[Webinar] How to better optimize an oncology drug discovery program
Register for this webinar discussing how conventional and innovative integrated technological skills should be incorporated into an oncology program.Read more
Nerina Dodic | Nicolas Ancellin | Morgane Bordessoules | Anne Bouillot | Nathan Butin | Cédric Charrier | Marie-Hélène Fouchet | Alain Laroze | Anne-Pascale Luzy | Alexandre Moquette | Julia Pilot | Guillaume Serin | Jean-François Mirjolet | Fabrice Viviani | Christophe Parsy
Small molecule macrocyclic kinase inhibitors have attracted significant attention in drug discovery over the past years with drugs approval such as lorlatinib demonstrating the clinical relevance of this approach. We developed our expertise to further optimize those cyclic molecules. Having low molecular weight, they favorably alter the biological and physiochemical properties as well as selectivity, as compared to their linear parent, yielding high-quality drug candidates. While focused on kinase inhibitors, macrocyclic derivatives could be potentially turned into bifunctional protein degrader molecules useful for selective cellular knockdown of targeted proteins and investigation of the pharmacological effects. With macrocyclic “probes” from our proprietary library in nanomolar range IC50, we turned our attention to CDK9 inhibitors with good selectivity profile against CDK1/2/5/7.
Previous CDK9 degraders based on acyclic inhibitors have used thalidomide as recruiter of the CRL4CRBN, resulting in successful ubiquitination and proteasomal degradation of the protein of interest. Applying a three-step approach, we first defined the vector of substitution on our macrocyclic “probes” by homology modeling with known acyclic ligands/CDK9 co-crystal structures. Having defined 2 substitution patterns, we synthesized and tested in a biochemical assay the novels “probes” vectorized with the pro-linker moieties, without loss of activity. ODS’7152 was further derived into fully bifunctional molecules with variation on the linker’s nature and length, keeping thalidomide as the E3 ligase recruiter. Biophysical and biological properties were further evaluated in an MTS assay against several cell lines (HCT116, Jurkat E6.2, MOLT-4 and MV4-11) to assess cell proliferation and viability. The E3 Ligase engagement along with cell penetration was measured in a NanoBRET™ assay and compared to MDR_MDCK permeability. Finally, a selection of 4 PROTACs were evaluated in a dose response assay on MV4-11 for protein degradation phosphorylation of CTD-pol II and selectivity among several CDKs. Our findings support the rapid derivatization of macrocyclic “probes” inhibitors into macrocyclic-PROTAC as a strategy to generate early chemical biology tools with maintained potency and selectivity between homologous targets. Optimization of the physico-chemical and ADME properties of molecules can lead to drug candidates with distinct pharmacological effects as compared to the parent linear kinase inhibitors.
Emma Renard2 | Olivier Raguin1 | Victor Goncalves2 | Céline Mothes1 | Mathieu Moreau2 | Claire Bernhard2 | Peggy Provent1 | Pierre Adumeau1,2 | Romane Vizier2 | Frederic Boschetti3 | Franck Denat2 | Cyril Berthet1
1 – Oncodesign | 2 – ICMUB UMR 6302, CNRS, Université Bourgogne Franche-Comté | 3 – CheMatech
Molecular Radiotherapy (MRT) targeting SSTR2 or PSMA have proven to be highly efficient for treatment of neuroendocrine or metastatic prostate cancer respectively. Beyond the leading radiopharmaceutical molecules 177Lu-DOTATATE or 177Lu-PSMA-617, a variety of vectors (small molecules, peptides, panel of biologics) have been developed on the same targets in order to improve the biodistribution within the tumor, the blood clearance, the route of elimination or the dosimetry.
Labeling of the targeting ligand, whatever its nature, is a crucial step as it may affect significantly the properties of the theranostic conjugate, i.e. its binding affinity, PK and biodistribution. The addition of linkers, such as albumin binding domain or PEG, and choice of chelating agents have a major impact on the chemical and biological properties of the vectors. Random or site-specific bioconjugation, click chemistry, have also to be considered in the early stage as the choice of the selected technology will modify your development plan and manufacturing.
New ligands and biological platforms are now being developed based on this historical knowledge, improved Target Product Profiles are built to conduct optimal lead optimization of MRT. Herein, we will present our lead optimization and preclinical evaluation process to select efficiently good radiolabeled molecules and list the key parameters to be checked. To date, it remains hard to predict the behavior of the modified bioconjugated molecules, and versatile synthesis strategies are needed to screen various combinations of radiometal complexes / linker / conjugation function, in order to converge rapidly to the optimized bioconjugate. For instance, we will present a study case where the conjugation of various bifunctional chelating agents on a small NTS1 receptor antagonist resulted in drastically different in vivo behavior of the resulting 68Ga-labeled compounds.
Once optimal in vivo tumor uptake has been achieved, preclinical evaluation requires the selection of appropriate and relevant models, driven by target expression, radioresistance, and potentially tumor immune infiltrate for combination studies with immunotherapies. The therapeutic evaluation should take into consideration the dose and specific activity, tolerance of the model related to ionizing radiations and the scheduling of treatment (cumulated dose, fractionation).
