AZD0466

While the BRAFV600E inhibitor encorafenib in combination with the EGFR inhibitor cetuximab (Enc+Cet) was recently approved for this indication, overall survival is only increased by 3.6 months and objective responses are observed in only 20% of patients. Combining encorafenib with DT2216 significantly increased apoptosis induction in vitro, while combining encorafenib with AZD0466 was well tolerated in mice and further reduced growth of BRAFV600E CRC xenografts compared to either agent alone. Collectively, these findings demonstrate that combined BRAF and BCL-XL inhibition significantly enhances apoptosis in pre-clinical models of BRAFV600E CRC and is a combination regimen worthy of clinical investigation to improve outcomes for these patients.

BCL-2 inhibition by venetoclax (VEN) is beneficial in many patients (pts) with acute myeloid leukemia (AML), but resistance is common due to upregulation of other pro-survival proteins such as BCL-extra-large (BCL-xL) and myeloid cell leukemia-1...Preclinically, AZD4320 showed greater inhibition of tumor growth than VEN and navitoclax in VEN-resistant xenograft models (Balachander et al... Given the observed DLTs of asymptomatic increase in troponin levels, and the lack of significant clinical benefit, the study was terminated.

Venetoclax is a selective BCL-2 inhibitor and is successfully used in CLL and AML, but heterogeneous sensitivity to venetoclax has been described in ALL and inhibitors of other BCL-2 family members including BCL-XL-selective A-1331852 and MCL-1 selective AZD5991 have been developed. Moreover, a dual inhibitor of the anti-apoptotic BCL-2 family members BCL-2 and BCL-XL (AZD4320) with its dendrimer conjugate (AZD0466) has recently shown anti-tumor activity in hematologic cancer models with manageable toxicity...Taken together, we found vulnerabilities of T-ALL to BCL-2 family inhibition, particularly to dual BCL-2/BCL-XL inhibition by AZD4320, and we demonstrated on-target activities. Using BH3-profiling, we identified BAD-priming as a marker of response for AZD4320 and MCL-1 dependence as a resistance mechanism that can be targeted by combination treatment, suggesting further clinical evaluation.

To evaluate if combination treatment of AZD0466 with BTK inhibitors would improve efficacy, we transplanted murine Eµ-TCL1 tumors into syngeneic recipient mice and randomized them for treatment with vehicle, ibrutinib (30mg/kg in drinking water), acalabrutinib (25mg/kg, p.o. QD), AZD0466 (70mg/kg, i.v., QW) and combination of AZD0466 with ibrutinib or acalabrutinib. Moreover, AZD4320 was highly efficacious in MAVER-1 and MINO cell line models where resistance to venetoclax mediated by BCL-XL upregulation was modelled by an in vitro dose escalation method. In summary, our pre-clinical study shows that the dual BCL2/BCL-XL inhibitor could represent an important treatment option for venetoclax resistance mediated by specific BCL2 mutations or BCL-XL upregulation and that its efficacy could be further improved upon combination treatment with BTKi.

P1/2, N=7, Terminated, AstraZeneca | N=50 --> 7 | Trial completion date: May 2025 --> Aug 2023 | Recruiting --> Terminated | Trial primary completion date: Oct 2024 --> Aug 2023; Due to safety reasons.

