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Simplifying Pharmacokinetics, Applying it to Drug and Dosage Form Development, and Making Drug Dosage Decisions in Clinical Medicine: The Adaptation of Kirchhoff’s Laws from Physics
No full textOver the past three years, we have published a series of nine manuscripts demonstrating that all relevant pharmacokinetic relationships may be simply derived independent of differential equations, offering an alternative to traditional pharmacokinetic analyses. These derivations are based on an understanding of parallel and in series rate-defining processes, and account for all relevant drivers, including organ blood flow and drug delivery clearance kinetics, across both linear and nonlinear scenarios. In this tutorial, we present the simple derivation of renal clearance and hepatic clearance directly relevant to clinical pharmacokinetics, as applied to making drug dosing decisions based on measures of systemic exposure. We further advocate for a more streamlined and practical approach to teaching and applying clinical pharmacokinetics, noting that compartmental modeling, protein binding in hypothetical compartments, trapezoidal AUC calculations, and alternative volume of distribution parameters, aside from (the unfortunately misnamed) volume of distribution steady-state, often overcomplicate pharmacokinetics in practice. The key advantage of this simplified methodology is the ability to directly incorporate clearance from the drug delivery site into systemic pharmacokinetic relationships. This enables a clear understanding of how entering clearance can influence systemic AUC, helping explain: enhanced pharmacodynamic outcomes of slow drug delivery versus immediate-release formulations; systemic bioavailability measures exceeding unity, statistically significant discrepancies between urinary and systemic bioavailability measures; and changes in renal clearance as a function of drug clearance from the delivery site. These key concepts are illustrated by applying the proposed methodology to an example drug, analyzing all relevant clinical pharmacokinetic relationships required for dosing decisions.Graphical Abstrac
Current challenges and opportunities in active and passive data collection for mobile health sensing: a scoping review
Get PDFObjectiveMobile and ubiquitous devices enable health data collection "in a free-living environment" to support applications such as remote patient monitoring and adaptive digital interventions using machine learning (ML). Despite their potential, significant data collection challenges persist, including issues related to user compliance with reporting data, passive data consistency, and authorization. This scoping review identifies and analyzes these challenges, focusing on barriers to effective data collection.Materials and methodsWe searched IEEE, ACM, and Web of Science for papers involving training ML models using both active and passive mobile sensing. We used the following search terms: "mobile OR ubiquitous", "EMA", "health", "passive", and "deep learning OR machine learning". We only included papers that collected both passive and active data and excluded papers that used a pre-existing dataset.ResultsA total of 77 studies met the inclusion criteria. These studies utilized smartphones, smartwatches, wearable devices, and environmental sensors for data collection. Several studies reported challenges with participant compliance in active data collection, while passive data collection faced data consistency and authorization issues. Efforts to address these challenges were documented in some but not all studies. Using this information, we outline current challenges and corresponding opportunities for data collection in mobile sensing studies.DiscussionML techniques can reduce participant burden in active data collection by optimizing prompt timing, auto-filling responses, and minimizing prompt frequency. Simplified interfaces such as user-friendly smartwatch prompts can further improve compliance. For passive data collection, techniques such as optimization of recording times to preserve battery life and motivational techniques to encourage proper device use can increase data consistency.ConclusionMobile sensing offers opportunities for developing intelligent mobile health applications but faces data collection challenges with respect to factors such as compliance, consistency, and authorization. Innovations in ML and user interface design show promise for addressing these barriers
Long-Distance Friendship in Roman Letters
