Lars Blumenstein

Abstract PD5-5: Phase I study of the PI3Kα inhibitor BYL719 plus fulvestrant in patients with PIK3CA-altered and wild type ER+/HER2- locally advanced or metastatic breast cancer

Background: BYL719 selectively inhibits the α-isoform of Class I PI3K. PI3Kα is encoded by PIK3CA, a frequently altered gene in human cancers. Preclinical data indicate BYL719 may be more effective in patients (pts) with PIK3CA-altered tumors; however there are data to suggest that PIK3CA-wild-type (wt) tumors may also be sensitive to BYL719. Here, we present updated data from the Phase I study of BYL719 + fulvestrant in pts with PIK3CA-altered or -wt ER+/HER2– locally advanced/metastatic breast cancer (BC) (NCT01219699). Methods: Adult women with PIK3CA-altered (mutation or amplification) ER+/HER2– BC received continuous oral BYL719 (300–400 mg/day; 28-day cycles) + fixed-dose fulvestrant (500 mg every 4 weeks, plus an additional dose 2 weeks after first dose) during dose escalation and expansion. Pts with PIK3CA-wt ER+/HER2– BC were enrolled into the dose expansion to receive BYL719 400 mg/day + fulvestrant. A Bayesian logistic regression model with overdose control guided dose escalation. Primary objective: to determine the maximum tolerated dose (MTD) and/or recommended phase II dose (RP2D) of BYL719 in combination with fulvestrant, which was declared previously as 400 mg/day. An expansion cohort at the MTD assessed safety (CTCAE v4.0), tolerability, pharmacokinetics (PK), and preliminary efficacy (RECIST v1.0). Results: As of May 2, 2014, 64 pts (PIK3CA-altered n=41; PIK3CA-wt n=16; PIK3CA status unknown/pending n=7) received BYL719 300–400 mg/day + fulvestrant. Median number of prior antineoplastic therapies: 5 (range: 1–12) for pts with PIK3CA-altered tumors and 5 (range: 4–16) for pts with PIK3CA-wt tumors. Prior fulvestrant treatment: 19 (46%) and 7 (44%) pts with PIK3CA-altered and -wt tumors, respectively. Overall, the most common (≥25%) adverse events (AEs; all grades/all doses) suspected to be study drug-related were hyperglycemia (41%), diarrhea (34%), nausea (30%), and vomiting (25%). The most common (>10%) study drug-related Grade 3/4 AEs (all doses) were maculopapular rash (14%) and hyperglycemia (13%). Preliminary antitumor activity was observed in this trial. At data cut-off, partial responses (PRs) were observed in 2 patients with PIK3CA-altered tumors evaluable for response (2/33, 6%), but no PRs were observed in the 15 evaluable patients with PIK3CA-wt tumors. Duration of exposure was >16 weeks in 24 (59%) patients with PIK3CA-altered tumors and in 5 (31%) patients with PIK3CA-wt tumors. PK and exposure of BYL719 + fulvestrant was similar to that observed with single-agent BYL719 at the same dose levels. At data cut-off, treatment was ongoing in 20 (49%) and 2 (13%) pts with PIK3CA-altered and -wt tumors, respectively. Conclusions: BYL719 + fulvestrant demonstrated a favorable safety profile in pts with PIK3CA-altered and -wt ER+/HER2– BC, with mostly on-target effects (i.e. hyperglycemia, rash). Preliminary clinical activity was seen in pts with PIK3CA-altered and -wt tumors, but confirmed PRs were only observed in pts with PIK3CA-altered tumors. The low number of pts with PIK3CA-wt tumors limits further conclusion. Citation Format: Filip Janku, Dejan Juric, Javier Cortes, Hope Rugo, Howard A Burris, Martin Schuler, Barbara Deschler-Baier, Mark R Middleton, Marta Gil-Martin, Jordan Berlin, Eric Winer, Douglas Bootle, Lars Blumenstein, David Demanse, Christina Coughlin, Cornelia Quadt, Jose Baselga. Phase I study of the PI3Kα inhibitor BYL719 plus fulvestrant in patients with PIK3CA-altered and wild type ER+/HER2- locally advanced or metastatic breast cancer [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr PD5-5.

