Kazia Therapeutics (Australia)

Abstract Background Breast cancer is the most commonly diagnosed cancer globally and remains the leading cause of cancer-related death in women. HER2-positive breast cancer, comprising 15-20% of cases, is defined by overexpression of the HER2 tyrosine kinase receptor. While HER2-targeted therapies have transformed clinical outcomes, resistance, recurrence, and metastasis continue to pose significant challenges. Immunotherapy has revolutionised the treatment of several solid tumors by enhancing anti-tumor immunity; however its efficacy in HER2-positive breast cancer has been limited. There is a pressing need for biomarkers that can both track metastatic burden and identify patients most likely to benefit from immunotherapeutic strategies. In this study, we present novel preclinical data from liquid biopsy analyses and propose a biomarker-driven approach targeting the PI3K-mTOR pathway to enhance immune reinvigoration in HER2-positive metastatic breast cancer. Methods Twenty five stage IV HER2-positive metastatic breast cancer (mBC) patients underwent liquid biopsy profiling using an epigenetic biomarker targeting the PI3K-mTOR signaling pathway. We applied digital pathology, Duolink® proximity ligation assays, global immune profiling, and flow cytometry to characterize biomarker expression and immune landscape features. Ex vivo drug screening with PI3K-mTOR inhibitors, including the brain-penetrant paxalisib, was performed to evaluate their effects on mesenchymal phenotypes, disruption of circulating tumor cell (CTC) clusters, and immune reinvigoration. Results We identified a novel epigenetic switch that drives the transition to a more aggressive, drug-resistant mesenchymal phenotype in HER2-positive breast cancer, associated with activation of the nuclear chromatinised PI3K signaling pathway. This phenotype, detectable via liquid biopsy, correlates with metastatic burden and immune exhaustion. In our pilot cohort, we evaluated the interplay between PI3K mutational burden and activation of this mesenchymal epigenetic program. Functional drug response profiling of patient-derived samples demonstrated that dual PI3K-mTOR inhibition not only facilitated mesenchymal-to-epithelial transition but also suppressed key metastatic signatures, including those linked to the highly aggressive persister cell phenotype (p65, FOXQ1, NRF2, and NNMT) and cancer drug resistance (ABCB5, SNAIL, and ALDH1). Importantly, PI3K-mTOR inhibition disrupted metastatic progenitors, including circulating tumor cell (CTC) clusters, and enhanced anti-tumor immune responses, as evidenced by modulation of both progenitor-like and terminally exhausted T cell populations. Conclusions Our preclinical findings support a biomarker-driven therapeutic strategy targeting the PI3K-mTOR pathway to induce epigenetic reprogramming and immune modulation. This dual mechanism may enhance anti-tumor efficacy by converting immunologically 'cold' tumors into 'hot', more immunogenic tumors. By reshaping the tumor microenvironment, this approach has the potential to improve the clinical utility of immunotherapy and expand treatment options for patients with HER2-positive metastatic breast cancer. Citation Format: M. Melino, W. Tu, S. Goh, A. Bain, J. Friend, T. Prasanna, D. Yip, A. Ives, M. Leahy, D. Lewis, M. Nottage, S. Rao. Liquid Biopsy Tracking of PI3K-mTOR Residual Disease Signatures in Metastatic Breast Cancer [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS2-10-02.

Abstract INTRODUCTION Partial or whole brain RT is a standard treatment for solid tumor BM and LM. However, tumors harboring PI3K pathway mutations have poorer CNS control after RT alone. Combining RT with paxalisib, a CNS-penetrant PI3K/mTOR inhibitor, may overcome radioresistance. METHODS This multi-center, phase I trial (NCT04192981) studied concurrent paxalisib with cranial RT (30Gy in 10 fractions) for PI3K pathway-altered BM/LM. Part A was a 3 + 3 dose escalation cohort of paxalisib at 45, 60 or 75mg to establish maximum tolerated dose (MTD). Part B was an expansion at MTD to further validate safety and early efficacy. CNS response was assessed using Response Assessment in Neuro-Oncology Brain Metastases (RANO-BM) criteria. RESULTS From 3/2020-2/2024, 21 patients (median age 58y, n=18 with BM, n=10 with breast cancer) were enrolled. PIK3CA mutations were the most common pathway alteration (n=15, 71%) followed by PTEN deletion (n=3, 14%). Part A enrolled 12 patients of which 9 were evaluable (3 did not complete protocol therapy). There were no dose-limiting toxicities (DLTs) at 45mg, and 2 DLTs at 60mg: grade 3 nausea and grade 4 neutropenic enterocolitis. Subsequently, 9 additional patients were enrolled during Part B at MTD of 45mg, of which 1 withdrew consent prior to completion of study interventions. No additional DLTs were observed during Part B. In total, there were 14 evaluable patients who received 45mg/day paxalisib concurrently with RT, two of whom died of intercurrent extracranial progression prior to CNS restaging. Of the remaining 12, there was robust response signal. At 1-month post-treatment, 6/12 (50%) had partial response (PR) and the remaining had stable disease (SD). To date, best CNS response is 67% PR (8/12) and 33% SD (4/12). CONCLUSIONS A MTD of 45 mg/day has been confirmed for paxalisib with concurrent brain RT for PI3K pathway-altered BM/LM. Given significant unmet need, this novel therapeutic combination warrants further investigation given promising activity signal.

