Cascade Mesophase Transitions of Multi-enzyme Responsive Polymeric Formulations.

Studying how synthetic polymer assemblies respond to sequential enzymatic stimuli can uncover intricate interactions in biological systems. Using amidase- and esterase-responsive PEG-based diblock (DBA) and triblock amphiphiles (TBAs), we created two distinct formulations: amidase-responsive DBA with esterase-responsive TBA and vice versa. We studied their cascade responses to the two enzymes and the sequence of their introduction. These formulations underwent cascade mesophase transitions upon the addition of the DBA-degrading enzyme, transitioning from (i) coassembled micelles to (ii) triblock-based hydrogel, and ultimately to (iii) dissolved polymers when exposed to the TBA hydrolyzing enzyme. The specific pathway of the two mesophase transitions depended on the compositions of the formulations and the enzyme introduction sequence. The results highlight the potential for designing polymeric formulations with programmable multistep enzymatic responses, mimicking the complex behavior of biological macromolecules.

Hydrogel Microneedles with Programmed Mesophase Transitions for Controlled Drug Delivery.

Microneedle-based drug delivery offers an attractive and minimally invasive administration route to deliver therapeutic agents through the skin by bypassing the stratum corneum, the main skin barrier. Recently, hydrogel-based microneedles have gained prominence for their exceptional ability to precisely control the release of their drug cargo. In this study, we investigated the feasibility of fabricating microneedles from triblock amphiphiles with linear poly(ethylene glycol) (PEG) as the hydrophilic middle block and two dendritic side-blocks with enzyme-cleavable hydrophobic end-groups. Due to the poor formation and brittleness of microneedles made from the neat amphiphile, we added a sodium alginate base layer and tested different polymeric excipients to enhance the mechanical strength of the microneedles. Following optimization, microneedles based on triblock amphiphiles were successfully fabricated and exhibited favorable insertion efficiency and low height reduction percentage when tested in Parafilm as a skin-simulant model. When tested against static forces ranging from 50 to 1000 g (4.9-98 mN/needle), the microneedles showed adequate mechanical strength with no fractures or broken segments. In buffer solution, the solid microneedles swelled into a hydrogel within about 30 s, followed by their rapid disintegration into small hydrogel particles. These hydrogel particles could undergo slow enzymatic degradation to soluble polymers. In vitro release study of dexamethasone (DEX), as a steroid model drug, showed first-order drug release, with 90% released within 6 days. Eventually, DEX-loaded MNs were subjected to an insertion test using chicken skin and showed full penetration. This study demonstrates the feasibility of programming hydrogel-forming microneedles to undergo several mesophase transitions and their potential application as a delivery system for self-administration, increased patient compliance, improved efficacy, and sustained drug release.

Reaching the Tumor: Mobility of Polymeric Micelles Inside an In Vitro Tumor-on-a-Chip Model with Dual ECM.

Degradable polymeric micelles are promising drug delivery systems due to their hydrophobic core and responsive design. When applying micellar nanocarriers for tumor delivery, one of the bottlenecks encountered in vivo is the tumor tissue barrier: crossing the dense mesh of cells and the extracellular matrix (ECM). Sometimes overlooked, the extracellular matrix can trap nanoformulations based on charge, size, and hydrophobicity. Here, we used a simple design of a microfluidic chip with two types of ECM and MCF7 spheroids to allow "high-throughput" screening of the interactions between biological interfaces and polymeric micelles. To demonstrate the applicability of the chip, a small library of fluorescently labeled polymeric micelles varying in their hydrophilic shell and hydrophobic core forming blocks was studied. Three widely used hydrophilic shells were tested and compared, namely, poly(ethylene glycol), poly(2-ethyl-2-oxazoline), and poly(acrylic acid), along with two enzymatically degradable dendritic hydrophobic cores (based on hexyl or nonyl end groups). Using ratiometric imaging of unimer:micelle fluorescence and FRAP inside the chip model, we obtained the local assembly state and dynamics inside the chip. Notably, we observed different micelle behaviors in the basal lamina ECM, from avoidance of the ECM structure to binding of the poly(acrylic acid) formulations. Binding to the basal lamina correlated with higher uptake into MCF7 spheroids. Overall, we proposed a simple microfluidic chip containing dual ECM and spheroids for the assessment of the interactions of polymeric nanocarriers with biological interfaces and evaluating nanoformulations' capacity to cross the tumor tissue barrier.

