Alex Matter

Safety and immunogenicity of a virus-like particle pandemic influenza A (H1N1) 2009 vaccine: Results from a double-blinded, randomized Phase I clinical trial in healthy Asian volunteers

METHODS A novel, fully bacterially produced recombinant virus-like particle (VLP) based influenza vaccine (gH1-Qbeta) against A/California/07/2009(H1N1) was tested in a double-blind, randomized phase I clinical trial at two clinical sites in Singapore. The trial evaluated the immunogenicity and safety of gH1-Qbeta in the presence or absence of alhydrogel adjuvant. Healthy adult volunteers with no or low pre-existing immunity against A/California/07/2009 (H1N1) were randomized to receive two intramuscular injections 21 days apart, with 100μg vaccine, containing 42μg hemagglutinin antigen. Antibody responses were measured before and 21 days after each immunization by hemagglutination inhibition (HAI) assays. The primary endpoint was seroconversion on Day 42, defined as percentage of subjects which reach a HAI titer ≥40 or achieve an at least 4-fold rise in HAI titer (with pre-existing immunity). The co-secondary endpoints were safety and seroconversion on Day 21. RESULTS A total of 84 Asian volunteers were enrolled in this study and randomized to receive the adjuvanted (n=43) or the non-adjuvanted (n=41) vaccine. Of those, 43 and 37 respectively (95%) completed the study. There were no deaths or serious adverse events reported during this trial. A total of 535 adverse events occurred during treatment with 49.5% local solicited symptoms, of mostly (76.4%) mild severity. The most common treatment-related systemic symptom was fatigue. The non-adjuvanted vaccine met all primary and secondary endpoints and showed seroconversion in 62.2% and 70.3% of participants respectively on Day 21 and Day 42. While the adjuvanted vaccine showed an increased seroconversion from 25.5% (Day 21) to 51.2% (Day 42), it did not meet the immunogenicity endpoint. CONCLUSION In summary, non-adjuvanted gH1-Qbeta showed similar antibody mediated immunogenicity and a comparable safety profile in healthy humans to commercially available vaccines. These results warrant the consideration of this VLP vaccine platform for the vaccination against influenza infection (HSA CTC1300092).

Enhanced Neutralizing Antibody Titers and Th1 Polarization from a Novel Escherichia coli Derived Pandemic Influenza Vaccine

Influenza pandemics can spread quickly and cost millions of lives; the 2009 H1N1 pandemic highlighted the shortfall in the current vaccine strategy and the need for an improved global response in terms of shortening the time required to manufacture the vaccine and increasing production capacity. Here we describe the pre-clinical assessment of a novel 2009 H1N1 pandemic influenza vaccine based on the E. coli-produced HA globular head domain covalently linked to virus-like particles derived from the bacteriophage Qβ. When formulated with alum adjuvant and used to immunize mice, dose finding studies found that a 10 µg dose of this vaccine (3.7 µg globular HA content) induced antibody titers comparable to a 1.5 µg dose (0.7 µg globular HA content) of the licensed 2009 H1N1 pandemic vaccine Panvax, and significantly reduced viral titers in the lung following challenge with 2009 H1N1 pandemic influenza A/California/07/2009 virus. While Panvax failed to induce marked T cell responses, the novel vaccine stimulated substantial antigen-specific interferon-γ production in splenocytes from immunized mice, alongside enhanced IgG2a antibody production. In ferrets the vaccine elicited neutralizing antibodies, and following challenge with influenza A/California/07/2009 virus reduced morbidity and lowered viral titers in nasal lavages.

Bridging academic science and clinical research in the search for novel targeted anti-cancer agents.

This review starts with a brief history of drug discovery & development, and the place of Asia in this worldwide effort discussed. The conditions and constraints of a successful translational R&D involving academic basic research and clinical research are discussed and the Singapore model for pursuit of open R&D described. The importance of well-characterized, validated drug targets for the search for novel targeted anti-cancer agents is emphasized, as well as a structured, high quality translational R&D. Furthermore, the characteristics of an attractive preclinical development drug candidate are discussed laying the foundation of a successful preclinical development. The most frequent sources of failures are described and risk management at every stage is highly recommended. Organizational factors are also considered to play an important role. The factors to consider before starting a new drug discovery & development project are described, and an example is given of a successful clinical project that has had its roots in local universities and was carried through preclinical development into phase I clinical trials.

