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In oncology, technologies for clinical molecular imaging are used to diagnose patients, establish the efficacy of treatments and monitor the recurrence of disease. Multiplexed methods increase the number of disease-specific biomarkers that can be detected simultaneously, such as the overexpression of oncogenic proteins, aberrant metabolite uptake and anomalous blood perfusion. The quantitative localization of each biomarker could considerably increase the specificity and the accuracy of technologies for clinical molecular imaging to facilitate granular diagnoses, patient stratification and earlier assessments of the responses to administered therapeutics. In this Review, we discuss established techniques for multiplexed imaging and the most promising emerging multiplexing technologies applied to the imaging of isolated tissues and cells and to non-invasive whole-body imaging. We also highlight advances in radiology that have been made possible by multiplexed imaging.

Cell surface marker expression in tumors dictates the selection of therapeutics, therapy response, and survival. However, biopsies are invasive, sample only a small area of the tumor landscape and may miss significant areas of heterogeneous expression. Here, we investigated the potential of antibody-conjugated surface-enhanced resonance Raman scattering nanoparticles (SERRS-NPs) to depict and quantify high and low tumoral surface marker expression, focusing on the surface markers epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) in an intracerebral and peripheral setting with an inter- and intratumoral comparison of Raman signal intensities.

Surface-enhanced Raman spectroscopy (SERS) nanotags hold a unique place among bioimaging contrast agents due to their fingerprint-like spectra, which provide one of the highest degrees of detection specificity. However, in order to achieve a sufficiently high signal intensity, targeting capabilities, and biocompatibility, all components of nanotags must be rationally designed and tailored to a specific application. Design parameters include fine-tuning the properties of the plasmonic core as well as optimizing the choice of Raman reporter molecule, surface coating, and targeting moieties for the intended application. This review introduces readers to the principles of SERS nanotag design and discusses both established and emerging protocols of their synthesis, with a specific focus on the construction of SERS nanotags in the context of bioimaging and theranostics.

Multiplexed imaging of 8 molecular targets with SERS nanoprobes in mice reveals spatially heterogeneous tumor responses to immunotherapy. Multichannel image segmentation enables classification of the tumor regions into responders and naïve.

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Purpose: Integrin αvβ3 plays an important role in tumor angiogenesis, growth, and metastasis. We have tested a targeted probe to visualize integrin receptor expression in glioblastomas using near-infrared fluorescent (NIRF) imaging.Experimental design: A transgenic glioblastoma mouse model (RCAS-PDGF-driven/tv-a glioblastoma, which mimics the infiltrative growth pattern of human glioblastomas) and two human orthotopic glioblastoma models (U-87 MG with high integrin β3 expression and TS543 with low integrin β3 expression) were studied. An integrin-targeting NIRF probe, IRDye 800CW-cyclic-RGD peptide (IRDye 800CW-RGD), was tested by in vivo and ex vivo NIRF imaging.Results: We show that the IRDye 800CW-RGD peptide: (i) specifically binds to integrin receptors; (ii) is selectively localized to glioblastoma tissue with overexpressed integrin receptors and is retained over prolonged periods of time; (iii) is associated with minimal autofluorescence and photobleaching because of imaging at 800 nm; (iv) provides delineation of tumor tissue with high precision because of a high tumor-to-normal brain fluorescence ratio (79.7 ± 6.9, 31.2 ± 2.8, and 16.3 ± 1.3) in the U-87 MG, RCAS-PDGF, and TS543 models, respectively; P < 0.01); and (v) enables fluorescence-guided glioblastoma resection. Importantly, small foci of residual fluorescence were observed after resection was completed using white light imaging alone, and these fluorescent foci were shown to represent residual tumor tissue by histology.Conclusions: NIRF imaging with the IRDye 800CW-RGD probe provides a simple, rapid, low-cost, nonradioactive, and highly translatable approach for improved intraoperative glioblastoma visualization and resection. It also has the potential to serve as an imaging platform for noninvasive cancer detection and drug efficacy evaluation studies. Clin Cancer Res; 18(20); 5731–40. ©2012 AACR.