Polymeric architecture as a tool for controlling the reactivity of palladium(II) loaded nanoreactors.

Self-assembled systems, like polymeric micelles, have become great facilitators for conducting organic reactions in aqueous media due to their broad potential applications in green chemistry and biomedical applications. Massive strides have been taken to improve the reaction scope of such systems, enabling them to perform bioorthogonal reactions for prodrug therapy. Considering these significant advancements, we sought to study the relationships between the architecture of the amphiphiles and the reactivity of their PdII loaded micellar nanoreactors in conducting depropargylation reactions. Towards this goal, we designed and synthesized a series of isomeric polyethylene glycol (PEG)-dendron amphiphiles with different dendritic architectures but with an identical degree of hydrophobicity and hydrophilic to lipophilic balance (HLB). We observed that the dendritic architecture, which serves as the main binding site for the PdII ions, has greater influence on the reactivity than the hydrophobicity of the dendron. These trends remained constant for two different propargyl caged substrates, validating the obtained results. Density functional theory (DFT) calculations of simplified models of the dendritic blocks revealed the different binding modes of the various dendritic architectures to PdII ions, which could explain the observed differences in the reactivity of the nanoreactors with different dendritic architectures. Our results demonstrate how tuning the internal architecture of the amphiphiles by changing the orientation of the chelating moieties can be used as a tool for controlling the reactivity of PdII loaded nanoreactors.

Architecture-Based Programming of Polymeric Micelles to Undergo Sequential Mesophase Transitions.

Di- and triblock amphiphiles can form different mesophases ranging from micelles to hydrogels depending on their chemical structures, hydrophilic to hydrophobic ratios, and their ratio in the mixture. In addition, their different architectures dictate their exchange rate between the assembled and unimer states and consequently affect their responsiveness toward enzymatic degradation. Here we report the utilization of the different reactivities of di- and triblock amphiphiles, having exactly the same hydrophilic to lipophilic balance, toward enzymatic degradation as a tool for programming formulations to undergo sequential enzymatically induced transitions from (i) micelles to (ii) hydrogel and finally to (iii) dissolved polymers. We show that the rate of transition between the mesophases can be programmed by changing the ratio of the amphiphiles in the formulation, and that the hydrogels can maintain encapsulated cargo, which was loaded into the micelles. The reported results demonstrate the ability of molecular architecture to serve as a tool for programming smart formulations to adopt different structures and functions.

Self-Assembly of Tunable Intrinsically Disordered Peptide Amphiphiles.

Intrinsically disordered peptide amphiphiles (IDPAs) present a novel class of synthetic conjugates that consist of short hydrophilic polypeptides anchored to hydrocarbon chains. These hybrid polymer-lipid block constructs spontaneously self-assemble into dispersed nanoscopic aggregates or ordered mesophases in aqueous solution due to hydrophobic interactions. Yet, the possible sequence variations and their influence on the self-assembly structures are vast and have hardly been explored. Here, we measure the nanoscopic self-assembled structures of four IDPA systems that differ by their amino acid sequence. We show that permutations in the charge pattern along the sequence remarkably alter the headgroup conformation and consequently alter the pH-triggered phase transitions between spherical, cylindrical micelles and hexagonal condensed phases. We demonstrate that even a single amino acid mutation is sufficient to tune structural transitions in the condensed IDPA mesophases, while peptide conformations remain unfolded and disordered. Furthermore, alteration of the peptide sequence can render IDPAs to become susceptible to enzymatic cleavage and induce enzymatically activated phase transitions. These results hold great potential for embedding multiple functionalities into lipid nanoparticle delivery systems by incorporating IDPAs with the desired properties.

Stimuli-Induced Architectural Transition as a Tool for Controlling the Enzymatic Degradability of Polymeric Micelles.

