultaneous inhibition of erbB receptor signaling could be of utility in the clinic, particularly in the majority of solid tumors that do not overexpress erbB2. Clin Cancer Res; 16(4); 1159–69

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Background: SERENA-1 (NCT03616587) is a Phase 1, multi-part, open-label study of camizestrant in women with ER+/HER2− advanced breast cancer. Parts A/B and C/D (escalation/expansion) examined camizestrant as monotherapy and in combination with palbociclib respectively and have been presented previously.1,2 Here we present data from parts G/H which examined camizestrant in combination with abemaciclib. Methods: The primary objective was to determine the safety and tolerability of camizestrant 75 mg once daily (QD) in combination with abemaciclib 150 mg twice daily (BID). Secondary objectives included investigation of anti-tumor response and pharmacokinetics (PK). Participants were previously treated women of any menopausal status (pre-menopausal women received this combination alongside ongoing ovarian function suppressors). Prior treatment with ≤2 lines of chemotherapy in the advanced setting was permitted. There was no limit on the number of lines of prior endocrine treatment in the advanced setting; previous treatment with CDK4/6 inhibitors (CDK4/6i) and fulvestrant was permitted. Results: As of 1st June 2022, 24 patients had received camizestrant in combination with abemaciclib with a median 7.7 month follow up. Tolerability of the combination of camizestrant and abemaciclib was consistent with that of each drug individually. No patient required camizestrant dose reduction. All camizestrant-related heart rate decreases were Grade 1 (asymptomatic). PK data for camizestrant in combination with abemaciclib were consistent with camizestrant as monotherapy and published abemaciclib steady-state PK data, indicating no clinically relevant drug-drug interaction. In these heavily pre-treated patients (46% prior chemotherapy, 75% prior CDK4/6i, 54% prior fulvestrant; all in the advanced disease setting) and of whom 67% had visceral metastases, the objective response rate was 5/19 (26.3%), the clinical benefit rate at 24 weeks was 16/24 (66.7%) and the median progression-free survival had not been reached, with 8/24 patients experiencing a progression event. These data support the use of camizestrant 75 mg QD combined with the approved abemaciclib dose. Conclusions: Camizestrant 75 mg QD in combination with abemaciclib 150 mg BID was well tolerated with encouraging clinical activity. The inclusion of this regimen in the ongoing Phase 3, SERENA-6 trial 3, of camizestrant combined with CDK4/6i versus an aromatase inhibitor, will further clarify the role of this combination in the treatment of patients with ER+/HER2− advanced breast cancer with tumors expressing ESR1 mutations. References 1. Baird R, Oliveira M, Ciruelos Gil EM, et al. SABCS 2020 Virtual Meeting. Abstract PS11-05. 2. Oliveira M, Hamilton EP, Incorvati J, et al. J Clin Oncol 40, 2022 (suppl 16; abstr 1032). 3. SERENA-6 trial. Available at https://clinicaltrials.gov/ct2/show/NCT04964934 We acknowledge Helen Heffron, PhD, from InterComm International who provided medical writing support funded by AstraZeneca. Citation Format: Nicholas Turner, Christos Vaklavas, Emiliano Calvo, Javier Garcia-Corbacho, Jason Incorvati, Manuel Ruiz Borrego, Chris Twelves, Anne Armstrong, Begoña Bermejo, Erika Hamilton, Mafalda Oliveira, Eva Ciruelos, Peter Kabos, Manish R Patel, Maria Borrell, Howard Burris, Bruno de Paula, Alejandro Falcon, Cristina Hernando, Irene Moreno, Ciara S. O’Brien, Elena Shagisultanova, Ivan Victoria Ruiz, Judy S. Wang, Mei Wei, Tim Brier, Danielle Carroll, Carmela Ciardullo, Lisa Gibbons, itziar irurzun-Arana, Tony Jack, bistra kirova, Teresa Klinowska, Justin Lindemann, Julie Maidment, Alastair Mathewson, Rhiannon Maudsley, Robert McEwen, Christopher Morrow, Andy Sykes, Richard D. Baird. SERENA-1: Updated analyses from a Phase 1 study of the next generation oral selective estrogen receptor degrader camizestrant (AZD9833) combined with abemaciclib, in women with ER-positive, HER2-negative advanced breast cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-07-28.