Optimization of Selective Mitogen-Activated Protein Kinase Interacting Kinases 1 and 2 Inhibitors for the Treatment of Blast Crisis Leukemia

Chronic myeloid leukemia (CML) is a myeloproliferative disease caused by bcr-abl1, a constitutively active tyrosine kinase fusion gene responsible for an abnormal proliferation of leukemic stem cells (LSCs). Inhibition of BCR-ABL1 kinase activity offers long-term relief to CML patients. However, for a proportion of them, BCR-ABL1 inhibition will become ineffective at treating the disease, and CML will progress to blast crisis (BC) CML with poor prognosis. BC-CML is often associated with excessive phosphorylated eukaryotic translation initiation factor 4E (eIF4E), which renders LSCs capable of proliferating via self-renewal, oblivious to BCR-ABL1 inhibition. In vivo, eIF4E is exclusively phosphorylated on Ser209 by MNK1/2. Consequently, a selective inhibitor of MNK1/2 should reduce the level of phosphorylated eIF4E and re-sensitize LSCs to BCR-ABL1 inhibition, thus hindering the proliferation of BC LSCs. We report herein the structure-activity relationships and pharmacokinetic properties of a selective MNK1/2 inhibitor clinical candidate, ETC-206, which in combination with dasatinib prevents BC-CML LSC self-renewal in vitro and enhances dasatinib antitumor activity in vivo.

Drugs’ development in acute heart failure: what went wrong?

Acute heart failure (AHF) is a major burden disease, with a complex physiopathology, unsatisfactory diagnosis, treatment and a very poor prognosis. In the last two decades, a number of drugs have progressed from preclinical to early and late clinical development, but only a few of them have been approved and added to a stagnant pharmacological armamentarium. We have reviewed the data published on drugs developed for AHF since early 2000s, trying to recognise factors that have worked for a successful approval or for the stoppage of the program, in an attempt to delineate future trajectories for AHF drug development. Our review has identified limitations at both preclinical and clinical levels. At the preclinical level, the major shortcoming is represented by animal models looking at short-term endpoints which do not recapitulate the complexity of the human disease. At the clinical level, the main weakness is given by the disconnect between short-term endpoints assessed in the early stage of drug development, and medium–long-term endpoints requested in Phase 3 for regulatory approval. This is further amplified by the lack of validation and standardisation of short- and long-term endpoints; absence of predictive biomarkers; conduct of studies on heterogeneous populations; and use of different eligibility criteria, time of assessments, drug schedules and background therapies. Key goals remain a better understanding of AHF and the construction of a successful drug development program. A reasonable way to move forward resides in a strong collaboration between main stakeholders of therapeutic innovation: scientific community, industry and regulatory agencies.

Abstract 1172:In vivopharmacokinetic properties and antitumor efficacy of porcupine lead inhibitors in the orthotopic murine MMTV-Wnt1 breast tumor model and the human HPAF-II pancreatic xenograft mouse model

Porcupine (PORCN), a muti-pass integral membrane-bound-O-Acyl acyltransferase (MBOAT), resides in the in the endoplasmic reticulum (ER) and is required for biogenesis of Wnt ligands. The secreted mature Wnt ligands bind to their cognate receptors (Frizzled, LRP5/6 and transmembrane receptor ROR) to form the ligand - receptor complex which is capable to activate the Wnt-β-catenin signalling cascade and downstream signalling pathways such as mTOR, GSK3, Akt, and PKC. The deregulation of and aberrant activation of the various components of Wnt-β-catenin signalling pathway have been implicated in tumorigenesis, cell proliferation, survival and differentiation. Hyperactivity of PORCN is found to be associated with cancerous cell growth. Knockdown of Porcn mRNA significantly reduced the proliferation of breast cancer cells and resulted in the delay of MDA-MB-231 tumor formation in mouse xenograft models (Covey et al 2012). Loss of function mutations of RNF43, a negative regulation of Wnt-signalling via Frizzled receptor, is recently reported to be involved with pancreatic ductal adenocarcinoma. Inhibition of the PDAC cell lines bearing RNF43 mutations enhanced Wnt-β-catenin signalling and resulted in suppression of proliferation and differentiation of PDAC tumor cells (Jiang et al 2013). Taken together porcupine could be an attractive therapeutic approach for a particular Wnt-driven cancer population. We have identified the porcupine lead compounds (ETC-159, ETC-535, ETC-611 and ETC-539) from different novel chemical scaffolds. The aim of this study was to evaluate their pharmacokinetic properties and antitumor efficacy in different cancer mouse models, the murine MMTV-Wnt1 breast cancer and the human pancreatic HPAF-II cancer. All porcupine lead compounds had good oral pharmacokinetic properties with the absolute oral bioavailability greater than 42%. They had the maximum tolerated dose (MTD-7d) up to 200 mg/kg. They produced antitumor efficacy ranging from 24% to 79% at 1 mg/kg, 38% to 89% at 3 mg/kg, and 58 to 97% at 10 mg/kg in MMTV-Wnt1 tumor mouse model. In vivo inhibition of PORCN led to reduce the expression level of Axin2 in MMTV-Wnt tumors upto 8h. At 100 mg/kg, they produced antitumor efficacy ranging from 34% to 91% in human HPAF-II pancreatic xenograft mouse model. Of 4 porcupine lead compounds, ETC-159 demonstrated great oral pharmacokinetic properties and produced significantly antitumor efficacy (p value Citation Format: Vishal Pendharkar, Yun Shan Chew, Vithya Manoharan, Choon Bing Low, Hongqian Esther Ong, Soo Yei Ho, Wei Ling Wang, Jeyaraj Duraiswamy Athisayamani, Babita Madan, David Virshup, Thomas Hugo Keller, May Ann Lee, Alex Matter, Jeffrey Hill, Kanda Sangthongpitag. In vivo pharmacokinetic properties and antitumor efficacy of porcupine lead inhibitors in the orthotopic murine MMTV-Wnt1 breast tumor model and the human HPAF-II pancreatic xenograft mouse model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1172. doi:10.1158/1538-7445.AM2017-1172