Enzyme-responsive polymeric micelles hold great potential as drug delivery systems due to the overexpression of disease-associated enzymes. To achieve selective and efficient delivery of their therapeutic cargo, micelles need to be highly stable and yet disassemble when encountering their activating enzyme at the target site. However, increased micellar stability is accompanied by a drastic decrease in enzymatic degradability. The need to balance between stability and enzymatic degradation has severely limited the therapeutic applicability of enzyme-responsive nanocarriers. Here, we report a general modular approach for designing stable enzyme-responsive micelles whose enzymatic degradation can be enhanced on demand. The control over their response to the activating enzyme is achieved by stimuli-induced splitting of triblock amphiphiles into two identical diblock amphiphiles, which have the same hydrophilic-lipophilic balance as the parent amphiphile. This architectural transition drastically affects the micelle-unimer equilibrium and therefore increases the sensitivity of the micelles toward enzymatic degradation. As a proof of concept, we designed UV- and reduction-activated splitting mechanisms, demonstrating the ability to use architectural transition as a tool for tuning amphiphile-protein interactions, providing a general solution toward overcoming the stability-degradability barrier for enzyme-responsive nanocarriers.

Dendron-Polymer Hybrids as Tailorable Responsive Coronae of Single-Walled Carbon Nanotubes.

Functional composite materials that can change their spectral properties in response to external stimuli have a plethora of applications in fields ranging from sensors to biomedical imaging. One of the most promising types of materials used to design spectrally active composites are fluorescent single-walled carbon nanotubes (SWCNTs), noncovalently functionalized by synthetic amphiphilic polymers. These coated SWCNTs can exhibit modulations in their fluorescence spectra in response to interactions with target analytes. Hence, identifying new amphiphiles with interchangeable building blocks that can form individual coronae around the SWCNTs and can be tailored for a specific application is of great interest. This study presents highly modular amphiphilic polymer-dendron hybrids, composed of hydrophobic dendrons and hydrophilic polyethylene glycol (PEG) that can be synthesized with a high degree of structural freedom, for suspending SWCNTs in aqueous solution. Taking advantage of the high molecular precision of these PEG-dendrons, we show that precise differences in the chemical structure of the hydrophobic end groups of the dendrons can be used to control the interactions of the amphiphiles with the SWCNT surface. These interactions can be directly related to differences in the intrinsic near-infrared fluorescence emission of the various chiralities in a SWCNT sample. Utilizing the susceptibility of the PEG-dendrons toward enzymatic degradation, we demonstrate the ability to monitor enzymatic activity through changes in the SWCNT fluorescent signal. These findings pave the way for a rational design of functional SWCNTs, which can be used for optical sensing of enzymatic activity in the near-infrared spectral range.

Order from Disorder with Intrinsically Disordered Peptide Amphiphiles.

Amphiphilic molecules and their self-assembled structures have long been the target of extensive research due to their potential applications in fields ranging from materials design to biomedical and cosmetic applications. Increasing demands for functional complexity have been met with challenges in biochemical engineering, driving researchers to innovate in the design of new amphiphiles. An emerging class of molecules, namely, peptide amphiphiles, combines key advantages and circumvents some of the disadvantages of conventional phospholipids and block copolymers. Herein, we present new peptide amphiphiles composed of an intrinsically disordered peptide conjugated to two variants of hydrophobic dendritic domains. These molecules, termed intrinsically disordered peptide amphiphiles (IDPA), exhibit a sharp pH-induced micellar phase-transition from low-dispersity spheres to extremely elongated worm-like micelles. We present an experimental characterization of the transition and propose a theoretical model to describe the pH-response. We also present the potential of the shape transition to serve as a mechanism for the design of a cargo hold-and-release application. Such amphiphilic systems demonstrate the power of tailoring the interactions between disordered peptides for various stimuli-responsive biomedical applications.

Using High Molecular Precision to Study Enzymatically Induced Disassembly of Polymeric Nanocarriers: Direct Enzymatic Activation or Equilibrium-Based Degradation?

Enzyme-responsive polymers and their assemblies offer great potential to serve as key materials for the design of drug delivery systems and other biomedical applications. However, the utilization of enzymes to trigger the disassembly of polymeric amphiphiles, such as micelles, also suffers from the limited accessibility of the enzyme to moieties that are hidden inside the assembled structures. In this Perspective, we will discuss examples for the utilization of high molecular precision that dendritic structures offer to study the enzymatic degradation of polymeric amphiphiles with high resolution. Up to date, several different amphiphilic systems based on dendritic blocks have all shown that small changes in the hydrophobicity and amphiphilicity strongly affected the degree and rate of enzymatic degradation. The ability to observe the huge effects due to relatively small variations in the molecular structure of polymers can explain the limited enzymatic degradation that is often observed for many reported polymeric assemblies. The observed trends imply that the enzymes cannot reach the hydrophobic core of the micelles, and instead, they gain access to the amphiphiles by the unimer-micelle equilibrium, making the unimer exchange rate a key parameter in tuning the enzymatic degradation rate. Several approaches that are aimed at overcoming the stability-responsiveness challenge are discussed as they open the way to the design of stable and yet enzymatically responsive polymeric nanocarriers.