Visualizing the presence and distribution of multiple specific molecular markers within a tumor can reveal the composition of its microenvironment, inform diagnosis, stratify patients, and guide treatment. Raman imaging with multiple molecularly-targeted surface enhanced Raman scattering (SERS) nanoprobes could help investigate emerging cancer treatments preclinically or enable personalized treatment assessment. Here, we report a comprehensive strategy for multiplexed imaging using SERS nanoprobes and machine learning (ML) to monitor the early effects of immune checkpoint blockade (ICB) in tumor-bearing mice. We used antibody-functionalized SERS nanoprobes to visualize 7 + 1 immunotherapy-related targets simultaneously. The multiplexed images were spectrally resolved and then spatially segmented into superpixels based on the unmixed signals. The superpixels were used to train ML models, leading to the successful classification of mice into treated and untreated groups, and identifying tumor regions with variable responses to treatment. This method may help predict treatment efficacy in tumors and identify areas of tumor variability and therapy resistance.

Rationale: Most contemporary cancer therapeutic paradigms involve initial imaging as a treatment roadmap, followed by the active engagement of surgical operations. Current approved intraoperative contrast agents exemplified by indocyanine green (ICG) have a few drawbacks including the inability of pre-surgical localization. Alternative near-infrared (NIR) dyes including IRDye800cw are being explored in advanced clinical trials but often encounter low chemical yields and complex purifications owing to the asymmetric synthesis. A single contrast agent with ease of synthesis that works in multiple cancer types and simultaneously allows presurgical imaging, intraoperative deep-tissue three-dimensional visualization, and high-speed microscopic visualization of tumor margins via spatiotemporally complementary modalities would be beneficial. Methods: Due to the lack of commercial availability and the absence of detailed synthesis and characterization, we proposed a facile and scalable synthesis pathway for the symmetric NIR water-soluble heptamethine sulfoindocyanine IRDye78. The synthesis can be accomplished in four steps from commercially-available building blocks. Its symmetric resonant structure avoided asymmetric synthesis problems while still preserving the benefits of analogous IRDye800cw with commensurable optical properties. Next, we introduced a low-molecular-weight protein alpha-lactalbumin (α-LA) as the carrier that effectively modulates the hepatic clearance of IRDye78 into the preferred renal excretion pathway. We further implemented 89Zr radiolabeling onto the protein scaffold for positron emission tomography (PET). The multimodal imaging capability of the fluorophore-protein complex was validated in breast cancer and glioblastoma. Results: The scalable synthesis resulted in high chemical yields, typically 95% yield in the final step of the chloro dye. Chemical structures of intermediates and the final fluorophore were confirmed. Asymmetric IRDye78 exhibited comparable optical features as symmetric IRDye800cw. Its well-balanced quantum yield affords concurrent dual fluorescence and optoacoustic contrast without self-quenching nor concentration-dependent absorption. The NHS ester functionality modulates efficient covalent coupling to reactive side-chain amines to the protein carrier, along with desferrioxamine (DFO) for stable radiolabeling of 89Zr. The fluorophore-protein complex advantageously shifted the biodistribution and can be effectively cleared through the urinary pathway. The agent accumulates in tumors and enables triple-modal visualization in mouse xenograft models of both breast and brain cancers. Conclusion: This study described in detail a generalized strategic modulation of clearance routes towards the favorable renal clearance, via the introduction of α-LA. IRDye78 as a feasible alternative of IRDye800cw currently in clinical phases was proposed with a facile synthesis and fully characterized for the first time. This fluorophore-protein complex with stable radiolabeling should have great potential for clinical translation where it could enable an elegant workflow from preoperative planning to intraoperative deep tissue and high-resolution image-guided resection.