Background Camizestrant (C), a next-generation oral selective estrogen receptor (ER) antagonist and degrader (ngSERD) has shown promising clinical activity in ER+ breast cancer (BC) in the Phase 1 SERENA-1 study1,2 with a dose-dependent safety profile. The Phase 2 randomized SERENA-2 study (NCT04214288) initially assessed three doses of C vs fulvestrant (F) in post-menopausal women with ER+ HER2˗ BC with disease recurrence or progression after ≤1 endocrine therapy (ET) in the advanced setting. Methods SERENA-2 evaluated efficacy and safety of C 75, 150 or 300 mg monotherapy QD vs F (per label). Eligible patients were randomized 1:1:1:1. The Primary objective was to determine clinical efficacy of C vs F by investigator-assessed progression-free survival (PFS). Secondary endpoints included objective response rate, response duration, clinical benefit rate at 24 weeks, overall survival and safety. Patients had no prior F or oral SERD and ≤1 ET and ≤1 chemotherapy (CTX) in the advanced setting. To assess the impact of prior CDK4/6 inhibitor (CDK4/6i) treatment, randomization was stratified so that 50% of patients had prior CDK4/6i. Planned enrolment of 288 patients began in April 2020. The C 300 arm was closed after 20 patients were enrolled, changing target enrolment to 236. By August 2021, 240 patients had been randomized. Primary analysis was triggered when 108 progression events (75% maturity) had occurred in the best performing pair (C vs F) in August 2022. Efficacy analyses compared C 75 and 150 mg doses with F, with no formal analyses of C 300 vs F. 108 events for pairwise comparison vs F gave 86% power at the 2-sided 10% significance level. Primary analyses used a Cox proportional hazards model to compare PFS, adjusting for prior CDK4/6i and lung/liver metastases. ESR1 mutations (ESR1m) were detected in plasma samples using next-generation sequencing. Results 119/240 (49.6%) patients had had prior CDK4/6i therapy. At baseline, 88 (36.7%) patients had detectable ESR1m and 140 (58.3%) had lung/liver metastases. Prior CTX or ET rates in the advanced setting were 19.2 and 65.4%. Treatment-emergent adverse events (AEs) (grade ≥3) occurred in 77.0 (12.2), 90.4 (21.9) and 68.5 (13.7) % of patients in the C 75, C 150 and F arms. AEs leading to treatment discontinuation occurred in 2.7, 0 and 0% of patients in the C 75, C 150 and F arms. The most common AEs considered by the investigator to be causally related to study drug were photopsia (18.4%) and (sinus) bradycardia (13.6%) – all were grade 1 or 2. Hot flush (2.7%) and myalgia (2.7%) were the most common AEs related to F. Conclusions SERENA-2 is the first Phase 2 trial investigating multiple dose levels of an ngSERD vs F in post-menopausal women with advanced ER+ HER2˗ BC with disease recurrence or progression after ≤1 ET in the advanced setting. C at both 75 and 150 mg dose levels showed a statistically significant and clinically meaningful benefit in PFS vs F in the overall study population, and was well tolerated. The results of SERENA-2 support the further development of C in ER+ HER2- BC. Acknowledgements AstraZeneca sponsored this trial and funded medical writing support from Helen Heffron (InterComm International). References 1. Baird R, Oliveira M, Ciruelos Gil EM, et al. SABCS 2020 Virtual Meeting. Abstract PS11-05; 2. Oliveira M, Hamilton EP, Incorvati J et al. ASCO 2022 Annual Meeting, Chicago, IL, USA. Abstract 1032. Citation Format: Mafalda Oliveira, Denys Pominchuck, Zbigniew Nowecki, Erika Hamilton, Yaroslav Kulyaba, Timur Andabekov, Yevhen Hotko, Tamar Melkadze, Gia Nemsadze, Patrick Neven, Yuriy Semegen, Vladmir Vladmirov, Claudio Zamagni, Hannelore Denys, Frederic Forget, Zsolt Horvath, Alfiya Nesterova, Maxine Bennett, Bistra Kirova, Teresa Klinowska, Justin Lindemann, Delphine Lissa, Alastair Mathewson, Christopher Morrow, Zuzana Traugottova, Ruaan Van Zyl, Ekaterine Arkania. GS3-02 Camizestrant, a next generation oral SERD vs fulvestrant in post-menopausal women with advanced ER-positive HER2-negative breast cancer: Results of the randomized, multi-dose Phase 2 SERENA-2 trial [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr GS3-02.