Real-Time Ratiometric Imaging of Micelles Assembly State in a Microfluidic Cancer-on-a-Chip.

The performance of supramolecular nanocarriers as drug delivery systems depends on their stability in the complex and dynamic biological media. After administration, nanocarriers are challenged by physiological barriers such as shear stress and proteins present in blood, endothelial wall, extracellular matrix, and eventually cancer cell membrane. While early disassembly will result in a premature drug release, extreme stability of the nanocarriers can lead to poor drug release and low efficiency. Therefore, comprehensive understanding of the stability and assembly state of supramolecular carriers in each stage of delivery is the key factor for the rational design of these systems. One of the main challenges is that current 2D in vitro models do not provide exhaustive information, as they fail to recapitulate the 3D tumor microenvironment. This deficiency in the 2D model complexity is the main reason for the differences observed in vivo when testing the performance of supramolecular nanocarriers. Herein, we present a real-time monitoring study of self-assembled micelles stability and extravasation, combining spectral confocal microscopy and a microfluidic cancer-on-a-chip. The combination of advanced imaging and a reliable 3D model allows tracking of micelle disassembly by following the spectral properties of the amphiphiles in space and time during the crucial steps of drug delivery. The spectrally active micelles were introduced under flow and their position and conformation continuously followed by spectral imaging during the crossing of barriers, revealing the interplay between carrier structure, micellar stability, and extravasation. Integrating the ability of the micelles to change their fluorescent properties when disassembled, spectral confocal imaging and 3D microfluidic tumor blood vessel-on-a-chip resulted in the establishment of a robust testing platform suitable for real-time imaging and evaluation of supramolecular drug delivery carrier's stability.

Judging Enzyme-Responsive Micelles by Their Covers: Direct Comparison of Dendritic Amphiphiles with Different Hydrophilic Blocks.

Enzymatically degradable polymeric micelles have great potential as drug delivery systems, allowing the selective release of their active cargo at the site of disease. Furthermore, enzymatic degradation of the polymeric nanocarriers facilitates clearance of the delivery system after it has completed its task. While extensive research is dedicated toward the design and study of the enzymatically degradable hydrophobic block, there is limited understanding on how the hydrophilic shell of the micelle can affect the properties of such enzymatically degradable micelles. In this work, we report a systematic head-to-head comparison of well-defined polymeric micelles with different polymeric shells and two types of enzymatically degradable hydrophobic cores. To carry out this direct comparison, we developed a highly modular approach for preparing clickable, spectrally active enzyme-responsive dendrons with adjustable degree of hydrophobicity. The dendrons were linked with three different widely used hydrophilic polymers-poly(ethylene glycol), poly(2-ethyl-2-oxazoline), and poly(acrylic acid) using the CuAAC click reaction. The high modularity and molecular precision of the synthetic methodology enabled us to easily prepare well-defined amphiphiles that differ either in their hydrophilic block composition or in their hydrophobic dendron. The micelles of the different amphiphiles were thoroughly characterized and their sizes, critical micelle concentrations, drug loading, stability, and cell internalization were compared. We found that the micelle diameter was almost solely dependent on the hydrophobicity of the dendritic hydrophobic block, whereas the enzymatic degradation rate was strongly dependent on the composition of both blocks. Drug encapsulation capacity was very sensitive to the type of the hydrophilic block, indicating that, in addition to the hydrophobic core, the micellar shell also has a significant role in drug encapsulation. Incubation of the spectrally active micelles in the presence of cells showed that the hydrophilic shell significantly affects the micellar stability, localization, cell internalization kinetics, and the cargo release mechanism. Overall, the high molecular precision and the ability of these amphiphiles to report their disassembly, even in complex biological media, allowed us to directly compare the different types of micelles, providing striking insights into how the composition of the micelle shells and cores can affect their properties and potential to serve as nanocarriers.

Tuning the Reactivity of Micellar Nanoreactors by Precise Adjustments of the Amphiphile and Substrate Hydrophobicity.

