tolinapant (ASTX660)

P1, N=33, Not yet recruiting, National Cancer Institute (NCI)

P1/2, N=127, Not yet recruiting, National Cancer Institute (NCI)

P1/2, N=132, Active, not recruiting, Astex Pharmaceuticals, Inc. | Recruiting --> Active, not recruiting

P1, N=10, Recruiting, Emory University | Trial primary completion date: Oct 2024 --> Oct 2025

P1, N=78, Recruiting, Gustave Roussy, Cancer Campus, Grand Paris | Not yet recruiting --> Recruiting

Our data offer a cellular and molecular insight into a case of clinical pseudoprogression observed during tolinapant treatment. Understanding pseudoprogression is important for clinical investigators and patients to ensure treatment with tolinapant, or other IAP antagonists, are not discontinued prematurely.

IAP antagonists have shown great promise for head and neck cancer, especially in combination with radiation therapy. Here, we review recent preclinical and clinical studies on the use of these novel targeted agents for head and neck cancer.

P1b, N=78, Not yet recruiting, Gustave Roussy, Cancer Campus, Grand Paris

Therefore, future work will explore these pathways under hypoxic conditions. Additionally changes in cell proliferation and protein expression of IAPs will be measured following exposure of cervical cancer cell lines to different hypoxic conditions in combination with irradiation, cisplatin and tolinapant.

While there are limited studies using decitabine as a hypomethylating agent (HMA) in PTCL, a recent guadecitabine prospective study showed 40% ORR (Wong et al., Leukemia, 2022)...Subjects with CD30-positive disease must have received or be ineligible for brentuximab vedotin... The Study opened in June 2022 with primary completion estimated to be December 2025. Pharmacokinetic, Oral, Peripheral T-cell lymphoma

Subjects with CD30-positive disease must have received, be ineligible for, or intolerant to brentuximab vedotin. In Phase 2, there will be efficacy analysis for every 34 subjects, without a pause in enrollment. Study initiated in May 2022.

In addition, re-expression of RIPK3 in CT26 cells increased lytic cell death on treatment with tolinapant indicating its essential role in the necroptosis pathway.Objectives: 1) Characterize the single-agent activity of tolinapant in a range of TCL lines; 2) Determine the synergy of tolinapant in combination with drugs active against PTCL (romidepsin, pralatrexate) and HMAs (azacytidine-AZA; decitabine-DAC), and 3) Define the role of necroptosis in the mechanism of synergy. Thus, activation of the necroptosis pathway, is a possible mechanism for the high degree of synergy displayed when HMAs are used in combination with tolinapant in the TCL lines in vitro. These data provided the rationale for a phase 1-2 study of the combination of tolinapant and oral decitabine/cedazuridine treatment in relapsed/refractory PTCL (NCT05403450).

These results support the continued development of tolinapant for the treatment of R/R PTCL and CTCL. A drug combination study using tolinapant in R/R PTCL is being developed.

Aims We explored the sensitivity of a range of TCL lines to tolinapant, established the synergy coefficient and interrogated the role of necroptosis in the mechanism of synergy between tolinapant and drugs active against PTCL, the HMAs; azacytidine and decitabine Methods A panel of 10 human TCL lines were tested in proliferation assays (CellTiterGlo) for sensitivity to tolinapant in the presence or absence of 10 ng/ml of TNFa. Thus, activation of the necroptosis pathway, is a possible mechanism for the high degree of synergism displayed when HMAs are used in combination with tolinapant in the TCL lines in vitro . These data provide the rationale to initiate clinical trials with the combination of tolinapant and decitabine.

Methods On-target effects of decitabine and tolinapant were measured by analysing levels of DNMT1 and cIAP1, respectively, by Western blotting in mouse and human cell lines. In addition, modulation of cytokine response and cancer/testis antigen expression could enhance anti-tumour response. These findings provide a strong rationale to explore this combination clinically.

