Yasong Lu

Metabolism-Directed Design of Oxetane-Containing Arylsulfonamide Derivatives as γ-Secretase Inhibitors

A metabolism-based approach toward the optimization of a series of N-arylsulfonamide-based γ-secretase inhibitors is reported. The lead cyclohexyl analogue 6 suffered from extensive oxidation on the cycloalkyl motif by cytochrome P450 3A4, translating into poor human liver microsomal stability. Knowledge of the metabolic pathways of 6 triggered a structure-activity relationship study aimed at lowering lipophilicity through the introduction of polarity. This effort led to several tetrahydropyran and tetrahydrofuran analogues, wherein the 3- and 4-substituted variants exhibited greater microsomal stability relative to their 2-substituted counterparts. Further reduction in lipophilicity led to the potent γ-secretase inhibitor and 3-substituted oxetane 1 with a reduced propensity toward oxidative metabolism, relative to its 2-substituted isomer. The slower rates of metabolism with 3-substituted cyclic ethers most likely originate from reductions in lipophilicity and/or unfavorable CYP active site interactions with the heteroatom. Preliminary animal pharmacology studies with a representative oxetane indicate that the series is generally capable of lowering Aβ in vivo. As such, the study also illustrates the improvement in druglikeness of molecules through the use of the oxetane motif.

Cerebrospinal Fluid Amyloid-β (Aβ) as an Effect Biomarker for Brain Aβ Lowering Verified by Quantitative Preclinical Analyses

Reducing the generation of amyloid-β (Aβ) in the brain via inhibition of β-secretase or inhibition/modulation of γ-secretase has been pursued as a potential disease-modifying treatment for Alzheimers disease. For the discovery and development of β-secretase inhibitors (BACEi), γ-secretase inhibitors (GSI), and γ-secretase modulators (GSM), Aβ in cerebrospinal fluid (CSF) has been presumed to be an effect biomarker for Aβ lowering in the brain. However, this presumption is challenged by the lack of quantitative understanding of the relationship between brain and CSF Aβ lowering. In this study, we strived to elucidate how the intrinsic pharmacokinetic (PK)/pharmacodynamic (PD) relationship for CSF Aβ lowering is related to that for brain Aβ through quantitative modeling of preclinical data for numerous BACEi, GSI, and GSM across multiple species. Our results indicate that the intrinsic PK/PD relationship in CSF is predictive of that in brain, at least in the postulated pharmacologically relevant range, with excellent consistency across mechanisms and species. As such, the validity of CSF Aβ as an effect biomarker for brain Aβ lowering is confirmed preclinically. Meanwhile, we have been able to reproduce the dose-dependent separation between brain and CSF effect profiles using simulations. We further discuss the implications of our findings to drug discovery and development with regard to preclinical PK/PD characterization and clinical prediction of Aβ lowering in the brain.

Quantitative Pharmacokinetic/Pharmacodynamic Analyses Suggest That the 129/SVE Mouse Is a Suitable Preclinical Pharmacology Model for Identifying Small-Molecule γ-Secretase Inhibitors

Alzheimers disease (AD) poses a serious public health threat to the United States. Disease-modifying drugs slowing AD progression are in urgent need, but they are still unavailable. According to the amyloid cascade hypothesis, inhibition of β- or γ-secretase, key enzymes for the production of amyloid β (Aβ), may be viable mechanisms for the treatment of AD. For the discovery of γ-secretase inhibitors (GSIs), the APP-overexpressing Tg2576 mouse has been the preclinical model of choice, in part because of the ease of detection of Aβ species in its brain, plasma, and cerebrospinal fluid (CSF). Some biological observations and practical considerations, however, argue against the use of the Tg2576 mouse. We reasoned that an animal model would be suitable for GSI discovery if the pharmacokinetic (PK)/pharmacodynamic (PD) relationship of a compound for Aβ lowering in this model is predictive of that in human. In this study, we assessed whether the background 129/SVE strain is a suitable preclinical pharmacology model for identifying new GSIs by evaluating the translatability of the intrinsic PK/PD relationships for brain and CSF Aβ across the Tg2576 and 129/SVE mouse and human. Using semimechanistically based PK/PD modeling, our analyses indicated that the intrinsic PK/PD relationship for brain Aβx-42 and CSF Aβx-40 in the 129/SVE mouse is indicative of that for human CSF Aβ. This result, in conjunction with practical considerations, strongly suggests that the 129/SVE mouse is a suitable model for GSI discovery. Concurrently, the necessity and utilities of PK/PD modeling for rational interpretation of Aβ data are established.

