The serine/threonine kinase DYRK1A has been implicated in regulation of a variety of cellular processes associated with cancer progression, including cell cycle control, DNA damage repair, protection ...from apoptosis, cell differentiation, and metastasis. In addition, elevated-level DYRK1A activity has been associated with increased severity of symptoms in Down’s syndrome. A selective inhibitor of DYRK1A could therefore be of therapeutic benefit. We have used fragment and structure-based discovery methods to identify a highly selective, well-tolerated, brain-penetrant DYRK1A inhibitor which showed in vivo activity in a tumor model. The inhibitor provides a useful tool compound for further exploration of the effect of DYRK1A inhibition in models of disease.
The kinase DYRK1A is an attractive target for drug discovery programs due to its implication in multiple diseases. Through a fragment screen, we identified a simple biaryl compound that is bound to ...the DYRK1A ATP site with very high efficiency, although with limited selectivity. Structure-guided optimization cycles enabled us to convert this fragment hit into potent and selective DYRK1A inhibitors. Exploiting the structural differences in DYRK1A and its close homologue DYRK2, we were able to fine-tune the selectivity of our inhibitors. Our best compounds potently inhibited DYRK1A in the cell culture and in vivo and demonstrated drug-like properties. The inhibition of DYRK1A in vivo translated into dose-dependent tumor growth inhibition in a model of ovarian carcinoma.
Myeloid cell leukemia 1 (Mcl-1) has emerged as an attractive target for cancer therapy. It is an antiapoptotic member of the Bcl-2 family of proteins, whose upregulation in human cancers is ...associated with high tumor grade, poor survival, and resistance to chemotherapy. Here we report the discovery of our clinical candidate S64315, a selective small molecule inhibitor of Mcl-1. Starting from a fragment derived lead compound, we have conducted structure guided optimization that has led to a significant (3 log) improvement of target affinity as well as cellular potency. The presence of hindered rotation along a biaryl axis has conferred high selectivity to the compounds against other members of the Bcl-2 family. During optimization, we have also established predictive PD markers of Mcl-1 inhibition and achieved both efficient in vitro cell killing and tumor regression in Mcl-1 dependent cancer models. The preclinical candidate has drug-like properties that have enabled its development and entry into clinical trials.
Myeloid cell leukemia 1 (Mcl-1), an antiapoptotic member of the Bcl-2 family of proteins, whose upregulation when observed in human cancers is associated with high tumor grade, poor survival, and ...resistance to chemotherapy, has emerged as an attractive target for cancer therapy. Here, we report the discovery of selective small molecule inhibitors of Mcl-1 that inhibit cellular activity. Fragment screening identified thienopyrimidine amino acids as promising but nonselective hits that were optimized using nuclear magnetic resonance and X-ray-derived structural information. The introduction of hindered rotation along a biaryl axis has conferred high selectivity to the compounds, and cellular activity was brought on scale by offsetting the negative charge of the anchoring carboxylate group. The obtained compounds described here exhibit nanomolar binding affinity and mechanism-based cellular efficacy, caspase induction, and growth inhibition. These early research efforts illustrate drug discovery optimization from thienopyrimidine hits to a lead compound, the chemical series leading to the identification of our more advanced compounds S63845 and S64315.
Abstract
Background: OX40, a member of the tumor necrosis factor receptor superfamily, is a costimulatory molecule promoting survival and cytokine release by effector T cells and inhibiting human ...regulatory T-cell activity. OX40 activation has been explored as a potential treatment for solid malignancies, using mainly agonistic anti-OX40 antibodies. These antibodies rely on Fc gamma receptor (FcγR)-mediated crosslinking to activate the OX40 pathway and have shown so far only limited antitumoral efficacy in clinical settings. One underlying hypothesis is that FcγR crosslinking of these antibodies may not allow optimal activation of the OX40 pathway to fully exploit the potential of this target for immunotherapy of cancer. To overcome this limitation, we designed PRS-352/S095025, a strong PD-L1-dependent OX40 agonistic bispecific molecule, that allows combination of OX40 costimulation and PD-L1 blockade.