Our study highlights that ASTX660 can overcome the limitation of apoptosis induction via triggering necroptosis in BC cells. Therefore, our findings may provide some important clues for the design of a novel treatment strategy for BC.

Tolinapant synergizes with the anti-leukemic activity of ABT199 and DEX, establishing a therapeutic rationale for IAP antagonist in T-ALL.

Tolinapant has demonstrated potent cytotoxic effects against a broad range of TCL lines both as a monotherapy and in combination with the HDAC Inhibitor, romidepsin. In in vitro studies, T cell lymphoma cell lines demonstrated varying sensitivity to tolinapant with certain cell lines being more resistant, even in the presence of TNF alpha. Interestingly, the addition of romidepsin appeared to overcome the intrinsic resistance to tolinapant in the absence of TNF alpha.

A Phase 1 clinical study investigating the combination of tolinapant and ASTX727 (oral decitabine) in AML is currently in progress (NCT04155580). Conclusion : These data demonstrate that decitabine enhances immunogenic cell death induced by tolinapant through the re-expression of genes in the necroptotic pathway. This finding provides strong rationale to explore this combination clinically.

Mechanistically, we show that tolinapant can activate both the adaptive and the innate arms of the immune system through the induction of immunogenic forms of cell death. In summary, we describe a novel role for IAP antagonists as immunomodulatory molecules capable of promoting a robust anti-tumor immune response in TCL.

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.

We found that elevated expression of cIAP1 and cIAP2 (but not XIAP) significantly correlated with poor prognosis in patients with microsatellite stable (MSS) stage III colorectal cancer treated with 5-fluorouracil (5FU)-based adjuvant chemotherapy, suggesting their involvement in promoting chemoresistance...Moreover, the effects of FOLFOX could be phenocopied using the clinically relevant class I HDAC inhibitor, entinostat, which also induced acetylation of Ku70 and FLIP downregulation. Further analyses revealed that caspase-8 knockout RIPK3-positive colorectal cancer models were sensitive to tolinapant-induced necroptosis, an effect that could be exploited in caspase-8-proficient models using the clinically relevant caspase inhibitor emricasan. Our study provides evidence for immediate clinical exploration of tolinapant in combination with FOLFOX in poor prognosis MSS colorectal cancer with elevated cIAP1/2 expression.

Here, we demonstrate that ASTX660 has cell death promoting activity in colorectal cancer and provide a head-to-head comparison with birinapant, the clinically most advanced peptidomimetic IAP antagonist. Failure to activate the mitochondria-associated (intrinsic) apoptosis pathway attenuated the apoptosis-promoting effect of ASTX660. Further clinical studies are warranted to highlight the therapeutic potential of ASTX660 in colorectal cancer.

IAP1/XIAP antagonist ASTX660 sensitizes HPV(+) HNSCC to TNFα via a mechanism involving restoration of TP53. The present findings serve to motivate further studies of dual cIAP/XIAP antagonists and future clinical trials combining these antagonists with radiotherapy to treat both HPV(+) and (-) HNSCC.

The combination of the direct effects of ASTX660 on tumor cells and ASTX660 dependent effects on immune responses adds to the description of ASTX660’s mode of action and our ongoing understanding of the clinical efficacy in T cell Lymphoma(1).

Biomarker evaluation from this model indicated a potent immunogenic/necroptotic response after ASTX660 dosing and upregulation of immune effector cells. Conclusion The combination of the direct effects of ASTX660 on tumor cells and ASTX660 dependent effects on immune responses adds to the description of ASTX660’s mode of action and our ongoing understanding of the clinical efficacy in T cell Lymphoma (2).

Also, TNFα-blocking antibody Enbrel had little protective effect on SGI-110/ASTX660-induced cell death. Together, these events culminated in the activation of caspases-3/-7, nuclear fragmentation, and cell death. In conclusion, SGI-110 and ASTX660 cooperatively induced apoptosis in AML cells by engaging extrinsic and intrinsic apoptosis pathways, highlighting the therapeutic potential of this combination for AML.