Polymeric assemblies, such as micelles, are gaining increasing attention due to their ability to serve as nanoreactors for the execution of organic reactions in aqueous media. The ability to conduct organic transformations, which have been traditionally limited to organic media, in water is essential for the further development of important fields ranging from green catalysis to bioorthogonal chemistry. Considering the recent progress that has been made to expand the range of organometallic reactions conducted using nanoreactors, we aimed to gain a deeper understanding of the roles of the hydrophobicity of both the core of micellar nanoreactors and the substrates on the reaction rates in water. Toward this goal, we designed a set of five metal-loaded micelles composed of polyethylene glycol-dendron amphiphiles and studied their ability to serve as nanoreactors for a palladium-mediated depropargylation reaction of four substrates with different log P values. Using dendrons as the hydrophobic block, we could precisely tune the lipophilicity of the nanoreactors, which allowed us to reveal linear correlations between the rate constants and the hydrophobicity of the amphiphiles (estimated by the dendron's cLog P). While exponential dependence was obtained for the lipophilicity of the substrates, a similar degree of rate acceleration was observed due to the increase in the hydrophobicity of the amphiphiles regardless of the effect of the substrate's log P. Our results demonstrate that while increasing the hydrophobicity of the substrates may be used to accelerate reaction rates, tuning the hydrophobicity of the micellar nanoreactors can serve as a vital tool for further optimization of the reactivity and selectivity of nanoreactors.

Architectural Change of the Shell-Forming Block from Linear to V-Shaped Accelerates Micellar Disassembly, but Slows the Complete Enzymatic Degradation of the Amphiphiles.

Tuning the enzymatic degradation and disassembly rates of polymeric amphiphiles and their assemblies is crucial for designing enzyme-responsive nanocarriers for controlled drug delivery applications. The common methods to control the enzymatic degradation of amphiphilic polymers are to tune the molecular weights and ratios of the hydrophilic and hydrophobic blocks. In addition to these approaches, the architecture of the hydrophilic block can also serve as a tool to tune enzymatic degradation and disassembly. To gain a deeper understanding of the effect of the molecular architecture of the hydrophilic block, we prepared two types of well-defined PEG-dendron amphiphiles bearing linear or V-shaped PEG chains as the hydrophilic blocks. The high molecular precision of these amphiphiles, which emerges from the utilization of dendrons as the hydrophobic blocks, allowed us to study the self-assembly and enzymatic degradation and disassembly of the two types of amphiphiles with high resolution. Interestingly, the micelles of the V-shaped amphiphiles were significantly smaller and disassembled faster than those of the amphiphiles based on linear PEG. However, the complete enzymatic cleavage of the hydrophobic end groups was significantly slower for the V-shaped amphiphiles. Our results show that the V-shaped architecture can stabilize the unimer state and, hence, plays a double role in the enzymatic degradation and the induced disassembly and how it can be utilized to control the release of encapsulated or bound molecular cargo.

Tuning the molecular weight of polymeric amphiphiles as a tool to access micelles with a wide range of enzymatic degradation rates.

Enyzme-responsive polymeric assemblies hold great potential for biomedical applications due to the over-expression of disease-associated enzymes, which can be utilized to activate such systems only in afflicted tissues. Herein we demonstrate that the overall molecular weight of polymeric amphiphiles, which have the same hydrophilic/hydrophobic ratio, can be tuned to create polymeric micelles with an extreme range of degradation rates. This approach expands the available set of molecular parameters that can be adjusted to tune the degradation rate of polymeric assemblies, paving new possibilities for rational design of polymeric systems with controlled degradation rates.

A microfluidic chip containing multiple 3D nanofibrous scaffolds for culturing human pluripotent stem cells.

In microfluidics-based lab-on-a-chip systems, which are used for investigating the effect of drugs and growth factors on cells, the latter are usually cultured within the device's channels in two-dimensional, and not in their optimal three-dimensional (3D) microenvironment. Herein, we address this shortfall by designing a microfluidic system, comprised of two layers. The upper layer of the system consists of multiple channels generating a gradient of soluble factors. The lower layer is comprised of multiple wells, each deposited with 3D, nanofibrous scaffold. We first used a mathematical model to characterize the fluid flow within the system. We then show that induced pluripotent stem cells can be seeded within the 3D scaffolds and be exposed to a well-mixed gradient of soluble factors. We believe that utilizing such system may enable in the future to identify new differentiation factors, investigate drug toxicity, and eventually allow to perform analyses on patient-specific tissues, in order to fit the appropriate combination and concentration of drugs.