Phosphatidylinositol 3-Kinase α–Selective Inhibition With Alpelisib (BYL719) in PIK3CA-Altered Solid Tumors: Results From the First-in-Human Study

Purpose We report the first-in-human phase Ia study to our knowledge ( ClinicalTrials.gov identifier: NCT01219699) identifying the maximum tolerated dose and assessing safety and preliminary efficacy of single-agent alpelisib (BYL719), an oral phosphatidylinositol 3-kinase α (PI3Kα)-selective inhibitor. Patients and Methods In the dose-escalation phase, patients with PIK3CA-altered advanced solid tumors received once-daily or twice-daily oral alpelisib on a continuous schedule. In the dose-expansion phase, patients with PIK3CA-altered solid tumors and PIK3CA-wild-type, estrogen receptor-positive/human epidermal growth factor receptor 2-negative breast cancer received alpelisib 400 mg once daily. Results One hundred thirty-four patients received treatment. Alpelisib maximum tolerated doses were established as 400 mg once daily and 150 mg twice daily. Nine patients (13.2%) in the dose-escalation phase had dose-limiting toxicities of hyperglycemia (n = 6), nausea (n = 2), and both hyperglycemia and hypophosphatemia (n = 1). Frequent all-grade, treatment-related adverse events included hyperglycemia (51.5%), nausea (50.0%), decreased appetite (41.8%), diarrhea (40.3%), and vomiting (31.3%). Alpelisib was rapidly absorbed; half-life was 7.6 hours at 400 mg once daily with minimal accumulation. Objective tumor responses were observed at doses ≥ 270 mg once daily; overall response rate was 6.0% (n = 8; one patient with endometrial cancer had a complete response, and seven patients with cervical, breast, endometrial, colon, and rectal cancers had partial responses). Stable disease was achieved in 70 (52.2%) patients and was maintained > 24 weeks in 13 (9.7%) patients; disease control rate (complete and partial responses and stable disease) was 58.2%. In patients with estrogen receptor-positive/human epidermal growth factor receptor 2-negative breast cancer, median progression-free survival was 5.5 months. Frequently mutated genes (≥ 10% tumors) included TP53 (51.3%), APC (23.7%), KRAS (22.4%), ARID1A (13.2%), and FBXW7 (10.5%). Conclusion Alpelisib demonstrated a tolerable safety profile and encouraging preliminary activity in patients with PIK3CA-altered solid tumors, supporting the rationale for selective PI3Kα inhibition in combination with other agents for the treatment of PIK3CA-mutant tumors.

Synthesis and biological evaluation of new cytotoxic indazolo[4,3-gh]isoquinolinone derivatives

A series of indazolo[4,3-gh]isoquinolinones derivatives have been synthesized to decrease cardiotoxic side effects in comparison to Mitoxantrone. The antiproliferative effects of different side chains were investigated and tested on at least four different cell lines of cervix, ovarian, CNS, NSCLC (non-small-cell lung cancer) and colon carcinoma. In addition to antiproliferative activities, influence on cell cycle and intercalation behavior have been tested.

Abstract 871: Dual inhibition of PI3K and Erk1/2 shows synergy and efficacy in human tumor cells, either by using drug combinations or novel dual PI3K/Erk inhibitors