No full textThis dissertation examines letters as a site for the maintenance of long-distance friendship through four case studies of letter-writers spanning both poetry and prose as well as classical Latin through late antiquity: Cicero, Ovid, Fronto, and Paulinus. My first chapter argues that in letters, geographical distance can function as a cipher for other forms of distance and difference, allowing correspondents to both paper over and deemphasize inequalities of age, status, wealth, religion, commitment to the friendship, and more. Ausonius and Paulinus, for example, use the physical space between Paulinus’s family estates in Spain and Ausonius’s home in Bordeaux to talk around power differentials (Ausonius is forty years older and taught Paulinus when he was a young boy) and, perhaps most significantly, the growing chasm between their religious beliefs as Paulinus becomes more devoutly Christian. My second chapter observes that while intimacy in letters is often expressed through anticipation of physical reunion, in many cases the letter is more a site of nostalgia than of anticipation. Long-distance friends may or may not look forward to an upcoming reunion, but they frequently mythologize and look back on an idealized version of their past in-person friendship. The letter, I argue, allows fantasy to substitute for actual co-presence. Finally, my third chapter examines how the asynchronicity of letters shapes the kinds of intimacy they can foster. Ovid laments that it can take a full year for him to receive a letter in Tomis from Rome, so when he learns that Gallio’s wife has died, the asynchronicity of the letter fundamentally changes the kind of friend he can be. Such a belated consolation, he frets, could simply resurface old wounds—or Gallio could have remarried, dooming Ovid to offend the new couple by sending condolences instead of congratulations. Even in cases where the gap between written and lived time is only a matter of weeks or days, this untimeliness can create a sense of freedom that leads to greater candor, but it can also allow correspondents to avoid or ignore one another more effectively than they could in person. Once again, then, the letter becomes a way of talking about and around other forms of distance; temporal distance can exacerbate other gaps that exist between correspondents
Synthetic Nanocarrier Systems for the Intracellular Delivery of Large Cargoes to Enable Genomic Integration of Whole Genes and the Treatment of Genetic Diseases and Cancers
No full textWe report the engineering of an integrated cargo-carrier system for gene therapy that leverages lipid nanoparticles (LNPs) to transport CRISPR/Cas9 payloads, including double-stranded DNA (dsDNA) donor templates, designed for homology directed repair (HDR)-mediated site-specific insertion of the cystic fibrosis transmembrane conductance regulator (CFTR) gene to correct cystic fibrosis (CF) in diseased airway epithelium. We screened a panel of single guide RNAs (sgRNAs) targeting the 5’ untranslated region (5’UTR) of endogenous CFTR, selecting constructs according to their cutting efficiencies and off-target activity in human bronchial epithelial cells (16HBE). We furthermore optimized DNA donor templates according to homology arm length, 5’ modification, and codon optimization strategy to maximize allelic integration efficiencies and functional activity of the integrated gene. In parallel, we screened various nanoparticle formulations, adjusting ratios of Cas9-encoding mRNA, single guide RNAs (sgRNAs), and dsDNA donor templates to optimize gene editing using16HBE cells harboring a CF-causing mutation (G542X). Populations of G542X cells edited via LNP delivery of CFTR donors achieved 3 – 3.5% gene integration and yielded comparable CFTR protein expression compared to normal 16HBE14o- controls. These edited populations exhibit restoration of CFTR-dependent Cl- current to ca. 80% of values measured in normal 16HBE14o- cell monolayers. This LNP platform adds capabilities for transporting large gene editing machinery to airway epithelial cells for genomic integration of entire genes, enabling therapeutic solutions that achieve correction of any CF-causing mutation. In preliminary steps toward clinical translation of this LNP platform, we have assessed in vitro models of intravenous administration of LNPs in CFTR inhibited tissues. To this end, we have applied models of CFTR-inhibited endothelium to evaluate the interplay between CFTR inhibition and LNP exposure on endothelial barrier integrity. Transcriptomic and phosphoproteomic analyses of human umbilical vein endothelial cells (HUVECs) exposed to either CFTR inhibitor 172 (CFTRinh-172), LNPs, or the two in combination (Combo) implicate a Rho GTPase-mediated mechanism for a transient loss in endothelial barrier integrity under Combo conditions. Elucidating the synergistic impacts of commonly used CFTRinh-172 and LNPs on endothelium is critical to interpreting data that may be produced in studies that apply the two together. Taken together, the tools developed in this dissertation lay the groundwork for safe, effective, and mutation-agnostic translation of nanoparticle-based platforms to treat cystic fibrosis. The modularity of these tools enables their facile adaptation for alternative disease targets such as cancer, ultimately redefining the landscape of translatable gene therapies
Enhancing Accuracy and Plausibility of Unsupervised Deep-Learning-Based Deformable Image Registration with Registration-Specific Designs and Anatomical Priors
