Clinical Experience of a Novel Optical Coherence Tomography-Guided Coronary Chronic Total Occlusion Re-Entry Device.

We demonstrated a first-in-human case of successful antegrade dissection and re-entry using an image-guided re-entry catheter that enables real-time high-resolution visualization with graphical augmentation, and precision steering and advancement of a guidewire. The total time from over-the-wire deployment in the proximity of the distal cap to successful re-entry was <20 minutes. (Level of Difficulty: Advanced.).

Time-resolved fluorescence microscopy with phasor analysis for visualizing multicomponent topical drug distribution within human skin.

Understanding a drug candidate's pharmacokinetic (PK) parameters is a challenging but essential aspect of drug development. Investigating the penetration and distribution of a topical drug's active pharmaceutical ingredient (API) allows for evaluating drug delivery and efficacy, which is necessary to ensure drug viability. A topical gel (BPX-05) was recently developed to treat moderate to severe acne vulgaris by directly delivering the combination of the topical antibiotic minocycline and the retinoid tazarotene to the pilosebaceous unit of the dermis. In order to evaluate the uptake of APIs within human facial skin and confirm accurate drug delivery, a selective visualization method to monitor and quantify local drug distributions within the skin was developed. This approach uses fluorescence lifetime imaging microscopy (FLIM) paired with a multicomponent phasor analysis algorithm to visualize drug localization. As minocycline and tazarotene have distinct fluorescence lifetimes from the lifetime of the skin's autofluorescence, these two APIs are viable targets for distinct visualization via FLIM. Here, we demonstrate that the analysis of the resulting FLIM output can be used to determine local distributions of minocycline and tazarotene within the skin. This approach is generalizable and can be applied to many multicomponent fluorescence lifetime imaging targets that require cellular resolution and molecular specificity.

Imaging and quantifying drug delivery in skin - Part 2: Fluorescence andvibrational spectroscopic imaging methods.

Understanding the delivery and diffusion of topically-applied drugs on human skin is of paramount importance in both pharmaceutical and cosmetics research. This information is critical in early stages of drug development and allows the identification of the most promising ingredients delivered at optimal concentrations to their target skin compartments. Different skin imaging methods, invasive and non-invasive, are available to characterize and quantify the spatiotemporal distribution of a drug within ex vivo and in vivo human skin. The first part of this review detailed invasive imaging methods (autoradiography, MALDI and SIMS). This second part reviews non-invasive imaging methods that can be applied in vivo: i) fluorescence (conventional, confocal, and multiphoton) and second harmonic generation microscopies and ii) vibrational spectroscopic imaging methods (infrared, confocal Raman, and coherent Raman scattering microscopies). Finally, a flow chart for the selection of imaging methods is presented to guide human skin ex vivo and in vivo drug delivery studies.

Visualizing topical drug uptake with conventional fluorescence microscopy and deep learning.

Mapping the uptake of topical drugs and quantifying dermal pharmacokinetics (PK) presents numerous challenges. Though high resolution and high precision methods such as mass spectrometry offer the means to quantify drug concentration in tissue, these tools are complex and often expensive, limiting their use in routine experiments. For the many topical drugs that are naturally fluorescent, tracking fluorescence emission can be a means to gather critical PK parameters. However, skin autofluorescence can often overwhelm drug fluorescence signatures. Here we demonstrate the combination of standard epi-fluorescence imaging with deep learning for the visualization and quantification of fluorescent drugs in human skin. By training a U-Net convolutional neural network on a dataset of annotated images, drug uptake from both high "infinite" dose and daily clinical dose regimens can be measured and quantified. This approach has the potential to simplify routine topical product development in the laboratory.

First Use of Optical Coherence Tomography on In Vivo Inflammatory Acne-Like Lesions: A Murine Model.

BACKGROUND AND OBJECTIVES: Successful outcomes of clinical studies for acne vulgaris depend greatly on achieving statistically significant reduction in acne lesion count and improvement in Investigator's Global Assessment score of the investigational drug product against its vehicle control. To date, there has not been a validated preclinical acne model to evaluate investigational drug products in order to improve the probability of clinical success. An inflammatory acne-like lesion mouse model developed in-house has previously been used for clinical guidance in our drug development program. In this study, we aim to implement and assess the adequacy of swept-source optical coherence tomography (SS-OCT) in quantifying the dynamic changes in inflammatory acne-like lesions. STUDY DESIGN/MATERIALS AND METHODS: Live Propionibacterium acnes bacteria were injected intradermally resulting in inflammatory acne-like lesions. Topical 1% and 2% minocycline gels were applied to the lesions in separate groups once daily for 2 weeks and compared with vehicle and untreated control groups. The growth of these lesions was monitored and measured with a ruler (height)/microcaliper (width)-an approach previously developed, and with SS-OCT. The reliability of the two methods were assessed. Acquired OCT images across the apex of these inflammatory lesions were statistically analyzed for lesion volume reduction from baseline as well as between the treatment groups and the control groups. RESULTS: The OCT technique allowed for reliable lesion volume analysis with varying conic profiles. After 14 days of topical minocycline treatments (1%, 2% minocycline), statistically significant reduction in lesion volume (P ≤ 0.05) based on OCT image analysis was observed compared with untreated and vehicle control groups as well as compared with baseline measurements. Under the right conditions, some morphological aspects of the P. acnes injection site were discernible within the skin in images captured with OCT. CONCLUSIONS: We demonstrated the first use of SS-OCT in evaluating in vivo inflammatory acne-like lesions in a murine model. Our findings support the use of OCT in assessing lesion size and evolution of P. acnes injection sites non-invasively in preclinical in vivo studies, which could potentially lead to more consistent and predictable outcomes in clinical development. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc.