Abstract Background: EVT801 is a highly selective, orally available VEGFR3 inhibitor that modulates tumor angiogenesis without inducing hypoxia, one of the main causes of cancer-associated immunosuppression. EVT801 has shown compelling efficacy as single agent in multiple in vivo models. In addition, combination of EVT801 and Immune Checkpoint Therapy (ICT) agents shows additive effects and thus holds promises for combination treatment without induction of hypoxia-induced immunosuppression. Clinical trial design: A phase I clinical study (NCT05114668) is underway to evaluate safety, tolerability, pharmacokinetics of EVT801 and explore patient characterization which is key for further development. The phase I trial consists in two sequential stages. During the first stage, currently ongoing, EVT801 is orally administered to patients with advanced solid tumors in a multiple ascending dose study using an accelerated 3+3 design (1 patient per dose until grade 2 toxicities are observed) in up to 48 patients in 8 dose levels. The primary objective is to determine the maximum tolerated dose (MTD) and recommended phase 2 dose (RP2D). Stage 2 will focus on validation of this RP2D in two 6-patient cohorts diagnosed with 2 indications. In addition to conventional measures of safety, tolerability, efficacy, and pharmacokinetics, the phase I study employs a rich suite of biomarkers analysis using histology, transcriptomics and immunomonitoring to provide early insights into the pharmacological activity of EVT801. At end of September 2023, 20 patients have been treated in five different cohorts for EVT801 doses ranging from 50mg QD to 400mg BID. At this date, only 1 dose limiting toxicity was declared at the higher dose. Conclusion: Collection of safety data on these first enrolled patients showed no safety alert which allows further dose escalation and reinforces the good safety profile of EVT801 observed during the preclinical toxicology studies. Citation Format: Carlos Gomez-Roca, Philippe Cassier, Michael Fitzgerald, Lise Davenne, Cristina Costantin, Philippe Rochaix, Jean-Pierre Delord, John Friend, Andrea Nizzardo, Alessia Tagliavini, Marco Pergher, Pierre Fons, Marie Mandron. EVT801, a novel selective VEGFR-3 inhibitor targeting tumor angiogenesis, is pursuing dose escalation stage of phase I first-in-human study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT088.

Abstract INTRODUCTION Radiotherapy (RT) is effective for brain metastases (BM) or leptomeningeal disease (LMD), but disease control is poor for tumors harboring PI3K pathway alterations (PI3Kalt). We hypothesized that combining RT with paxalisib, a CNS-penetrant PI3K/mTOR inhibitor, offers therapeutic synergy. The role of plasma circulating tumor DNA (ctDNA) to confirm study eligibility and response monitoring is unclear. METHODS This is a multi-institutional, phase I trial of concurrent paxalisib and whole brain RT (30 Gy in 10 fractions, NCT04192981) for CNS disease. To establish eligibility, patients had documented PI3Kalt performed using institutional tumor targeted sequencing (MSK-IMPACT) of primary or metastatic tumors (eligibility sequencing). Plasma ctDNA was collected at baseline and 3-, 6- and 9-months post-treatment and sequenced using institutional plasma ctDNA targeted sequencing platform (MSK-ACCESS). Our objectives were to a) determine PI3Kalt concordance between eligibility sequencing and baseline ctDNA and b) analyze quantitative post-treatment plasma ctDNA dynamics. RESULTS Between 3/2020-5/2023, 15 patients were enrolled to receive daily paxalisib (45mg, n = 11 or 60mg, n = 4) and WBRT. Twelve were treated for BM and 3 for LMD. Eligibility sequencing was determined a median of 8.1months (range: 2.4-38.6) pre-study enrollment and the most common PI3Kalt were PIK3CA (n = 7), INPP4B (n = 2), INPPL1 (n = 1) and PIK3CG (n = 1), and PTEN copy number alterations (homdel n = 2). Baseline ctDNA was sequenced from 13/15 patients (18 samples: baseline = 13, 3-months = 4 and 6-months = 1). We confirmed 100% of PIK3CA mutations using baseline ctDNA. One breast cancer BM patient demonstrated significant variant allele frequency reduction from 50% (baseline) to 0.1% (3-months post-treatment) for two tracked PIK3CA mutations (p.E545K and p.A1066V), which correlated with a partial response per RANO-BM. CONCLUSIONS For patients on genomically-driven targeted therapy and RT combination trials, alterations detected using plasma ctDNA, especially PIK3CA, may be used to rapidly assess eligibility. Serial ctDNA collection may serve as a potential response biomarker with further validation.