Get PDFDeformable image registration (DIR) is the computational process of aligning different images to a unified coordinate system through locally varying, non-linear (hence deformable) displacement fields. It is fundamental to many medical image analysis workflows where images from different time points, modalities, or patients need to be compared after spatial alignment. Conventional optimization-based DIR methods have seen limited clinical adoption due to persistent challenges in accuracy and efficiency. Recently, deep-learning-based deformable image registration (DL-DIR) has attracted interest for its speed and ability to leverage rich, data-driven feature representation. Unsupervised and weakly supervised approaches have become the main paradigms of DL-DIR.DIR faces three core challenges: the absence of ground truth, the problem’s inherent ill-posedness, and its non-convexity. These intrinsic limitations not only complicate the evaluation of registration accuracy but also hinder the assessment of deformation plausibility. The most common accuracy surrogate for DIR is segmentation label matching (e.g., Dice score) due to its wide availability. As such, weak supervision via contour matching losses becomes popular to boost the Dice metric. Despite improving the apparent accuracy, this strategy often leads to unrealistic deformations and compromises generalizability, especially when applied to datasets with different labeling protocols. Consequently, unsupervised DL-DIR has regained prominence. However, current literature lacks fair, standardized comparisons among unsupervised DL-DIR methods, and offers little consensus on best practices for unsupervised DL-DIR. It is even unclear whether unsupervised DL-DIR truly outperforms conventional methods, and if so, what drives that advantage. Moreover, most DL-DIR methods remain intensity-driven with only simple smoothness regularization, neglecting rich physiological priors in human anatomy. This oversight not only limits accuracy but can yield anatomically implausible deformation fields, significantly hindering clinical translation.
To address these challenges, we propose three specific aims. We first conduct a comprehensive ablation-type study to identify the true drivers for accurate unsupervised DL-DIR, which highlights the importance of registration-specific designs. Building upon that, we then tailor DL-DIR to two radiation-therapy applications at two distinct anatomical sites, enhancing accuracy and anatomical plausibility by integrating relevant anatomical information into the registration framework. The three aims are:
Specific Aim 1: Enhance accuracy of unsupervised DL-DIR with registration-specific designs. We hypothesize that registration-specific designs such as the multi-resolution pyramid, correlation calculation, and inverse-consistency constraints are more important than complex network architectures. We propose a comprehensive ablation-type study to identify the key modules for unsupervised mono-modal DL-DIR and demonstrate that simple models, when properly equipped with these design choices, can achieve state-of-the-art performance.
Specific Aim 2: Improve bladder trigone MRI registration by integrating anatomical context through multi-task learning. We hypothesize that the features learned in landmark prediction and segmentation tasks will improve registration accuracy. We propose a multi-task learning framework for joint landmark regression, segmentation, and deformation registration for the bladder trigone and demonstrate improved registration accuracy.
Specific Aim 3: Improve accuracy and plausibility in head-and-neck CT registration using a MUsculo-Skeleton-Aware (MUSA) framework. We hypothesize registration of the complex and heterogeneous head-and-neck deformations can benefit from distinguishing the tissue types and motion types. We propose to decompose such complex deformations into bulk posture changes and residual fine deformations and distinguish between the rigidity of bone structures and the flexibility of soft tissues. With such anatomical knowledge incorporated into the DIR optimization process, we demonstrate both enhanced accuracy and markedly better plausibility of deformation fields
After the Final Bell: A Critical Ethnography with a Youth-Serving Community-Based Education Non-Profit
No full textOut-of-school time (OST) educational non-profits can offer positive and affirming programming to students who attend chronically underfunded public schools. Despite this, OST non-profits, similarly to public schools, have been impacted by the privatized restructuring and reform of the past several decades. One striking difference, though, is the impact of OST non-profits’ reliance on the philanthropy of individuals, foundations, and corporations to run their programming. This critical and participatory ethnography investigates an OST non-profit from the top-down and the bottom-up to illuminate the complex racialized systems and structures that sustain and limit these organizations, and highlights the power of participatory methods to transform scholarship, communities, and individuals. The questions guiding my study are: 1) How do youth-serving community-based non-profits manage the tension between private philanthropy and the needs of their students?