Imaging and quantifying drug delivery in skin - Part 1: Autoradiography and mass spectrometry imaging.

In this two-part review we present an up-to-date description of different imaging methods available to map the localization of drugs on skin as a complement of established ex-vivo absorption studies. This first part deals with invasive methods which are grouped in two classes according to their underlying principles: i) methods using radioactivity such as autoradiography and ii) mass spectrometry methods such as MALDI and SIMS. For each method, a description of the principle is given along with example applications of imaging and quantifying drug delivery in human skin. Thanks to these techniques a better assessment of the fate of drugs is obtained: its localization on a particular skin structure, its potential accumulation, etc. A critical comparison in terms of capabilities, sensitivity and practical applicability is included that will help the reader to select the most appropriate technique depending on the particular problem to be solved.

Topical minocycline formulations: Evaluation and comparison of dermal uptake efficacy.

Acne vulgaris is a clinically distinct skin condition with evidence suggesting that inflammation plays a critical role in the pathogenesis of this disorder. Treatment of severe inflammatory acne often involves the use of oral antibiotics, sometimes in combination with topical products. Oral antibiotics often result in systemic side effects and the risks of antibiotic resistance, but no commercial topical minocycline is currently available. We have developed a unique, stable, hydrophilic topical gel formulation with fully solubilized minocycline (MNC-H). Minocycline delivered in our hydrophilic gel remained more stable in situ, resulting in less degradation product (4-epiminocycline) than a lipophilic formulation (MNC-L). The hydrophilic nature of our formulation enabled 2-3 fold increase in delivery into the skin ex vivo compared to a lipophilic counterpart, mostly seen in the epidermis and pilosebaceous units. The lipophilic formulation also appeared to be more occlusive, resulting in higher sebum production in minipigs, which may exacerbate acne vulgaris. As our results indicate, a 1, 2% minocycline hydrophilic gel may deliver sufficient drug (>15 µg/g) to potentially demonstrate clinical efficacy. These findings suggest that topical hydrophilic minocycline gel may provide a novel tool for topical acne therapy.

Characterization of human cutaneous tissue autofluorescence: implications in topical drug delivery studies with fluorescence microscopy.

In pharmacokinetic studies of topical drugs, fluorescence microscopy methods can enable the direct visualization and quantification of fluorescent drugs within the skin. One potential limitation of this approach, however, is the strong endogenous fluorescence of skin tissues that makes straightforward identification of specific drug molecules challenging. To study this effect and quantify endogenous skin fluorescence in the context of topical pharmacokinetics, an integrating sphere-based screening tool was designed to collect fluorescence yield data from human skin specimens. Such information could be utilized to select specific donors in the investigation of drug uptake and distribution. Results indicated human facial skin specimens from a group of more than 35 individuals exhibited an at least 6-fold difference in endogenous fluorescence. In visualizing drug distributions, the negative impact of autofluorescence could be exacerbated in cases where there are overlapping spatial distributions or spectral emission profiles between endogenous fluorophores and the exogenous fluorophore of interest. We demonstrated the feasibility of this approach in measuring the range of tissue endogenous fluorescence and selecting specimens for the study of drug pharmacokinetics with fluorescence microscopy.

Visualization of drug distribution of a topical minocycline gel in human facial skin.

Acne vulgaris is a common chronic skin disease in young adults caused by infection of the pilosebaceous unit, resulting in pimples and possibly permanent scarring on the skin. Minocycline, a common antibiotic, has been widely utilized as a systemic antimicrobial treatment for acne via oral administration. Recently, a topical minocycline gel (BPX-01) was developed to directly deliver minocycline through the epidermis and into the pilosebaceous unit to achieve localized treatment with lower doses of drug. As the effectiveness of the drug is directly related to its successful delivery, there is a need to evaluate the pharmacokinetics at the cellular level within tissue. Advantageously, minocycline is naturally fluorescent and can be directly visualized using microscopy-based approaches. Due to high endogenous autofluorescence, however, imaging of weakly emitting fluorescent molecules such as minocycline in skin tissue can be challenging. Here, we demonstrate a method for the selective visualization of minocycline within human skin tissue by utilizing two-photon excitation fluorescence (TPEF) microscopy and fluorescence lifetime imaging microscopy (FLIM). To demonstrate the feasibility of this approach, ex vivo human facial skin samples treated with various concentrations of BPX-01 were investigated. From the TPEF analysis, we were able to visualize relatively high levels of drug uptake within facial skin. However, minocycline fluorescence could be overwhelmed by endogenous fluorescence that complicates TPEF quantitative analysis, making FLIM more advantageous for visualizing drug uptake. Importantly, we found a unique signature of minocycline uptake via FLIM analysis that enabled the successful differentiation of the drug and enabled the extraction of drug local distribution from the endogenous fluorescence using a non-Euclidean phasor analysis method. Based on these results, we believe that the drug local distribution visualization method using TPEF and FLIM with phasor analysis can play an important role in studying the pharmacokinetics and pharmacodynamics of a topically applicable drug.