A novel BACE inhibitor (PF-05297909): A two-part adaptive design to evaluate safety, pharmacokinetics and pharmacodynamics for modifying beta-amyloid in a first-in-human study

BackgroundThe accumulation of amyloid beta (Aβ) peptides is believed to be a central contributor to the neurodegeneration seen in the Alzheimers disease (AD) brain. Given the central role of Aβ42 in AD pathogenesis, a therapeutic strategy to lower central Aβ42 (and Aβ40) levels via inhibition of BACE was adopted in a first in human trial in a 2-part adaptive design.MethodsPart 1 evaluated PF-05297909 plasma PK and the PK/PD relationship for the reduction of plasma Aβ40, Aβ42 and AβX levels; Part 2 evaluated the exposure-response relationship between PF-05297909 and CSF levels of Aβ40, Aβ42 and AβX. Sufficient safety and tolerability, plasma exposure and reduction in plasma Aβ were necessary to initiate Part 2. Part 1 was a sequential parallel group dose escalation (25, 100, 250 and 325 mg) with n=8 (6:2, active:placebo) healthy volunteers (HV) in each cohort. Part 2 consisted of 3 cohorts of n=8 (6:2, active:placebo) HV. Doses selected for Part 2 started with the highest safe dose in Part 1 and then adapted for subsequent cohorts. The PK/PD relationship between PF-05297909 and Aβ42 was determined using a non-linear mixed effects (NLME) analysis. The doses for Part 2 - cohort 2 and 3 were to be chosen to improve the relative standard error in the estimate of the BACE IC50 as quantified by evaluating the determinant of the Fisher information matrix for the NLME model.ResultsPF-05297909 was well-tolerated. Reduction in plasma Aβ (Aβ40 and Aβ42) was exposure related with an apparent maximum at the 250 mg dose with a greater duration of activity at the 325 mg dose of PF-05297909. A 325 mg dose was selected for Part 2 - cohorts 1 and 2 without further cohorts being run, as stopping criteria for futility were met following analysis of cohort 2. A PK/PD relationship in CSF was not observed.ConclusionsThe adaptive designed PF-05297909 FIH study allowed efficient testing of safety and of the PK/PD relationship between PF-05297909 exposure and Aβ (Aβ40 and Aβ42). PF-05297909 was safe and well tolerated in HV at exposures tested. A robust effect on plasma Aβ did not translate to CSF pharmacodynamic effects.

A Translational Systems Pharmacology Model for Aβ Kinetics in Mouse, Monkey, and Human

A mechanistic model of amyloid beta production, degradation, and distribution was constructed for mouse, monkey, and human, calibrated and externally verified across multiple datasets. Simulations of single‐dose avagacestat treatment demonstrate that the Aβ42 brain inhibition may exceed that in cerebrospinal fluid (CSF). The dose that achieves 50% CSF Aβ40 inhibition for humans (both healthy and with Alzheimers disease (AD)) is about 1 mpk, one order of magnitude lower than for mouse (10 mpk), mainly because of differences in pharmacokinetics. The predicted maximal percent of brain Aβ42 inhibition after single‐dose avagacestat is higher for AD subjects (about 60%) than for healthy individuals (about 45%). The probability of achieving a normal physiological level for Aβ42 in brain (1 nM) during multiple avagacestat dosing can be increased by using a dosing regimen that achieves higher exposure. The proposed model allows prediction of brain pharmacodynamics for different species given differing dosing regimens.

An updated Aβ systems kinetic model for mouse, monkey and human

icity in Alzheimer’s disease (AD) has been focused on sensory deficit, especially impaired olfactory sensibility. AD might provide pathogenic ally significant causes of olfactory dysfunction, which designated even nonfibrillar Ab peptide deposition within olfactory bulb prior to the deposition within any region of brain. In addition, the olfactory receptor neurons (ORNs) might be affected by Ab peptides and contribute to atypical olfactory sensory dysfunction. Therefore, approaches allowing the simple, direct detection of the AD marker within the ORNs in vitro would be beneficial for the assessment of the diseases status. Methods: We suggest a new technical concept that is capable of evaluating as small as a few femtomolar Ab peptides by using a photo-sensitive field-effect transistor (p-FET) integrated with a selectively optical-transmissible filter. Selenium filter has optically adequate properties for the quantum dots for labeling Ab peptides, which reflects the unwanted range (<600 nm) of the excitation light and is only able to transmit the emitted light (>650 nm). The ORNs including Ab peptide s were lysed and placed on the sensing area of the p-FET.Results:We observed destructively decayed photo-current by the selenium-filtered p-FET compared with the bare p-FET, which confirmed that the selenium filter rarely transmit the excitation beam. Thereafter, we measured the photo-currents generated from the transmitted fluorescent beam through the selenium filter when the quantum dots conjugated with Ab on the filter w ere excited at 550 nm and emitted at 655 nm. Compared with no Ab peptide cases, t he difference of the photo-currents with Ab peptide s is as much apparent as about 10 mA. Conclusions: This study showed that with even slightly small amount of Ab peptides the selenium-filtered p-FET is simply applicable to differentiating the optically tenuous fluorescent beam. In addition, the correlation between the p-FET photo-currents and the various amounts of the quantum dots (Ab peptides) represented a linearity which showed a good conceptual agreement with the theoretically induced equation, consequently assuming that the photo-current would potentially evaluate the small quantity of Ab peptides on the p-FET sensing area.