2) How do organizational stakeholders define success? Using critical theory I find three overlapping tensions facing the non-profit of focus. First, I capture a tension of measuring success—how wealthy actors exert their definitions of success that are defined by a desire for easy-to-digest quantifiable data points rather than built on the collective or collaborative ideas of the on-the-ground stakeholders. Second, I introduce how the pressure of expansion influences the organization to grow beyond its community-based mission in order to be more visible to white elite donors. Finally, I demonstrate a tension of performance—how the organization performs for its donors, and how its donors perform for the organization. Drawing on rich ethnographic data, I demonstrate how power is reproduced to those at the top of a structurally and socially designed hierarchy—where elite donors and board members, defined by whiteness and capital, influence and transform the non-profit.This dissertation highlights an overlooked educational learning and developmental site while capturing how the privatized restructuring of public education has infiltrated the OST non-profit landscape. This research deepens scholarship, practice, and policy on democratic goods, privatization, the non-profit sector, and philanthropy. Finally, my participatory methods disrupt conventional ethnography by collectively building towards more justice and solidarity as defined by community partners themselves
Modeling gene regulatory logic in the innate immune response
No full textThis thesis highlights the regulation of the interferon β (IFNβ) enhancer by NFκB and IRF as well as epigenomic changes driven by NFκB and IRF. Precise regulation of the immune cytokine IFNβ is essential for human health. Classic studies established that the transcription factors NFκB and IRF function synergistically in activating IFNβ expression. The regulation of IFNβ promoter activity has been a touchpoint of mammalian gene control research since the discovery of functional synergy between two stimulus-responsive transcription factors (TFs) nuclear factor kappa B (NFκB) and interferon regulatory factors (IRF). However, subsequent gene knockout studies revealed that this synergy is condition-dependent such that either NFκB or IRF activation can be dispensable, leaving the precise regulatory logic of IFNβ transcription an open question.In Chapter 2, we develop a series of quantitative enhancer states models of IFNβ expression control and evaluate them with stimulus-response data from TF knockouts. Our analysis confirmed that TF synergy is a hallmark of the regulatory logic but that it need not involve NFκB, as synergy between two adjacent IRF dimers is sufficient.
In Chapter 3, we evaluate the stimulus specificity of the IFNβ enhancer. We found that a sigmoidal binding curve at the distal site renders the dual IRF synergy mode ultrasensitive, allowing it only in conditions of high IRF activity upon viral infection. In contrast, the proximal site has high affinity and enables expression in response to bacterial exposure through synergy with NFκB. However, its accessibility is controlled by the competitive repressor p50:p50, which prevents basal IRF levels from synergizing with NFκB, such that NFκB-only stimuli do not activate IFNβ expression. The enhancer states model identifies two commonly used modes of synergy that are accessed differentially in response to different immune threats, enabling a highly stimulus-specific but also versatile regulatory logic for stimulus-specific IFNβ expression.