Immunohistochemical evaluation of the heat shock response to nonablative fractional resurfacing.

Despite the emergence of nonablative fractional resurfacing (NFR) as a new therapeutic modality for skin photoaging, little is known about the molecular events that underlie the heat shock response to different treatment parameters. Human subjects are treated with a scanned 1550-nm fractional laser at pulse energies spanning 6 to 40 mJ and a 140-µm spot size. The heat shock response is assessed immunohistochemically immediately through 7 days posttreatment. At the immediately posttreatment time point, we observe subepidermal clefting in most sections. The basal epidermis and dermal zones of sparing are both found to express HSP47, but not HSP72. By day 1, expression of HSP72 is detected throughout the epidermis, while that of HSP47 remains restricted to the basal layer. Both proteins are detected surrounding the dermal portion of the microscopic treatment zone (MTZ). This pattern of expression persists through day 7 post-NFR, although neither protein is found within the MTZ. Immediately posttreatment, the mean collagen denaturation zone width is 50 µm at 6 mJ, increasing to 202 µm at 40 mJ. The zone of cell death exceeds the denaturation zone by 19 to 55% over this pulse energy range. The two zones converge by day 7 posttreatment.

In vivo confocal imaging of epidermal cell migration and dermal changes post nonablative fractional resurfacing: study of the wound healing process with corroborated histopathologic evidence.

In vivo wound healing response post nonablative fractional laser treatment is evaluated. Seven healthy subjects receive treatments with a Fraxel re:store laser system on the forearm with pulse energies ranging from 10 to 70 mJ. The treatment sites are imaged at 1-h increments up to 40 h using confocal microscope z-stacks using 10-mum-depth spacing. At least five individual microscopic treatment zones are imaged per subject, time point, and treatment energy. Images are analyzed for tissue structure and morphology to classify each lesion as healed or not healed, depending on epidermal re-epithelialization at each time point and treatment energy. Probit analysis is used to statistically determine the ED(50) and ED(84) probabilities for a positive dose response (healed lesion) as a function of treatment energy. Confocal observations reveal epidermal keratinocyte migration patterns confirmed with histological analysis using hematoxylin and eosin (HE) and lactate dehydrogenase (LDH) staining at 10 mJ at 0, 7, 16, and 24-h post-treatment. Results indicate that more time is required to conclude re-epithelialization with larger lesion sizes (all less than 500 mum) corresponding to higher treatment energies. For the entire pulse energy range tested, epidermal re-epithelialization concludes between 10 to 22-h post-treatment for ED(50) and 13 to 28 h for ED(84).

Fractional deep dermal ablation induces tissue tightening.

BACKGROUND AND OBJECTIVE: Due to the significant risk profile associated with traditional ablative resurfacing, a safer and less invasive treatment approach known as fractional deep dermal ablation (FDDA) was recently developed. We report the results of the first clinical investigation of this modality for treatment of photodamaged skin. STUDY DESIGN/MATERIALS AND METHODS: Twenty-four subjects received treatments on the inner forearm with a prototype fractional CO(2) laser device (Reliant Technologies Inc., Mountain View, CA) at settings of 5-40 mJ/MTZ and 400 MTZ/cm(2). Clinical and histological effects were assessed by study investigators 1 week, 1 month, and 3 months following treatment. Thirty subjects were then enrolled in a multi-center study for treatment of photodamage using the same device. Subjects received 1-2 treatments on the face and neck, with energies ranging from 10 to 40 mJ/MTZ and densities ranging from 400 to 1,200 MTZ/cm(2). Study investigators assessed severity of post-treatment responses during follow-up visits 48 hours, 1 week, 1 month, and 3 months following treatment. Using a standard quartile improvement scale (0-4), subjects and investigators assessed improvement in rhytides, pigmentation, texture, laxity and overall appearance 1 and 3 months post-treatment. RESULTS: Clinical and histologic results demonstrated that fractional delivery of a 10,600 nm CO(2) laser source offers an improved safety profile with respect to traditional ablative resurfacing, while still effectively resurfacing epidermal and dermal tissue. Forearm and facial treatments were well-tolerated with no serious adverse events observed. Eighty-three percent of subjects exhibited moderate or better overall improvement (50-100%), according to study investigator quartile scoring. CONCLUSIONS: FDDA treatment is a safe and promising new approach for resurfacing of epidermal and deep dermal tissue targets.