Methods: We generated an Anticalin protein that binds with high affinity to human OX40. Anticalin® proteins are approximately 18-20 kDa protein therapeutics derived from human lipocalins that can be engineered to bind with high affinity and specificity to different targets. The PRS-352/S095025 bispecific fusion protein was obtained by genetic fusion of OX40-targeting Anticalin protein to a PD-L1-targeting monoclonal antibody with a modified IgG4 backbone, allowing an optimal activation of OX40 in the presence of PD-L1, but not FcγRs.
Results: We showed that PRS-352/S095025 retains binding to PD-L1 like the anti PD-L1 antibody backbone alone and binds with high affinity to both human and cynomolgus OX40. We showed that PRS-352/S095025 inhibited the PD-1/PD-L1 pathway, as the parental PD-L1 targeting antibody and an approved anti PD-L1 antibody. In addition to its PD-L1 blocking properties, we showed that PRS-352/S095025 activates OX40, and that this activity is dependent on the expression of PD-L1 and not on FcγR, consistent with the desired mechanism of action. Furthermore, we demonstrated that PRS-352/S095025 activity is superior to both PD-L1 benchmark antibodies and to the combination of clinically relevant OX40 and PD-L1 benchmark antibodies. We could also show that PRS352/S095025 strongly stimulated human CD4 T cells. In vivo, we showed that PRS-352/S095025 has a PK profile similar to that of the PD-L1 antibody alone.
Conclusions: We provide here the preclinical characterization of the fusion protein PRS-352/S095025, a bispecific molecule composed of a PD-L1 blocking moiety and an Anticalin protein agonizing OX40. In vitro, this molecule showed the desired MoA, PD-L1 blocking and potent OX40 agonism driven by binding to PD-L1, with superior activity to a clinical stage OX40 agonist.
Citation Format: Lucia Pattarini, Marina Pavlidou, Aizea Kastresana Morales, Janet Peper-Gabriel, Matthieu Riviere, Didier Demarles, Alix Scholer-Dahirel, Veronique Blanc, Shane Olwill. The anticalin-antibody bispecific PRS-352/S095025 strongly stimulates human CD4+ T cells in a PD-L1-dependent manner 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 LB220.
Study Objective. To investigate the steady‐state pharmacokinetics of a triple combination tablet containing abacavir (ABC) 300 mg, lamivudine (3TC) 150 mg, and zidovudine (ZDV) 300 mg taken ...twice/day, and those of ABC 300 mg twice/day plus a double combination tablet containing 3TC 150 mg and ZDV 300 mg twice/day (ABC‐COM).
Design. Open‐label, crossover study.
Setting. Two hospital‐based clinical research units.
Patients. Twelve men infected with human immunodeficiency virus‐1.
Intervention. Steady‐state pharmacokinetics of ABC, 3TC, and ZDV were assessed after dosing with ABC‐COM and the triple combination tablet.
Measurements and Main Results. Steady‐state pharmacokinetics of ABC, 3TC, and ZDV were similar for the triple combination tablet versus ABC‐COM for the following: geometric mean (GM) area under the serum concentration‐time curve, ABC 6.08 versus 5.87, 3TC 5.51 versus 5.53, and ZDV 1.38 versus 1.46 μg·hr/ml; GM maximum serum concentration (Cmax‐ss), ABC 3.09 versus 3.19, 3TC 1.26 versus 1.40, and ZDV 1.19 versus 1.15 μg/ml; median time to Cmax‐ss, ABC 0.75 versus 0.75, 3TC 1.50 versus 1.24, and ZDV 0.75 versus 0.75 hours; and GM oral clearance, ABC 51 versus 49, 3TC 27 versus 27, and ZDV 217 versus 206 L/hour. The GM half‐lives of ABC and ZDV were similar for both treatments, 1.69 versus 1.58 and 2.30 versus 2.08 hours, respectively.
Conclusion. Steady‐state pharmacokinetics of ABC, 3TC, and ZDV were similar in patients who took them as ABC‐COM or as a triple combination tablet.
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