Abstract B13: ETC-159 is a novel PORCN inhibitor effective for treatment of Wnt-addicted genetically defined cancers

Pathologic expression of Wnts is found in cancers, fibrosis and other diseases, hence there is considerable interest in blocking Wnt signaling to achieve therapeutic benefit. Post-translational palmitoleation of Wnts at a highly conserved serine residue is essential for their secretion and function. We have pursued a strategy to block all Wnt-dependent pathways concomitantly using a small molecule inhibitor of the enzyme PORCN that is required for Wnt O-palmitoleation. As Wnt pathway inhibitors advance to clinical trials, the paucity of well-defined biomarkers makes it challenging to monitor response to therapeutics. We have developed a novel PORCN inhibitor ETC-159, which has remarkable efficacy in pre-clinical models of RNF43-mutant pancreatic and ovarian cancers and RSPO3-translocation bearing colorectal cancer patient derived xenografts. ETC-159 reverses Wnt dependent pathology by promoting cellular differentiation. Consistent with the role of Wnt/β-catenin signaling in regulation of diverse cellular functions, RNA sequencing of these ETC-159 sensitive, Wnt driven cancers has revealed significant remodeling of the transcriptome on treatment with ETC-159 with a decrease of cell cycle, stem cell, and proliferation genes and an increase in differentiation markers. Our studies show that coordinate regulation of NOTUM and AXIN2 gene expression may be a useful predictor of sensitivity to PORCN inhibitors. NOTUM is a secreted protein and its levels in serum correlate with tumor growth. Notum has potential to be a pharmacodynamic biomarker that will reflect a response to PORCN and other Wnt pathway inhibitors. Citation Format: Babita Madan, Zhiyuan Ke, Nathan Harmston, Enrico Petretto, Jeffrey Hill, Thomas H. Keller, May Ann Lee, Alex Matter, David M. Virshup. ETC-159 is a novel PORCN inhibitor effective for treatment of Wnt-addicted genetically defined cancers. [abstract]. In: Proceedings of the AACR Special Conference: Developmental Biology and Cancer; Nov 30-Dec 3, 2015; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(4_Suppl):Abstract nr B13.

Abstract 2134: Rational design of selective MNK 1 and 2 kinase inhibitors for the treatment of blast crisis chronic myeloid leukemia patients