Among 52 patients with hematological malignancies the maximum tolerated dose (MTD) was not reached and no clinical TLS was observed.20 In this study of 22 evaluable patients with RR CLL there was an overall response rate (ORR) of 63.6% (14/22).20 A larger study of 114 patients with RR CLL evaluated lisaftoclax in combination with either the Bruton's tyrosine kinase inhibitor (BTKi) acalabrutinib or the anti CD20 monoclonal antibody rituximab (NCT04215809)...In CLL an overall response rate (ORR) to BGB-1147 of 100% as monotherapy was observed among 8 patients with RR disease.25 In 12 patients with RR MM, the ORR to BGB-1147 in combination with dexamethasone varied from 0–68% depending on the dose cohort.26 In combination with azacytidine in AML the ORR to BGB-11417 was 74% among those with treatment naïve (TN) disease (n=20/27) and 65% (n=13/20) in the cohort with RR disease.27 BGB-11417 monotherapy in 23 patients with RR NHL demonstrated responses in 2 patients with diffuse large B cell lymphoma (DLBCL) and one patient with marginal zone lymphoma (MZL).28 In 11 patients with RR MCL treated with BGB-11417 in combination with zanubrutinib there was a 55% (6/11) ORR.28 S55746 is a potent BAX/BAK dependent BCL2 inhibitor administered orally.18,29 Developed by Servier, it has been tested in phase I studies in CLL, NHL and AML. In a study of S55746 among 37 patients with RR NHL, no dose limiting toxicities (DLT) or TLS was observed after a medium duration of treatment of 42 days.30 FCN-338 is another orally available selective BCL2 inhibitor developed by Fochon currently undergoing phase I testing in RR CLL.18,31 Clinical outcomes with this drug are yet to be publicly reported...More recently navitoclax has been tested in RR myelofibrosis (MF) in combination with ruxolinitib with evidence of clinical response to the combination.35 AZD0466 by Astrazeneca is a nanomedicine potent dual BCL2/ BCLxL inhibitor that mediates BAX/BAK induced apoptosis36,37 and is administered intravenously...Among 9 patients reported undergoing testing for RR hematological malignancy the DLT had not yet been reached.38 Pelcitoclax or APG1252 by Ascentage is a more recent BAX/BAK dependent dual BCL2 and BCLxL inhibitor18,39,40 currently in phase I trials in RR NHL (NCT05186012)...There are multiple emerging BCL2 inhibitors currently undergoing clinical trial testing in hematological malignancies, and it remains too early to appreciate the differential efficacy and toxicity profiles that these agents may carry compared with venetoclax. It is hoped that the results seen with venetoclax can be improved upon across a raft of disease groups over the coming years.

Background: The BCL2 family of proteins induce apoptosis via caspase activation and are being increasingly targeted in hematologic malignancies by small molecules such as venetoclax (VEN)...Preclinically, AZD4320 showed activity in patient (pt)-derived AML xenografts and superior tumor growth inhibition to VEN and navitoclax in VEN-resistant xenograft models (Balachander et al...Module 2 investigates drug-drug interactions between AZD0466 and voriconazole... Treatment with AZD0466 is well tolerated in pts with R/R acute leukemia, with AEs matching expected toxicity from preclinical data. Preclinical efficacy modeling and clinical PK data suggest further dose escalation is warranted to explore clinical activity.

The active moiety, AZD4320, is a potent dual inhibitor of BCL-2 and BCL-XL...AZD0466 outperformed the selective BCL-2 inhibitor venetoclax in 6/10 models. Notably, AZD0466 was active in models resistant to platinum/etoposide chemotherapy, the standard-of-care for SCLC...AZD0466 exhibits linear PK, consistent across solid tumor and leukemia patients. The doses tested are in line with preclinical studies in SCLC.

P1/2, N=50, Recruiting, AstraZeneca | Trial primary completion date: May 2025 --> Oct 2024

P1/2, N=50, Recruiting, AstraZeneca | Trial completion date: Nov 2024 --> May 2025 | Trial primary completion date: Nov 2024 --> May 2025

Specific Bcl-2 inhibition using venetoclax is beneficial in patients with acute myeloid leukemia (AML), but resistance often develops due to upregulation of anti-apoptotic proteins such as Bcl-xL and Mcl-1...Following IV infusion, released AZD4320 exhibited a dose-proportional increase in area under the concentration time curve (AUC) and maximum serum concentration (Cmax), moderate PK variability (≈50% coefficient of variation), and a half-life of ≈18 hours. AZD0466 has been well tolerated, with no DLTs to date and no discontinuations due to treatment-related AEs. The trial continues to enroll, further dose escalation is planned, and updated data will be presented.