Cross-species analysis of cerebrospinal fluid (CSF) beta-amyloid reductions by the BACE1 inhibitor PF-05297909 indicates species differences in PK/PD relationships: Relevance to clinical translation

reversed Ab oligomer-damaged long-term potentiation (LTP) at concentrations that did not interfere normal high frequency stimulation-induced LTP. Moreover, bis(heptyl)-cognitin prevented Ab oligomer-induced reduction of neurite length and synaptic quantity in mature hippocampal neurons. In contrast, tacrine could not reverse synaptic impairments in these models. Under oligomerization condition, bis(heptyl)-cognitin reduced the amount of Ab oligomer as evidenced by dot blot assay and immunoblot analysis. Finally, bis(heptyl)-cognitin was shown to alter Ab self-assembling as demonstrated by circular dichroism spectroscopy and transmission electron microscopy. Conclusions: All these results not only offer a modality as to how dimeric agents protect against Ab oligomer-induced synaptic impairments, but also offers a strong support for the beneficial therapeutic effects of bis(heptyl)-cognitin in the treatment of AD.

Understanding the target and evaluating the consequences of gamma-secretase modulation from in vitro models to higher-order species

P4-298 UNDERSTANDING THE TARGETAND EVALUATING THE CONSEQUENCES OF GAMMASECRETASE MODULATION FROM IN VITRO MODELS TO HIGHER-ORDER SPECIES Kathleen Wood, Nikolay Pozdnyakov, Emily Sylvain, Michael Marconi, Ashley Robshaw, Leslie Pustilnik, Stefanus Steyn, Patrick Trapa, Yasong Lu, Douglas Johnson, Martin Pettersson, Kelly Bales, Pfizer Inc., Cambridge, Massachusetts, United States; Pfizer, Cambridge, Massachusetts, United States; Pfizer, Groton, Connecticut, United States; Pfizer Inc., Cambridge, Massachusetts, United States; Pfizer Inc., Groton, Connecticut, United States. Contact e-mail: kathleen.m.wood@pfizer.com

Gamma-secretase modulators demonstrate similar exposure-response relationship in the rat and guinea pig

Background: Compounds that modulate g -secretase (GSMs) activity to selectively lower brain Ab42 have been pursued as potential therapies for individuals with Alzheimer’s disease. The Ab sequence in humans and guinea pigs is identical but differs from that in rat. To probe the pharmacodynamic (PD) effects of GSMs, the guinea pig seems to be a reasonable model; however, rat is the preferred species since pharmacokinetic (PK) and safety characterization is conducted in the rat. Therefore, it is critical to understand whether the exposure-response relationship of GSMs vary between the rat and guinea pig. Methods:We completed comprehensive time-course studies for measurement of compound concentrations and Ab levels in the plasma, cerebrospinal fluid (CSF), and brain in the rat and guinea pig. Each species was administered with two structurally distinct GSMs, GSM67 and GSM09. The data enabled a PK/PDmodel, which accounts for the synthesis as well as clearance of Ab, to derive an intrinsic PK/PD relationship that was devoid of confounds from PK or Ab turnover. Results: GSM67 and GSM09 significantly reduced Ab42 in a dose-dependent fashion in the plasma, CSF, and brain in the rat and guinea pig. For all three compartments, the intrinsic PK/PD relationship for Ab42 lowering was similar between these two species. Conclusions: Our quantitative PK/PD analysis support utilizing the rat as a suitable model for preclinical profiling of GSMs. Moreover, the quantitative approach described here can be used for further assessment of PK/PD translatability across species.

Abeta systems pharmacology: An initial model for wild type mouse, monkey, and human

P4-296 ABETA SYSTEMS PHARMACOLOGY: AN INITIAL MODEL FORWILD TYPE MOUSE, MONKEY, AND HUMAN Hugh Barton, Ellen Wang, Yasong Lu, Alina Efremenko, Nikki Etxgoien, Harvey Clewell, Melvin Andersen, Jerry Campbell, Timothy Nicholas, Pfizer, Groton, Connecticut, United States; Pfizer, New York, New York, United States; 3 The Hamner Institutes for Health Sciences, Research Triangle Park, North Carolina, United States.