Background: AZD9833 is an oral selective estrogen receptor (ER) antagonist and degrader (SERD) in Phase 2 clinical development for the treatment of ER+ HER2− breast cancer. Here we report data from Parts C and D of the ongoing Phase 1 study (SERENA-1) examining AZD9833 in combination with palbociclib, together with updated data from Parts A and B examining AZD9833 monotherapy. Methods: SERENA-1 (NCT03616587) is an ongoing open-label Phase 1 study of AZD9833 in pre- and post-menopausal women with ER+, HER2− advanced breast cancer who have previously received ≥1 endocrine therapy and ≤2 prior chemotherapies. Prior treatment with fulvestrant and/or CDK4/6 inhibitors was permitted. The primary objective is to determine the safety and tolerability of once daily (QD) AZD9833, with dose-limiting toxicities (DLTs) in the first 28 days defining the maximum tolerated dose. Secondary objectives include anti-tumor response (including circulating tumor [ct] DNA response) and pharmacokinetics. Parts A (escalation) and B (expansion) assess AZD9833 as a monotherapy, and Parts C (escalation) and D (expansion) assess AZD9833 in combination with palbociclib. Results: At a data cut-off of March 24 2020, 17 patients had received either 150 mg or 300 mg AZD9833 in combination with palbociclib, given according to its product labeling. Eighty patients had received AZD9833 monotherapy at doses of 25, 75, 150, 300, and 450 mg QD. In patients treated with AZD9833 and palbociclib, treatment-related adverse events (AEs; experienced by ≥10% of patients) included visual disturbances*, bradycardia*, asthenia, anemia, QTcF prolongation, nausea, neutropenia, decreased white blood cell count, and vomiting (*combined terms). All instances of AZD9833-related bradycardia were Grade 1. One DLT was observed in the 150 mg cohort: CTCAE Grade 2 visual disturbances, which began on Cycle 1 Day 8 and resolved by Cycle 1 Day 9 following dose interruption. The patient restarted treatment on Cycle 1 Day 15 at 75 mg and continued this dose until data cut-off. No causally related AEs led to discontinuation of AZD9833. The tolerability of AZD9833 with palbociclib was consistent with the observed tolerability profile of AZD9833 monotherapy, and the known tolerability profile of palbociclib. Pharmacokinetic analysis showed similar AZD9833 exposure for monotherapy and palbociclib combination therapy. Similarly, palbociclib exposure was comparable with simulations using a published population pharmacokinetic model. In Part A, ESR1 hotspot mutations were detected in ctDNA at baseline in 26/56 (46%) patients; 13/26 (50%) of these patients achieved a partial response or stable disease at 24 weeks, including 5/10 (50%) with a Y537S ESR1 mutation. Further, in patients with ESR1 mutations and samples available for longitudinal ctDNA analysis, 17/20 (85%) exhibited a reduction or loss of mutant ESR1 on treatment with AZD9833. Efficacy data to be presented include objective response rate and clinical benefit rate at 24 weeks. Of note, unconfirmed partial responses have been observed in Part C after the data cut-off for this abstract. Conclusions: AZD9833 continues to show an encouraging efficacy and dose-dependent safety profile as a monotherapy or in combination with palbociclib. A Phase 2 study comparing the efficacy and safety of three doses of AZD9833 versus fulvestrant (NCT04214288), and a Phase 2 pre-surgical ‘window of opportunity’ study (EUDRA-CT; 2019-003706-2) are ongoing. Citation Format: Richard Baird, Mafalda Oliveira, Eva Maria Ciruelos Gil, Manish R Patel, Begoña Bermejo de las Heras, Manuel Ruiz-Borrego, Javier García-Corbacho, Anne Armstrong, Udai Banerji, Chris Twelves, Valentina Boni, Jason Incorvati, Peter Kabos, Adam L Cohen, Bruno de Paula, Marta Capelán Rodríguez, Judy S Wang, Christina Hernando, Alejandro Falcón Gonzalez, Ivan Victoria Ruiz, Julia Lai-Kwon, Anosheh Afghani, Christos Vaklavas, Tim Brier, Steven Fox, Bistra Kirova, Teresa Klinowska, Chris Leach, Justin PO Lindemann, Richard Mather, Rhiannon Maudsley, Christopher J Morrow, Nitharsan Sathiyayogan, Andy Sykes, Li Zhang, Erika Hamilton. Updated data from SERENA-1: A Phase 1 dose escalation and expansion study of the next generation oral SERD AZD9833 as a monotherapy and in combination with palbociclib, in women with ER-positive, HER2-negative advanced breast cancer [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS11-05.