Here we examine the impact of SC dosing on lymphatic exposure, pharmacokinetics (PK), and efficacy of AZD0466, a small molecule dual Bcl-2/Bcl-xL inhibitor conjugated to a 'DEP®' G5 poly-l-lysine dendrimer...SC administration improves lymphatic exposure and facilitates higher dose administration due to improved tolerability. Higher dose SC administration also results in improved efficacy, suggesting that drug delivery systems that access both plasma and lymph hold significant potential for the treatment of haematological cancers where lymphatic and extranodal dissemination are poor prognostic factors.

P1/2, N=50, Recruiting, AstraZeneca | Not yet recruiting --> Recruiting | Initiation date: Feb 2022 --> Jul 2022

AZD4320 was developed as a dual inhibitor of BCL-2 and BCL-XL, and its dendrimer conjugate (AZD0466) was recently reported to demonstrate anti-tumor activity in hematological cancer models, while showing only a transient thrombocytopenia. Aims In this study, the anti-leukemia activities of the dual BCL-2 and BCL-XL inhibitor AZD4320 and of MCL-1-selective AZD5991 were evaluated and compared to the effects of other BH3-mimetics (BCL-2-selective venetoclax, BCL-XL-selective A-1331852 and MCL-1-selective S63845)...Importantly, the highest synergism was found at low concentrations of both inhibitors, suggesting efficacy at moderate concentrations, which could potentially be achieved in vivo . Conclusion In summary, our study demonstrates sensitivity, on-target activity and synergism of the dual BCL-2 and BCL-XL inhibitor AZD4320 with inhibition of MCL-1, thereby providing strong evidence for further clinical evaluation in ALL.

P1/2, N=50, Not yet recruiting, AstraZeneca

Dual BCL2/xL inhibition with AZD4320 has potential for broader activity than BCL2-specific inhibition with venetoclax (Balachander et al, Clinical Cancer Research 2020)...Treatment with hydroxyurea during screening and cycle 1 is permitted to control white blood cell count...Module 2 is a drug-drug interaction (DDI) study that will investigate the safety and establish the sensitivity of AZD0466 to voriconazole, a strong inhibitor of CYP3A4...This study is actively enrolling patients at sites in the USA, and will be opening at multiple sites in Australia, Europe and South Korea. An update of preliminary safety and pharmacokinetics will be presented.

This is largely due to frequent relapses after therapies including paradigm shifting therapies BTK inhibitors (BTKi), such as ibrutinib and acalabrutinib, and Bcl-2 inhibitor (Bcl-2i) venetoclax after long-term treatment in the clinic. Conclusion Compared to AZD4320/AZD0466 and acalabrutinib, combination therapy demonstrated anti-MCL synergy both in vitro and in vivo . These findings suggest that targeting Bcl-2/X L and BTK is promising to overcome multiple acquired resistance phenotypes, including CD19 CAR T-cell therapy.