The Ras/Raf/Mek/Erk and the PI3K/Akt signaling pathways are frequently deregulated in cancer due to activation of upstream receptors, decreased expression of tumor suppressors like PTEN or activating mutations of Ras, B-Raf or PI3K. Several clinical studies with kinase inhibitors targeting single members of the Ras/Raf/Mek/Erk or the PI3K/Akt pathway are ongoing. However, preclinical and clinical trials indicated limited success by single pathway inhibition and different resistance mechanisms were defined. Further clinical trials with drug combinations of Ras/Raf/Mek/Erk and PI3K/Akt suppressors have been initiated, suggesting a more favorable outcome than targeting only one single pathway, Ras/Raf/Mek/Erk or PI3K/Akt. We performed in-vitro combination experiments with Raf, Mek or Erk inhibitors and PI3K or Akt inhibitors in several human tumor cell lines. Strong synergy was achieved with various combinations including the Mek inhibitor CI-1040 and the PI3K inhibitor GDC-0941 (CI = 0.17) or the Erk inhibitor AEZS-131 and the PI3K inhibitor D-117073 (CI = 0.23) in A549 cells. Also Raf inhibitors like Sorafenib and Zelboraf were combined with different PI3K inhibitors, resulting in synergistic anti-proliferative activity. Due to the attractiveness of parallel inhibition of the Ras/Raf/Mek/Erk and PI3K/Akt pathways, we developed AEZS-136 that concurrently inhibits Erk1/2 (IC50 ∼ 50nM) and PI3K (IC50 ∼ 100nM) by an ATP competitive mode of action. Derivatives of the dual PI3K/Erk inhibitor were co-crystallized with Erk2 and PI3Kα enabling an optimization process by SAR driven medicinal chemistry. The anti-proliferative efficacy of AEZS-136 was evaluated in more than 40 human tumor cell lines and physico-chemical as well as in-vitro ADMET properties were widely assessed. Furthermore, the in-vivo pharmacokinetics and anti-tumor efficacy was explored. AEZS-136 was well tolerated and showed dose dependent inhibition of human colon tumor growth of up to 74% in a Hct116 mouse model. Here we present the concept of dual targeting of the Ras/Raf/Mek/Erk and the PI3K/Akt pathways, either by using drug combinations or our novel dual PI3K/Erk inhibitor. AEZS-136 is a small molecule in preclinical development showing a uniquely advantageous kinase inhibition profile. Broad clinical anti-tumor activity is expected for AEZS-136 in tumors with deregulated Ras/Raf/Mek/Erk and PI3K-Akt signaling. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 871. doi:1538-7445.AM2012-871

Abstract 5476: LHRH receptor targeting as mechanism of anti-tumor activity for cytotoxic conjugates of Disorazol Z with the LHRH receptor agonistic peptide D-Lys6-LHRH.

Background For drug-targeting aimed at the treatment of LHRH receptor overexpressing cancers the LHRH receptor agonistic peptide D-Lys6-LHRH has been conjugated to the novel highly cytotoxic natural compound Disorazol Z. As shown previously by early proof of concept in an ovary cancer xenograft model, differentially linked conjugates possess a high potential regarding the treatment of LHRH-R positive tumors [1]. Here we present further characterization of these conjugates with respect to PK/PD parameter and provide evidence that LHRH receptor targeting significantly contributes to their mechanism of action. Materials and Methods LHRH-R competitive binding, calcium release and cytotoxic activity were measured by Tag-Lite technology (Cisbio), and Fluo-4 (Invitrogen) or Resazurin-based detection, respectively. PK parameter were assessed by standard procedures followed by LC-MS/MS analysis. For the xenograft studies, tumor fragments were transplanted subcutaneously in female nude mice and treatment was started at a tumor size of approx. 100 mm3. Results Disorazol Z - D-Lys6-LHRH conjugates showed varying cytotoxic activity from single digit nanomolar to higher submicromolar EC50 values but comparable LHRH receptor binding and activation in the low nanomolar EC50 range. Comparison in ovarian and triple negative breast cancer xenograft models revealed potent inhibition of tumor growth for the conjugates, whereas equimolar dosing of Disorazol Z failed to reach statistical significance. PK analysis showed substantial plasma levels for the conjugates with only minor release of Disorazol Z, pointing to stabilization by conjugation and demonstrating reasonable half-life of the intact conjugates as prerequisite for tumor targeting. In the same tumor models, competition by previous administration of D-Lys6-LHRH provides evidence for LHRH receptor targeting as mechanism of action. Increased sensitivity of LHRH receptor overexpressing cells towards conjugate cytotoxicity, i.e. leading to about 30 fold decreased EC50 values for the conjugate AEZS-125, further supports the LHRH-R dependency of conjugate efficacy. Conclusions The presented LHRH receptor-dependent efficacies of Disorazol Z - D-Lys6-LHRH conjugates in vitro and in mouse xenograft models support the principle of tumor targeting by the LHRH receptor as already employed by the drug candidate AEZS-108, which is currently in phase II clinical studies. Preclinical development of Disorazol Z conjugates will be started in the first half of 2013. Citation Format: Babette Aicher, Tilmann Schuster, Lars Blumenstein, Antje Schubert, Carsten Grundker, Joerg B. Engel, Olaf Ortmann, Rolf Mueller, Eckhard Guenther, Matthias Gerlach, Michael Teifel. LHRH receptor targeting as mechanism of anti-tumor activity for cytotoxic conjugates of Disorazol Z with the LHRH receptor agonistic peptide D-Lys6-LHRH. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5476. doi:10.1158/1538-7445.AM2013-5476