Purpose: The vast majority of advanced ER POS breast cancers eventually cease responding to endocrine (ET) and other therapies leading to evolution of lethal disease. However, timely monitoring of the molecular events associated with response/progression in tissue biopsies is logistically difficult. The use of liquid biopsies, such as CTC and ctDNA, in this context has been of recent interest. Patients and Methods: Individual CTC and ctDNA were obtained at different time points from patients with advanced ER POS/HER2 NEG breast cancer enrolled in a Phase I trial of AZD9496, an oral selective estrogen receptor degrader (SERD) ET. The CTC, purified using tandem CellSearch®/DepArray™ technologies, were genomically profiled by DNA single cell next generation sequencing (scNGS). Plasma ctDNA was isolated from blood collected in Streck BCT tubes. Genomic profiling was performed by targeted gene panel scNGS for CTC and ddPCR for ERα gene (ESR1) mutations in ctDNA. Results: 123 high-quality CTCs from 12 patients profiled by scNGS showed 100% concordance with ctDNA in detection of driver ESR1 somatic mutations. CTC scNGS additionally revealed extensive intra-patient heterogeneity of driver alterations, that would have been unresolvable by bulk ctDNA profiling, including separate subclonal CTC populations emerging within the same patient. ScNGS revealed potential opportunities for targeted therapies in 73% of patients, directed at alterations in PIK3CA, FGFR2, KIT and BRAF, at times present as 2 or more targets in the same or different cell populations. In one patient, an emergent, distinct, BRAF p.V600E targetable alteration was detected in a subpopulation of CTCs collected at the progression time point but not at baseline. Conclusion: Serial monitoring of CTC and ctDNA genomic alterations is feasible and should enable real-time tracking of response/progression, tumor evolution and opportunities for precision medicine interventions. Citation Format: Andi K. Cani, Emily M. Dolce, Elizabeth P. Darga, Kevin Hu, Martha Brown, Chia-Jen Liu, Jackie Pierce, Kieran Bradbury, Kimberly Aung, Gaia Schiavon, Danielle Carroll, T. H. Carr, Teresa Klinowska, Justin Lindemann, Gayle Marshall, Vicky Rowlands, Elizabeth A. Harrington, J. Barrett, Anne Armstrong, Richard Baird, Erika Hamilton, Seock-Ah Im, Komal Jhaveri, Manish R. Patel, Caroline Dive, Scott A. Tomlins, Aaron M. Udager, Daniel F. Hayes, Costanza Paoletti. Monitoring circulating tumor cell (CTC) and circulating tumor DNA (ctDNA) genomic alterations in ER positive (POS)/HER2 negative (NEG) advanced breast cancer during endocrine therapy: correlative study of AZD9496 oral SERD phase I trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3143.

[Image: see text] Herein we report the use of metathesis to construct a novel tetracyclic core in a series of estrogen receptor degraders. This improved the chemical stability, as assessed using an NMR-MS based assay, and gave a molecule with excellent physicochemical properties and pharmacokinetics in rat. X-ray crystallography established minimal perturbation of the bridged compounds relative to the unbridged analogues in the receptor binding pocket. Unfortunately, despite retaining excellent binding to ERα, this adversely affected the ability of the compounds to degrade the receptor.