Micellar Stability in Biological Media Dictates Internalization in Living Cells.

The dynamic nature of polymeric assemblies makes their stability in biological media a crucial parameter for their potential use as drug delivery systems in vivo. Therefore, it is essential to study and understand the behavior of self-assembled nanocarriers under conditions that will be encountered in vivo such as extreme dilutions and interactions with blood proteins and cells. Herein, using a combination of fluorescence spectroscopy and microscopy, we studied four amphiphilic PEG-dendron hybrids and their self-assembled micelles in order to determine their structure-stability relations. The high molecular precision of the dendritic block enabled us to systematically tune the hydrophobicity and stability of the assembled micelles. Using micelles that change their fluorescent properties upon disassembly, we observed that serum proteins bind to and interact with the polymeric amphiphiles in both their assembled and monomeric states. These interactions strongly affected the stability and enzymatic degradation of the micelles. Finally, using spectrally resolved confocal imaging, we determined the relations between the stability of the polymeric assemblies in biological media and their cell entry. Our results highlight the important interplay between molecular structure, micellar stability, and cell internalization pathways, pinpointing the high sensitivity of stability-activity relations to minor structural changes and the crucial role that these relations play in designing effective polymeric nanostructures for biomedical applications.

Reversible Dimerization of Polymeric Amphiphiles Acts as a Molecular Switch of Enzymatic Degradability.

Enzyme-responsive polymeric micelles have great potential as drug delivery systems due to the high selectivity and overexpression of disease-associated enzymes, which could be utilized to trigger the release of active drugs only at the target site. We previously demonstrated that enzymatic degradation rates of amphiphilic PEG-dendron hybrids could be precisely tuned by gradually increasing the hydrophobic to hydrophilic ratio. However, with the increase in hydrophobicity, the micelles rapidly became too stable and could not be degraded, as often encountered for many other amphiphilic assemblies. Here we address the challenge to balance between stability and reactivity of enzymatically degradable assemblies by utilizing reversible dimerization of diblock polymeric amphiphiles to yield jemini amphiphiles. This molecular transformation serves as a tool to control the critical micelle concentration of the amphiphiles in order to tune their micellar stability and enzymatic degradability. To demonstrate this approach, we show that simple dimerization of two polymeric amphiphiles through a single reversible disulfide bond significantly increased the stability of their micellar assemblies toward enzymatic degradation, although the hydrophilic to hydrophobic ratio was not changed. Reduction of the disulfide bond led to dedimerization of the polymeric hybrids and allowed their degradation by the activating enzyme. The generality of the approach is demonstrated by designing both esterase- and amidase-responsive micellar systems. This new molecular design can serve as a simple tool to increase the stability of polymeric micelles without impairing their enzymatic degradability.

Modular Synthetic Approach for Adjusting the Disassembly Rates of Enzyme-Responsive Polymeric Micelles.

Self-assembled nanostructures and their stimuli-responsive degradation have been recently explored to meet the increasing need for advanced biocompatible and biodegradable materials for various biomedical applications. Incorporation of enzymes as triggers that can stimulate the degradation and disassembly of polymeric assemblies may be highly advantageous owing to their high selectivity and natural abundance in all living organisms. One of the key factors to consider when designing enzyme-responsive polymers is the ability to fine-tune the sensitivity of the platform toward its target enzyme in order to control the disassembly rate. In this work, a series of enzyme-responsive amphiphilic PEG-dendron hybrids with increasing number of hydrophobic cleavable end-groups was synthesized, characterized, and compared. These hybrids were shown to self-assemble in aqueous media into nanosized polymeric micelles, which could encapsulate small hydrophobic guests in their cores and release them upon enzymatic stimulus. Utilization of dendritic scaffolds as the responsive blocks granted ultimate control over the number of enzymatically cleavable end-groups. Remarkably, as we increased the number of end-groups, the micellar stability increased significantly and the range of enzymatic sensitivity spanned from highly responsive micelles to practically nondegradable ones. The reported results highlight the remarkable role of hydrophobicity in determining the micellar stability toward enzymatic degradation and its great sensitivity to small structural changes of the hydrophobic block, which govern the accessibility of the cleavable hydrophobic groups to the activating enzyme.