Abstract 3563: A highly selective Erk1/2 Inhibitor with in-vivo anti tumor potency

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Mutations in receptor tyrosine kinases, Ras or B-Raf frequently manifest constitutive activation of the Ras/Raf/Mek/Erk pathway in a variety of human cancers. The mitogen activated protein kinase Erk1/2 is an important regulator of cell proliferation and was shown to be over-activated by these dysregulated or mutated upstream signals. AEZS-131 selectively inhibits Erk1/2 with an IC50 of 4 nM, blocks cellular Rsk-1 phosphorylation, modulates downstream cellular substrate activation, arrests tumor cells in G1 and inhibits the growth of multiple human tumor cell lines in the nanomolar range. We have studied the response of human cancer cells (Hct116) in in-vivo mouse xenograft experiments. AEZS-131 significantly inhibited tumor growth at daily doses of 30mg/kg. Currently, AEZS-131 is undergoing in-vivo combination experiments with PI3K inhibitors. Since pharmacological inhibition of Erk1/2 reverses Ras and Raf activation also in cells demonstrating resistancy to common Raf inhibitors like GDC-0879 and PLX4720, it appears likely to put more attention on the downstream kinase Erk1/2 as therapeutic target. Furthermore, in instances of coincident activation of the Raf and PI3K pathways, combinations of AEZS-131 and PI3K inhibitors may prove efficacious. Here we present the in-vitro and in-vivo characterisation of the first in-class Erk1/2 inhibitor showing in-vivo efficacy as single therapy agent. Early development of the Erk1/2 inhibitor AEZS-131 by AEterna Zentaris is integral part of our in-house kinase research program comprising the investigation of different compounds for single Erk1/2 inhibition, single PI3K inhibition and dual kinase inhibitions. All compounds are exclusively synthesized by our Drug Discovery Department. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3563. doi:10.1158/1538-7445.AM2011-3563

Abstract 4474: Dual inhibitors for PI3K and Erk induce growth inhibition of tumor cells

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC The Ras/Raf/Mek/Erk and the PI3K-Akt signaling pathways are prime targets for drug discovery in proliferative diseases such as cancer. The results of research to date indicate that both the MAPK and the PI3K signaling pathways represent therapeutic intervention points for the clinical treatment of malignant tumors. Our multi-parameter optimization program for kinase inhibitor selectivity, cellular efficacy, physico-chemical and in-vitro ADMET properties has led to the identification of a small molecular compound class with an uniquely advantageous dual kinase inhibition profile. These ATP competitive compounds inhibit Erk and PI3K in the nanomolar range and exert high selectivity against other serine threonine and tyrosine kinases. The anti-tumor efficacy of these dual kinase inhibitors was evaluated in diverse human tumor cell lines like HCT116, A549, MDA-MB 468, PC-3 and others. Physicochemical and in-vitro ADMET and safety parameters have been widely assessed. Furthermore in-vivo pharmacokinetic experiments showed plasma profiles expected to result in beneficial in-vivo anti-tumor efficacy. Here we present the key characteristics of the compound class that led to the selection of AEZS-132 for in-vivo experiments with tumor bearing nude mice. The optimization of ADME and physicochemical properties such as solubility, permeability and metabolic stability by medicinal chemistry is ongoing. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4474.