1032 Background: SERENA-1 (NCT03616587) is a Phase 1, multi-part, open-label study of camizestrant in women with ER+, HER2− advanced breast cancer (ABC). Parts A/B (escalation/expansion) assessed camizestrant monotherapy and have been presented previously. Parts C/D examine camizestrant in combination with palbociclib; here we present mature data from camizestrant 75 mg in combination with palbociclib; 75 mg being the camizestrant dose currently under investigation in the Phase 3 studies SERENA-4 (NCT04711252) and SERENA-6 (NCT04964934). Methods: The primary objective was to determine the safety and tolerability of camizestrant once daily (QD) with palbociclib. Secondary objectives included anti-tumor response and pharmacokinetics (PK). Prior treatment with < 2 lines of chemotherapy in the advanced setting was permitted. There was no limit on the number of lines of prior endocrine treatment in the advanced setting; prior treatment with CDK4/6 inhibitors and fulvestrant (F) was permitted. Results: As of 9 September 2021, 25 patients had received camizestrant 75 mg QD in combination with palbociclib. Tolerability of the combination of camizestrant 75 mg and palbociclib was consistent with that of each drug individually. No patient required camizestrant dose interruption/reduction/discontinuation due to a camizestrant-related adverse event (AE); moreover, none experienced a Grade ≥3 camizestrant-related AE. All camizestrant-related heart rate reductions were Grade 1 (asymptomatic). All camizestrant-related visual effects were Grade 1 (mild), apart from one patient who experienced transient Grade 2 (moderate) visual effects that resolved to Grade 1 without dose modification. Camizestrant-related gastrointestinal disorders were all Grade 1, except one instance of Grade 2 nausea lasting one day. PK data for camizestrant 75 mg QD and palbociclib combined were broadly consistent with camizestrant as monotherapy and published palbociclib steady-state PK data, further supporting the use of camizestrant 75 mg QD (Phase 3 dose) in combination with the approved palbociclib doses. In these heavily pre-treated patients (48% prior chemotherapy, 80% prior CDK4/6i, 68% prior F; all in advanced disease setting) and of whom 60% had visceral metastases, the clinical benefit rate at 24 weeks was 7/25 (28%). Conclusions: The PK and safety profile of camizestrant 75 mg QD in combination with palbociclib is favorable in this mature Phase 1 dataset. Despite extensive pre-treatment - including chemotherapies, CDK4/6i, and F - camizestrant 75 mg QD in combination with palbociclib exhibits encouraging clinical activity. The results from the ongoing Phase 3 studies, SERENA-4 and SERENA-6, will further elucidate the role of this combination in the treatment of patients with HR+/HER2− ABC. Clinical trial information: NCT03616587.

Suppressive myeloid cells mediate resistance to immune checkpoint blockade. PI3Kγ inhibition can target suppressive macrophages, and enhance efficacy of immune checkpoint inhibitors. However, how PI3Kγ inhibitors function in different tumor microenvironments (TME) to activate specific immune cells is underexplored. The effect of the novel PI3Kγ inhibitor AZD3458 was assessed in preclinical models. AZD3458 enhanced antitumor activity of immune checkpoint inhibitors in 4T1, CT26, and MC38 syngeneic models, increasing CD8+ T-cell activation status. Immune and TME biomarker analysis of MC38 tumors revealed that AZD3458 monotherapy or combination treatment did not repolarize the phenotype of tumor-associated macrophage cells but induced gene signatures associated with LPS and type II INF activation. The activation biomarkers were present across tumor macrophages that appear phenotypically heterogenous. AZD3458 alone or in combination with PD-1–blocking antibodies promoted an increase in antigen-presenting (MHCII+) and cytotoxic (iNOS+)-activated macrophages, as well as dendritic cell activation. AZD3458 reduced IL-10 secretion and signaling in primary human macrophages and murine tumor-associated macrophages, but did not strongly regulate IL-12 as observed in other studies. Therefore, rather than polarizing tumor macrophages, PI3Kγ inhibition with AZD3458 promotes a cytotoxic switch of macrophages into antigen-presenting activated macrophages, resulting in CD8 T-cell–mediated antitumor activity with immune checkpoint inhibitors associated with tumor and peripheral immune activation.

Herein we report the optimization of a series of tricyclic indazoles as selective estrogen receptor degraders (SERD) and antagonists for the treatment of ER+ breast cancer. Structure based design together with systematic investigation of each region of the molecular architecture led to the identification of N-[1-(3-fluoropropyl)azetidin-3-yl]-6-[(6S,8R)-8-methyl-7-(2,2,2-trifluoroethyl)-6,7,8,9-tetrahydro-3H-pyrazolo[4,3-f]isoquinolin-6-yl]pyridin-3-amine (28). This compound was demonstrated to be a highly potent SERD that showed a pharmacological profile comparable to fulvestrant in its ability to degrade ERα in both MCF-7 and CAMA-1 cell lines. A stringent control of lipophilicity ensured that 28 had favorable physicochemical and preclinical pharmacokinetic properties for oral administration. This, combined with demonstration of potent in vivo activity in mouse xenograft models, resulted in progression of this compound, also known as AZD9833, into clinical trials.