Immunotherapy has revolutionized the treatment of several malignancies. Notwithstanding the encouraging results, many patients do not respond to treatments. Evaluation of the efficacy of treatments is challenging and robust methods to predict the response to treatment are not yet available. The outcome of immunotherapy results from changes that treatment evokes in the tumor immune landscape. Therefore, a better understanding of the dynamics of immune cells that infiltrate into the tumor microenvironment may fundamentally help in addressing this challenge and provide tools to assess or even predict the response. Noninvasive imaging approaches, such as PET and SPECT that provide whole-body images are currently seen as the most promising tools that can shed light on the events happening in tumors in response to treatment. Such tools can provide critical information that can be used to make informed clinical decisions. Here, we review recent developments in the field of noninvasive cancer imaging with a focus on immunotherapeutics and nuclear imaging technologies and will discuss how the field can move forward to address the challenges that remain unresolved.

In recent years, Raman spectroscopy has undergone major advancements in its ability to probe deeply through turbid media such as biological tissues. This progress has been facilitated by the advent of a range of specialist techniques based around spatially offset Raman spectroscopy (SORS) to enable non-invasive probing of living tissue through depths of up to 5 cm. This represents an improvement in depth penetration of up to two orders of magnitude compared to what can be achieved with conventional Raman methods. In combination with the inherently high molecular specificity of Raman spectroscopy, this has therefore opened up entirely new prospects for a range of new analytical applications across multiple fields including medical diagnosis and disease monitoring. This article discusses SORS and related variants of deep Raman spectroscopy such as transmission Raman spectroscopy (TRS), micro-SORS and surface enhanced spatially offset Raman spectroscopy (SESORS), and reviews the progress made in this field during the past 5 years including advances in non-invasive cancer diagnosis, monitoring of neurotransmitters, and assessment of bone disease., Recent advances in non-invasive biomedical analysis using SORS are discussed.

Theranostic agents should ideally be renally cleared and biodegradable. Here, we report the synthesis, characterization and theranostic applications of fluorescent ultrasmall gold quantum clusters that are stabilized by the milk metalloprotein alpha-lactalbumin. We synthesized three types of these nanoprobes that together display fluorescence across the visible and near-infrared spectra when excited at a single wavelength through optical colour coding. In live tumour-bearing mice, the near-infrared nanoprobe generates contrast for fluorescence, X-ray computed tomography and magnetic resonance imaging, and exhibits long circulation times, low accumulation in the reticuloendothelial system, sustained tumour retention, insignificant toxicity and renal clearance. An intravenously administrated near-infrared nanoprobe with a large Stokes shift facilitated the detection and image-guided resection of breast tumours in vivo using a smartphone with modified optics. Moreover, the partially unfolded structure of alpha-lactalbumin in the nanoprobe helps with the formation of an anti-cancer lipoprotein complex with oleic acid that triggers the inhibition of the MAPK and PI3K–AKT pathways, immunogenic cell death and the recruitment of infiltrating macrophages. The biodegradability and safety profile of the nanoprobes make them suitable for the systemic detection and localized treatment of cancer. Renally clearable gold quantum clusters that are stabilized by the milk metalloprotein alpha-lactalbumin and display multicolour fluorescence aid the detection, resection and treatment of breast cancer in mice.