However, 10–50% of newly diagnosed patients with AML may not respond to venetoclax and HMA or LDAC, and 3–15% patients may not respond to venetoclax with intensive or non-intensive chemotherapy.1–6 In addition, up to 40% of responding patients may relapse with low rates of response of 20% to salvage therapy and poor overall survival of 2 months after relapse.7 Clinical and biological factors associated with primary and acquired resistance to venetoclax include secondary AML, monocytic differentiation, complex cytogenetics, mutations in TP53, BAX, dependence on other anti- apoptotic proteins, altered metabolism of nicotinamide, fatty acids, and oxidative phosphortylation.3,8–14 Several novel inhibitors of BCL-2 are currently being tested in clinic, including BGB 11417, APG-2575, LP-108 and others...There is strong pre-clinical rationale for targeting MCL-1 alone as well as in conjunction with BCL-2 inhibition in AML.15 Recently several selective and highly potent MCL-1 inhibitors have entered pre-clinical and clinical development including S63845, AZD5991, AMG397, and others. Questions remain regarding the therapeutic window of these inhibitors given the important physiologic role of MCL-1 in vital organs and early reports of cardiac adverse events from the AMG176 phase 1 trial.15,16 Multiple pre-clinical studies have expectedly shown synergism between BCL- 2 and MCL-1 inhibition making it a promising path for clinical development of these agents.17,18 Multifactorial challenges in design of specific MCL-1 inhibitors also led to interest in compounds which downregulate MCL-1 expression. Cyclin dependent kinase (CDK) inhibitors including alvocidib, dinaciclib, voruciclib are in various stages of evaluation. Although addition of alvocidib to intensive chemotherapy improved response rates but failed to improve event-free or overall survival.19 Novel CDK inhibitors are currently in early phase trials including AZD4573, CYC065, TG02-101, and others. Inhibition of Nedd8 activating enzyme has complex repercussions for the intrinsic apoptotic pathway with eventual increase in Noxa leading to MCL-1 neutralization.20 Pevonedistat has shown promising early results in AML and myelodysplastic syndrome and is being investigated multiple clinical trials for solid tumors as well. BCL-xL Inhibition Another anti-apoptotic protein BCL-xL had been long recognized as a potential therapeutic target in AML, in particular AML from preceding MPN and AML recurrent post venetoclax failure, but toxicity of earlier inhibitors precluded clinical development.21–23 Recently, AZD0466, a dual BCL-2/xL inhibitor with a favorable therapeutic index and robust activity has been developed and is undergoing pre-clinical development and planned for phase iin hematological malignancies.24 Targeting the Extrinsic Apoptosis Pathway Inhibitor of apoptosis protein (IAP) inhibition: X-linked IAP (XIAP), cellular IAP (cIAP) and survivin have been of long- standing interest in AML. Prior clinical trials with XIAP inhibitor AEG35156, cIAP targeting agent birinapant, and survivin targeting agent LY2181308 have not succeeded in clinc.16,25 ASTX660 is a dual antagonist of XIAP and cIAP which is currently being investigated in phase 1/2 trials in solid tumors and in combination with HMA in relapsed or refractory AML.26,27 TRAIL Agonism Agonists of the TNF-related apoptosis-inducing ligand (TRAIL) receptors have been tested in AML with low response rates.28,29 Previous agents have had limited success in part due to suboptimal clustering of TRAIL receptors.30 Novel antibodies against TRAILR1 and TRAILR2 including an IgM molecule IGM-8444, a tetravalent compound INBRX-109, and HLX56 are currently in phase 1 trials and preclinical data suggests potential synergy with venetoclax.31 FLIP Inhibition FLICE-like inhibitor protein (FLIP or CFLAR) is a key regulator of the death-inducing signaling complex (DISC) involved in the extrinsic apoptotic pathway...This can be augmented by inhibiting p53 degradation via MDM2, which is often upregulated in AML.34 Idasanutlin in combination with venetoclax showed anti- Figure 1 leukemic activity in the dose finding stage in R/R AML.35 Several other inhibitors of MDM2 and dual MDM2/X inhibitors are currently in various stages of pre-clinical and clinical development including HDM-201, KRT-232, BI-9078282, and others.34 Conclusions Opportunities to target the apoptosis machinery in AML has considerably evolved in the last decade. While venetoclax heralded a paradigm shift for patients, we are now faced with challenges in patients who relapse or remain refractory. We have novel clinical stage compounds to methodologically target different facets of the apoptotic pathway and optimize novel combinations with the goal to improve the cure rates in AML patients.