Purpose:Fulvestrant, the first-in-class selective estrogen receptor (ER) degrader (SERD), is clinically effective in patients with ER+ breast cancer, but it has administration and pharmacokinetic limitations. Pharmacodynamic data suggest complete ER degradation is not achieved at fulvestrant's clinically feasible dose. This presurgical study (NCT03236974) compared the pharmacodynamic effects of fulvestrant with AZD9496, a novel, orally bioavailable, nonsteroidal, potent SERD, in treatment-naïve patients with ER+ HER2− primary breast cancer awaiting curative intent surgery.Patients and Methods:Patients were randomized 1:1 to receive AZD9496 250 mg twice daily from day 1 for 5–14 days, or fulvestrant 500 mg on day 1. On-treatment imaging-guided core tumor biopsies were taken between day 5 and 14 and compared with pretreatment diagnostic biopsies. The primary objective was to compare the effects of AZD9496 and fulvestrant on ER expression. Secondary objectives included changes in progesterone receptor (PR) and Ki-67 pharmacokinetic/pharmacodynamic relationships and safety.Results:Forty-six women received treatment (AZD9496 n = 22; fulvestrant n = 24); 35 paired biopsies were evaluable (AZD9496 n = 15; fulvestrant n = 20). The least square mean estimate for ER H-score reduction was 24% after AZD9496 versus 36% after fulvestrant treatment (P = 0.86). AZD9496 also reduced PR H-scores (−33.3%) and Ki-67 levels (−39.9%) from baseline, but was also not superior to fulvestrant (PR: −68.7%, P = 0.97; Ki-67: −75.4%, P = 0.98). No new safety findings were identified.Conclusions:This was the first presurgical study to demonstrate that an oral SERD affects its key biological targets. However, AZD9496 was not superior to fulvestrant at the dose tested.

Loss of the tumor suppressor PTEN confers a tumor cell dependency on the PI3Kβ isoform. Achieving maximal inhibition of tumor growth through PI3K pathway inhibition requires sustained inhibition of PI3K signaling; however, efficacy is often limited by suboptimal inhibition or reactivation of the pathway. To select combinations that deliver comprehensive suppression of PI3K signaling in PTEN-null tumors, the PI3Kβ inhibitor AZD8186 was combined with inhibitors of kinases implicated in pathway reactivation in an extended cell proliferation assay. Inhibiting PI3Kβ and mTOR gave the most effective antiproliferative effects across a panel of PTEN-null tumor cell lines. The combination of AZD8186 and the mTOR inhibitor vistusertib was also effective in vivo controlling growth of PTEN-null tumor models of TNBC, prostate, and renal cancers. In vitro, the combination resulted in increased suppression of pNDRG1, p4EBP1, as well as HMGCS1 with reduced pNDRG1 and p4EBP1 more closely associated with effective suppression of proliferation. In vivo biomarker analysis revealed that the monotherapy and combination treatment consistently reduced similar biomarkers, while combination increased nuclear translocation of the transcription factor FOXO3 and reduction in glucose uptake. These data suggest that combining the PI3Kβ inhibitor AZD8186 and vistusertib has potential to be an effective combination treatment for PTEN-null tumors. Mol Cancer Ther; 17(11); 2309–19. ©2018 AACR.

Fulvestrant, the first-in-class selective estrogen receptor (ER) degrader (SERD), is clinically effective in patients with ERPatients were randomized 1:1 to receive AZD9496 250 mg twice daily from day 1 for 5-14 days, or fulvestrant 500 mg on day 1. On-treatment imaging-guided core tumor biopsies were taken between day 5 and 14 and compared with pretreatment diagnostic biopsies. The primary objective was to compare the effects of AZD9496 and fulvestrant on ER expression. Secondary objectives included changes in progesterone receptor (PR) and Ki-67 pharmacokinetic/pharmacodynamic relationships and safety.Forty-six women received treatment (AZD9496This was the first presurgical study to demonstrate that an oral SERD affects its key biological targets. However, AZD9496 was not superior to fulvestrant at the dose tested.