Free functioning muscle transfer is a reconstructive option to restore elbow flexion in brachial plexus injuries. The authors determined the impact of body mass index, age, and location of distal tendon attachment on elbow flexion strength after free functioning muscle transfer in traumatic brachial plexus injury patients.A retrospective review of patients who underwent free functioning muscle transfer for elbow flexion as part of their brachial plexus injury reconstruction with a minimum 2-year follow-up were evaluated. Outcomes assessed included elbow flexion strength (British Medical Research Council grade) and change in Disabilities of the Arm, Shoulder and Hand questionnaire and visual analogue scale pain scores.One hundred six patients met inclusion criteria. The average age was 32 years, and the average body mass index was 27.1 kg/m2; 56.5 percent of patients achieved M3 or greater muscle grade using the authors' strict modification of the British Medical Research Council scale. Disabilities of the Arm, Shoulder and Hand questionnaire scores improved from 45.7 to 38.8 (p0.05). Visual analogue scale pain scores decreased, but this trend did not obtain significance. Age and body mass index both had a significant negative impact on final free functioning muscle transfer grade (p0.05). Use of a distal tendon insertion led to improved muscle grade outcomes, with targeting of wrist extension being superior to finger flexion (p0.05). Simultaneous musculocutaneous nerve grafting did not significantly alter final elbow flexion strength.Increasing age and body mass index both imparted a deleterious effect on free functioning muscle transfer muscle grade. Distal muscle targets had better strength outcomes than when the biceps tendon was used.Risk, III.

Targeted imaging and therapy for triple-negative breast cancer (TNBC) in the perioperative period are believed to be imperative for better disease management and improved life expectancy. Still, they are not yet available in clinical settings, and only a few nanoparticle-based theranostic agents potentially offer these capabilities. Herein, we develop an innovative class of biocompatible triple-modality nanoprobes (TMNPs) that offer optical imaging using optoacoustic, fluorescence, and surface-enhanced Raman scattering (SERS), as well as photothermal therapy (PTT) with near-infrared (NIR) light. The TMNPs are fabricated by immobilizing positively charged NIR fluorophores on negatively charged DNA-coated gold nanorods (AuNR), then silica encapsulation. The DNA-based design allows the screening of commercially available positively charged NIR fluorophores for the optimum fluorescence and SERS signals. After the design optimization, we functionalize TMNPs with folate groups to target folate receptor1 (FOLR1)-overexpressing TNBC in vitro and in vivo. Our results reveal that TMNPs preferentially accumulate in the FOLR1 positive tumors in TNBC patient-derived xenograft mouse models and show excellent imaging capabilities with all three imaging modalities. Selective exposure of the tumor with NIR laser further shows efficient thermal tissue ablation without causing systemic toxicity. Collectively, TMNP holds great promise for real-time multiplexed imaging of cancer biomarkers and therapeutic capability.

We sought to identify predictors of failed ulnar nerve fascicle (to flexor carpi ulnaris) to biceps motor branch transfer. A retrospective review of adult brachial plexus patients treated with flexor carpi ulnaris to biceps transfer with a minimum 1-year follow-up was performed. Failure, defined as modified British Medical Research Council grade M3 flexion met criteria. Eighteen failures and 18 controls, with similar follow-up (20 vs 23 months) were evaluated. Preoperative flexor carpi ulnaris weakness (M Level of evidence: III

In the last two decades, DNA has attracted significant attention toward the development of materials at the nanoscale for emerging applications due to the unparalleled versatility and programmability of DNA building blocks. DNA‐based artificial nanomaterials can be broadly classified into two categories: DNA nanostructures (DNA‐NSs) and DNA‐functionalized nanoparticles (DNA‐NPs). More importantly, their use in nanotheranostics, a field that combines diagnostics with therapy via drug or gene delivery in an all‐in‐one platform, has been applied extensively in recent years to provide personalized cancer treatments. Conveniently, the ease of attachment of both imaging and therapeutic moieties to DNA‐NSs or DNA‐NPs enables high biostability, biocompatibility, and drug loading capabilities, and as a consequence, has markedly catalyzed the rapid growth of this field. This review aims to provide an overview of the recent progress of DNA‐NSs and DNA‐NPs as theranostic agents, the use of DNA‐NSs and DNA‐NPs as gene and drug delivery platforms, and a perspective on their clinical translation in the realm of oncology., DNA‐based artificial nanomaterials, namely, DNA nanostructures (DNA‐NSs) and DNA‐functionalized nanoparticles (DNA‐NPs), are emerging as an all‐in‐one platform for imaging and therapy of different cancers. The ease of attachment of both imaging and therapeutic molecules, biostability, biocompatibility has augmented the growth of the field. This review aims at providing an overview of the recent progress in this rapidly expanding field.