Olivier Duchamp | Gaël Krysa | Adélaïde Ferment | Loïc Morgand | Hugo Quillery | Robin Artus | Maxime Ramelet | Jeremy Odillard | Peggy Provent | Sylvie Maubant | Caroline Mignard | Fabrice Viviani | Samira Benhamouche-Trouillet
Hepatocellular carcinoma (HCC) is the 4th leading cause of cancer-related death and accounts for over 80% of primary liver cancer worldwide. Early stage HCC can be treated by local ablation, surgical resection or liver transplantation. Systemic pharmacological options are limited (kinase and immune-checkpoint inhibitors). Most cases of liver cancer occur in the setting of chronic liver diseases. Risk factors include chronic Hepatitis B and C, alcohol addiction and metabolic diseases. HCC is a multistep process comprising chronic liver injury, inflammation, fibrosis/cirrhosis and cancer formation. Therefore, providing palliative and curative options remains a high medical need. And with the recent success of immunotherapies in HCC, mouse models that better recapitulate the human disease and antitumor immune response are needed. In order to better evaluate new preventive and curative treatments of liver cancers we developed complementary and integrated strategies to mimic the liver cancer initiation and progression steps in mouse models.
These models involve chemotoxic agents, diet-induced disorders and syngeneic or xenogeneic tumor implantation strategies. We established an orthotopic syngeneic model using Hepa1.6 mouse liver hepatoma cells, characterized through liver index, alpha-fetoprotein measurement in serum and liver, and MRI. The response to chemotherapy (sorafenib) and immunotherapy (anti PD-1, anti-TLR) was also assessed and show moderate to high efficiency. Moreover, a panel of xenograft models including Patient-Derived Xenograft models (PDXs) are available to assess new treatment options in human HCC with regards of the genetic mutations and the variety of etiologies seen in human. But as xenograft models are not completely mimicking the human situation of both immune and liver microenvironment, we have recently initiated the development of a double humanized (immune and liver) transgenic mouse as a better host for human tumor engraftment. During the course of HCC formation, the liver undergoes cycles of inflammation, necrosis with regeneration, fibrosis, cell dysplasia and ultimately HCC. Thus, we developed a model induced by a low dose of streptozotocin and high fat diet regimen. In this model, mice develop metabolic syndrome NASH and fibrosis within 12 weeks and HCC within 16 weeks. Of interest, 100% of male mice develop HCC within 16 weeks, in accordance with studies showing that men had a 2-to 7-fold higher risk of developing HCC in human. Treatment of mice with lenvatinib alone or in combination with anti-PD1 increases survival and reduces tumor burden as shown with reduced liver weight/body weight ratio at 16 weeks. Altogether, these results demonstrate the usefulness of this comprehensive platform of preclinical in vivo HCC models to discover and identify novel therapeutic strategies that could circumvent the progression of liver cancers.
Sylvie Maubant l Marie Leblanc l Elisabeth Bertrand l Audrey Bertaux l Loïc Morgand l Maxime Ramelet l Marie Lux l Olivier Duchamp l Fabrice Viviani
Preclinical and clinical studies have shed light on the beneficial role of bacteria for cancer therapy. Indeed, these studies have demonstrated that these microorganisms have beneficial properties that allow them to selectively colonize tumor and that they could also be considered as predictive drug efficacy biomarkers. Based on these results, bacteria are now used for delivering therapeutic components or for shaping the gut microbiota. Ultimately these approaches lead to the activation of an immune response against the tumor.
Owing to our scientific and technological expertise in manipulating microbes, we propose tailor-made strategies for investigating the efficacy of bacteria-based treatments and/or the effect of therapies on the microbiota both in vitro and in vivo.
Different methods/analyses can be used for culturing, detecting, quantifying, identifying and localizing live bacteria (e.g. counting of CFUs, PMA-qPCR, 16 rRNA gene sequencing, mass spectrometry, bioluminescence) in simple or complex samples (e.g. from culture of single bacterial species to rodent/human stools or other tissues). In addition, a continuum of assays allows us to evaluate the impact of bacteria or derived products directly on tumor and/or immune cells (e.g. immune infiltrate and phenotyping, cytokine/chemokine profiling, tumor burden). We will highlight some results obtained in a cancer context such as the immunostimulatory properties of bacteria or their derivatives, the selective colonization of tumor tissue by bacteria, the benefits in delivery of therapeutic proteins or antigens by bacteria, the impact of tumor engraftment/growth on gut microbiota, the effects of chemotherapeutic agents on intestinal microflora, the consequences of supplementation with bacteria or antibiotics treatment on the response to immune checkpoint inhibitors. Altogether these data demonstrate that bacteria are now allies in the treatment of cancer and that our comprehensive platform is suitable for evaluating both in vitro and in vivo therapeutics developed from bacteria (individuals, consortium or derived products; native or modified). All these technologies can be also applied to develop novel therapeutic strategies for inflammatory and infectious diseases known to increase the risk of cancer development.