Aberrant activation of the PI3K-AKT pathway is common in many cancers, including melanoma, and AKT1, 2 and 3 (AKT1-3) are bona fide oncoprotein kinases with well-validated downstream effectors. However, efforts to pharmacologically inhibit AKT have proven to be largely ineffective. In this study, we observed paradoxical effects following either pharmacologic or genetic inhibition of AKT1-3 in melanoma cells. Although pharmacological inhibition was without effect, genetic silencing of all three AKT paralogs significantly induced melanoma cell death through effects on mTOR. This phenotype was rescued by exogenous AKT1 expression in a kinase-dependent manner. Pharmacological inhibition of PI3K and mTOR with a novel dual inhibitor effectively suppressed melanoma cell proliferation in vitro and inhibited tumor growth in vivo. Furthermore, this single-agent-targeted therapy was well-tolerated in vivo and was effective against MAPK inhibitor-resistant patient-derived melanoma xenografts. These results suggest that inhibition of PI3K and mTOR with this novel dual inhibitor may represent a promising therapeutic strategy in this disease in both the first-line and MAPK inhibitor-resistant setting.

Abstract Introduction: EVT801 is a highly selective, orally available VEGFR3 inhibitor that strongly inhibits angiogenesis without inducing hypoxia, considered one of the main causes of cancer-associated immunosuppression. EVT801 has shown compelling single agent efficacy in multiple in vivo models. In addition, combination of EVT801 and Immune Checkpoint Therapy (ICT) agents shows additive effects, and thus holds promise for combination treatment without induction of hypoxia-induced-immunosuppression. A phase I clinical trial is underway. Methods: The phase I trial will consist of two sequential stages. During the first stage, EVT801 will be administered to patients with advanced solid tumors in a multiple ascending dose study using an accelerated 3+3 design (1 patient per dose until grade 2 toxicities are observed) in up to 48 patients in 8 dose levels. The primary objective is to determine the maximum tolerated dose (MTD) and recommended phase 2 dose (RP2D). Stage 2 will focus on validation of this RP2D in two 6-patient cohorts diagnosed with soft tissue sarcoma or renal cell carcinoma. These patients will participate in intensive analyses involving several biomarkers to better understand the pharmacological activity of the drug. A potential third stage, consisting of a multiple ascending dose evaluation of the combination of EVT801 with immuno-oncology drugs, may be added to the ongoing trial, pending further technical discussions with physicians and regulators. In addition to conventional measures of safety, tolerability, efficacy, and pharmacokinetics, the phase I study will employ a rich suite of histological, immunological, and radiological biomarkers to provide early insights into the activity of EVT801. Citation Format: Carlos Gomez roca, Philippe Cassier, Marie Mandron, Myriam Estrabaut, Nathalie Delesque-Touchard, Adam C. Smith, Amy Klawitter, Leesa Gentry, Pierre Fons, Michael R. Paillasse, Lise Davenne, Michael Fitzgerald, James Garner, Jean-Pierre Delord. EVT801, a novel selective VEGFR-3 inhibitor targeting tumor angiogenesis, started enrollment for its phase I first-in-human study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr CT206.

Abstract BACKGROUND GDC-0084 is a potent, oral, selective small molecule inhibitor of class I phosphoinositide 3-kinase and mammalian target of rapamycin (PI3K/mTOR). GDC-0084 crosses the blood-brain barrier and achieves a brain / plasma ratio of approximately 1.0. GDC-0084 was given as once daily dosing in a phase 1 study (Wen et al, J Clin Oncol 34, 2016(15) suppl.2012) in 47 patients with recurrent high-grade gliomas. The adverse events were generally consistent with the established PI3K/mTOR inhibitor class-effects. The MTD identified was 45mg once daily. METHODS The current study is conducted in the newly diagnosed GBM patient with unmethylated MGMT promotor status upon completion of standard adjuvant XRT/TMZ. It has a 2-part design: an open-label, dose-escalation phase to assess the safety, tolerability, MTD (Part 1, followed by an expansion cohort (Part 2) commencing once MTD is established. Dose-escalation started at 60mg, and progressed in 15mg increments, per standard 3 + 3 rules. Part 2 recruits 20 patients, who are randomized to take GDC-0084 at the identified MTD, in fed and fasted states. RESULTS Part 1 of the study is complete. There were no DLTs among 3 pts treated at the 60mg. Among 6 pts treated at 75mg, DLTs were identified as hyperglycaemia (symptomatic) and oral mucositis. Adverse effects seen were generally modest, manageable and consistent with the PI3K-class. PK parameters are in line with phase 1 data. Part 2 recruitment is ongoing. CONCLUSION GDC-0084 displays a safety profile consistent with previous data in recurrent high-grade glioma but appears better tolerated in the newly diagnosed GBM setting. An MTD of 60mg is identified.