In Chapter 4, we investigate mechanisms of enhancer formation driven by NFκB and IRF. In response to specific pathogens, the NFκB pathway and the IRF pathway modify the chromatin state and epigenome, increasing chromatin accessibility and depositing new transcriptional enhancer marks. The mechanisms by which this epigenomic remodeling occurs remains unclear. It has been thought that only a distinct subset of transcription factors, called “pioneer factors,” can bind to and open condensed chromatin. However, recent evidence has shown that NFκB can bind to nucleosomal DNA and displace histone-DNA contacts. We test the hypothesis that NFκB increases chromatin accessibility solely through its DNA binding activity, without requiring additional proteins to be recruited by its transcriptional activation domain. We additionally profile the dynamic enhancer formation process and find evidence for different mechanisms involving various histone methyltransferases, histone acetyltransferases, RNA polymerase, and other cofactors
Palladium-Catalyzed Reactions of Cyclic Allenes and Strain-Promoted Reactions of Cyclic 1,2,3-Trienes
No full textThis dissertation describes the development of methodologies that engage strained cyclic intermediates in complexity-generating reactions. One major effort involves the transition metal-catalyzed interception of strained cyclic allenes, which has been accomplished using palladium catalysis. Additionally, the study of alternative minimally explored strained intermediates including cyclic 1,2,3-trienes and heterocyclic 1,2,3-trienes are reported. These studies contribute to fundamental understanding of structure and reactivity of transient strained compounds and give rise to polycyclic products. Computational studies relating to the mechanism of strained intermediate generation from Kobayashi precursors are also reported. Chapters one and two are related to the development of palladium-catalyzed reactions of strained cyclic allenes. Chapter one describes the development of a modular annulation reaction of strained allenes and arylpalladium species. This methodology employs aryl halides and cyclic allene precursors to generate fused heterocyclic products via the formation of two new bonds and a new center. Chapter two described the development of a catalyst-controlled annulation reaction of strained allenes and -allylpalladium species. This methodology employs vinyl benzoxazinones and cyclic allenes precursors to generate two isomeric products with high selectivity based on the ligand employed. The development of these palladium-catalyzed reactions demonstrates that despite their high reactivity and short lifetimes, strained cyclic allenes efficiently engaged in catalytic processes, to access complex products, including examples with absolute stereocontrol.
Chapter three describes the development of strained 1,2,3-cyclohexatrienes, which have remained underexplored historically, as synthetic building blocks. Studies of the reactivity of the unsubstituted 1,2,3-cyclohexatriene, as well as its mono- and disubstituted derivatives are reported, drastically expanding the scope of reactions known for such intermediates. Combined computational and experimental studies elucidate the factors controlling regioselectivity in reactions of an unsymmetrical strained triene. Furthermore, the potential utility of strained trienes in rapidly generating complex scaffolds is demonstrated through the integration of triene trapping reactions into multistep synthetic sequences to access polycyclic products. These studies highlight the potential of these traditionally avoided species for broader use in synthetic chemistry.
Chapter four details the study of six-membered heterocyclic 1,2,3-trienes, particularly 4,5-didehydropyridones. Computational studies of the structure of such species, as well as the development of a synthetic route to access precursors to the same, are reported. Scope studies demonstrate the utility of six-membered azacyclic 1,2,3-trienes for accessing complex nitrogen-containing heterocycles, and trends in the regioselectivity observed therein are explored through computational studies. Collectively, this study demonstrates the value of six-membered azacyclic 1,2,3-trienes, a previously unexplored class of strained cyclic intermediates, in heterocycle synthesis, while pushing the limits of strained intermediate chemistry.
Chapter five describes the reaction mechanisms of the fluoride-mediated generation of selected strained intermediates from Kobayashi precursors. We interrogate several mechanistic aspects using Density Functional Theory (DFT) calculations and find that the eliminations to form alkynes and alkenes can take place through primarily two different mechanisms. This study is one of the few theoretical studies on Kobayashi eliminations for the generation of strained intermediates. It is anticipated that this report will enable the rational design of new strained intermediate precursors for future uses in synthesis
Condensates as a Culprit in RAS Activation and Inhibitor Resistance.
Get PDFTherapy resistance is a significant cause of death in patients treated with targeted cancer therapy in diverse oncogene-driven cancers. A better understanding of resistance mechanisms can lay the foundation for improving existing and developing new therapies. A recent elegant study published in Nature Chemical Biology sheds light on a new resistance mechanism. The authors define a novel role for ARAF, a member of the RAF protein family (ARAF, BRAF, and CRAF), that is distinct from its previously understood role as a RAS effector and MEK protein kinase in the MAPK pathway. They describe how ARAF sequesters active RAS at the plasma membrane in phase-separated condensates to sustain signaling and prevent inactivation by the RAS GTPase-activating protein neurofibromin 1. This study underscores emerging roles for biomolecular condensates in cancer and highlights important implications for disrupting protein condensates to address treatment resistance to RAS (and RAS pathway)-targeted therapies. The study also illuminates evolutionary functional distinction between the RAF proteins and indicates unique biology for ARAF in normal physiology and disease