The marketed BCR-ABL tyrosine kinase inhibitor (TKI), imatinib (Gleevec™) is a very successful targeted anti-cancer therapy. It has revolutionized the treatment of early stage or chronic phase (CP) chronic myeloid leukemia (CML). Unfortunately, a proportion of CP patients experience suboptimal responses to BCR-ABL TKIs, and progress to blast crisis (BC) stage of CML with poor survival rate. A potential cause of the resistance to TKI is the elevated level of phosphorylated eukaryotic initiation factor 4E (eIF4E), which has been found to be a consistent feature in patient-derived BC-CML samples. Importantly, both in vivo and in vitro studies have demonstrated that the MAP kinase-interacting serine/threonine-protein kinases 1 and 2 (MNK1/2) phosphorylate eIF4E on Ser209, and that the overexpression of eIF4E drives oncogenesis in a variety of cancers including BC-CML. Furthermore, several reports have indicated that eIF4E phosphorylation at Ser209, as well as eIF4E overexpression, is critical to tumor progression. We found that a BC-CML cell line, K562, that expresses a serine to alanine phospho-mutant at position 209 of eIF4E, shows reduced ability to form tumors in mice compared to wildtype eIF4E. In addition, our recent work has demonstrated the importance of the MNK-eIF4E axis in activating BC leukemia stem cell (LSC) function (Lim et al., PNAS18; 110(25):E2298-307, 2013). These data highlight the critical importance of MNK1/2-dependent eIF4E phosphorylation in cancer progression and maintenance, and suggests that inhibition of MNK1/2 is an attractive therapeutic approach to treat BC-CML. Consequently, we set out to identify selective inhibitors of the MNK1/2 kinases to treat BC-CML patients. Here, we report our hit finding strategy, as well as our hit to lead optimization process. Results describing structure activity relationships, pharmacokinetics properties, and biochemical characteristics of a highly specific MNK1/2 inhibitor, are presented. Our data demonstrate that drug-like molecules can be developed to potently and specifically inhibit the MNK kinases. We also show that simultaneous inhibition of MNK and BCR-ABL is effective at inhibiting BCR-ABL-driven growth and proliferation, as well as inhibiting the MNK-eIF4E-dependent self-renewal function of BC-LSCs. A combination of selective MNK and BCR-ABL inhibitors may provide clinical benefit to BC-CML patients. Citation Format: Kassoum Nacro, Haiyan Yang, Melvyn Wai Tuck Ho, Yoon Sheng Yeap, Lohitha Rao Chennamaneni, Shi Hua Ang, Eldwin Sum Wai Tan, Athisayamani Jeyaraj Duraiswamy, Sharon Lim, Boping Liu, Esther Hongqian Ong, Meng Ling Choong, Shi Jing Tai, Vithya Manoharan, Vishal Pendharkar, Lijun Ding, Yun Shan Chew, Joma Kanikadu Joy, John LW Kuan, Perlyn Z. Kwek, Anders Poulsen, May Ann Lee, Kanda Sangthongpitag, Charles Chuah, Tiong S. Ong, Jeffrey Hill, Thomas H. Keller, Alex Matter. Rational design of selective MNK 1 and 2 kinase inhibitors for the treatment of blast crisis chronic myeloid leukemia patients. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2134.

Abstract 948: Discovery of dual MNK 1 and 2 and BCR-ABL kinase inhibitors for the treatment of blast crisis chronic myeloid leukemia

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Although targeting the BCR-ABL kinase with tyrosine kinase inhibitors (TKI) is effective in chronic phase (CP) CML, targeting BCR-ABL alone is not sufficient to treat late-stage or blast crisis (BC) CML. In BC CML, granulocyte macrophage progenitors (GMPs) acquire the ability to function as leukemic stem cells (LSCs), and are thought to act as a reservoir for TKI resistance. While long-term remission in BC CML is possible with allogeneic stem cell transplantation, only a minority of patients are eligible for transplantation, and may experience high morbidity and mortality. Safe and effective BC CML therapy will likely have to target the BC LSC population to ensure long-term disease control. Recent studies have shown the importance of the MAP kinase interacting serine/threonine kinase (MNK1/2)-eukaryotic translation initiation factor 4E (eIF4E) axis in sustaining BC LSC self renewal. Thus, a single agent which inhibits both BCR-ABL and MNK1/2 kinase simultaneously represents a rational approach to target BC LSCs. Such an agent should be able to distinguish between normal hematopoietic stem cells (HSC) and LSCs. Here, we report the discovery, structure activity relationships, pharmacokinetic properties, and biochemical characteristics of dual MNK1/2 and BCR-ABL inhibitors, ETC-027 and ETC-219. These compounds are able to prevent eIF4E phosphorylation, BCR-ABL-driven growth and proliferation of BC CML cells, as well as inhibit the MNK-eIF4E-dependent self-renewal function of BC LSCs, while leaving normal HSCs untouched. Citation Format: Joseph Cherian, Kassoum Nacro, Zhi Ying Poh, Samantha Guo, Melvyn Ho, Haiyan Yang, Sharon Lim, Meng Ling Choong, Jun Li Ding, Joma Kanikadu Joy, Zekui Perlyn Kwek, Boping Liu, Hongqian Esther Ong, Vishal Pendharkar, Anders Poulsen, May Ann Lee, Kanda Sangthongpitag, Charles Chuah, Tiong S. Ong, Jeffrey Hill, Thomas H. Keller, Alex Matter. Discovery of dual MNK 1 and 2 and BCR-ABL kinase inhibitors for the treatment of blast crisis chronic myeloid leukemia. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 948. doi:10.1158/1538-7445.AM2014-948