Malignant pleural mesothelioma (MPM) is an aggressive cancer with treatment limited to Cisplatin and Pemetrexed chemotherapy...In this study we show another inhibitor, AZD4320 that targets BCL-XL (and BCL-2), can also potently kill MPM tumor cells in vitro (EC values in the 200 nM range) and this effect is enhanced by co-inhibition of MCL-1 using AZD5991...Critically, the degree of thrombocytopenia, an on-target toxicity associated with BCL-XL inhibition, was significantly reduced throughout the treatment period compared to other BCL-XL-targeting BH3-mimetics. These pre-clinical findings provide a rationale for the future clinical evaluation for novel BH3-mimetic formulations in MPM, and indeed, other solid tumor types dependent on BCL-XL.

The optimised candidate is shown to be efficacious and better tolerated in preclinical models compared with AZD4320 alone. The AZD4320-dendrimer conjugate (AZD0466) identified, through mathematical modelling, has resulted in an improved therapeutic index and thus enabled progression of this promising dual Bcl-2/Bcl-x inhibitor into clinical development.

AZD0466, the nanomedicine formulation of AZD4320 (30mg/kg, weekly, IV), dramatically inhibited tumor growth and prolonged mouse survival in an ibrutinib-CAR-T dual-resistant PDX mouse model. The novel BCL-2/XL dual inhibitor AZD4320 demonstrated excellent anti-MCL activity in both ibrutinib/venetoclax-sensitive and -resistant MCL cells in vitro. This was further validated in vivo in a ibrutinib-CAR-T dual-resistant PDX model. These findings provide evidence that dual targeting of BCL-2 and BCL-XL by AZD4320 is promising as it may overcome therapeutic resistance in relapsed/refractory MCL.

AZD0466 is a novel drug-dendrimer conjugate, where the active moiety, AZD4320, is chemically conjugated to Starpharma’s DEP® dendrimer platform, a 5-generation PEGylated poly-lysine dendrimer via a hydrolytically labile linker...AZD0466 demonstrated greater monotherapy activity than platinum/etoposide chemotherapy regimen or venetoclax monotherapy in 6 out of 11 SCLC PDX models...In the SUDHL-4 GCB DLBCL model, 103 mg/kg AZD0466 with 10 mg/kg Rituximab resulted in complete and durable regressions in 5/6 animals. Finally, combination of 103 mg/kg AZD0466 with 12.5 mg/kg BID Acalabrutinib, a Bruton’s Tyrosine kinase inhibitor, was investigated in OCI-LY10 DLBCL model...These data show that AZD0466 has monotherapy activity and a differentiated response from Venetoclax in SCLC models. AZD0466 also has therapeutic potential as monotherapy and a combinatorial agent to increase the depth and duration of response to standard of care and BTK inhibitors in hematological tumors.

In this study, we tested BCL-2 and BCL-XL dependencies among different subtypes of ALL and the potential to revert these using AZD0466, a novel drug-dendrimer conjugate, of a BCL-2/ BCL-XL inhibitor AZD4320 and an FDA-approved BCL-2 inhibitor, ABT-199 (Venetoclax). Our findings suggest that the novel dual BCL-2/BCL-XL inhibitor AZD0466 outperforms single BCL-2 inhibition by ABT-199 in T-ALL and Ph-like B-ALL. These findings will facilitate translation of AZD0466 dual BCL-2/BCL-XL inhibitor into ALL clinical trials, alone or in combination with standard chemotherapy and monoclonal antibodies.

DEP irinotecan also showed significant benefit when dosed in combination with current standard of care therapies such as the anti-EGFR antibody, cetuximab, and the PARP inhibitor, olaparib...The fourth DEP candidate, AZD0466 is a promising Bcl-2/Bcl-xL inhibitor and is partnered with AstraZeneca...The dendrimer nanoparticle shows significant efficacy and survival benefits compared to current standard of care therapies when used either as a monotherapy or in combination. DEP irinotecan is currently in a phase 1 / 2 adaptive trial at leading UK hospitals; The Christie, The Royal Marsden and The Newcastle Upon Tyne.