There is controversy regarding the effectiveness of brachial plexus reconstruction in older patients, as outcomes are thought to be poor. The aim of this study is to determine the outcomes of shoulder abduction obtained after nerve reconstruction in patients over the age of 50 years and factors related to success.Forty patients over the age of 50 years underwent nerve surgery to improve shoulder function after a traumatic brachial plexus injury. Patients were evaluated pre- and postoperatively for shoulder abduction strength and range of motion (ROM); Disability of the Arm, Shoulder and Hand (DASH) scores; pain; age bracket; gender; body mass index (BMI); delay from injury to operation; concomitant trauma; severity of trauma; and type of reconstruction.The average age was 58.2 years (range 50-77 years) with an average follow-up of 18.8 months. The average modified British Medical Research Council (BMRC) shoulder abduction grade improved significantly from 0.23 to 2.03 (p 0.005). Fourteen patients achieved functional shoulder abduction of ≥ M3 postoperatively. There was no correlation between age or age range stratification and BMRC grade or those obtaining useful shoulder abduction ≥ M3. Active shoulder abduction improved significantly from 18.25° to 40.64°, with no difference on the basis of age or age stratification. There were improved modified BMRC grades with nerve transfers versus nerve grafts. Less patients achieved ≥ M3 function if surgery was delayed 6 months. The mean DASH score decreased from 45.3 to 40.7 postoperatively, and the average pain score decreased from 3.7 to 3.0. Patients with a higher postoperative BMRC grade for shoulder abduction had improved postoperative DASH scores and VAS for pain (p = 0.011 and 0.005, respectively).Brachial plexus nerve reconstruction for shoulder abduction in patients over the age of 50 years can yield useful BMRC scores and ROM, and age should not be used to exclude nerve reconstruction in these patients.

Objective: Monitoring emerging vascular-targeted photodynamic therapy (VTP) and understanding the time-dynamics of treatment effects remains challenging. We interrogated whether handheld multispectral optoacoustic tomography (MSOT) could noninvasively monitor the effect of VTP using WST11, a vascular-acting photosensitizer, on tumor tissues over time using a renal cell cancer mouse model. We also investigated whether MSOT illumination can induce VTP, to implement a single-modality theranostic approach. Materials and Methods: Eight BalB/c mice were subcutaneously implanted with murine renal adenocarcinoma cells (RENCA) on the flank. Three weeks later VTP was performed (10 min continuous illumination at 753 nm following intravenous infusion) using WST11 or saline as control. Handheld MSOT images were collected prior to VTP administration and subsequently thereafter over the course of the first hour, at 24 and 48 h. Data collected were unmixed for blood oxygen saturation in tissue (SO 2 ) based on the spectral signatures of deoxy- and oxygenated hemoglobin. Changes in oxygen saturation over time, relative to baseline, were examined by paired t-test for statistical significance (p < 0.05). In-vivo findings were corroborated by histological analyses of the tumor tissue. Results: MSOT is shown to prominently resolve changes in oxygen saturation in tumors within the first 20 min post WST11-VTP treatment. Within the first hour post-treatment, SO 2 decreased by more than 60% over baseline (p < 0.05), whereas it remained unchanged (p > 0.1) in the sham-treated group. Moreover, unlike in the control group, SO 2 in treated tumors further decreased over the course of 24 to 48 h post-treatment, concomitant with the propagation of profound central tumor necrosis present in histological analysis. We further show that pulsed MSOT illumination can activate WST11 as efficiently as the continuous wave irradiation employed for treatment. Conclusion: Handheld MSOT non-invasively monitored WST11-VTP effects based on the SO 2 signal and detected blood saturation changes within the first 20 min post-treatment. MSOT may potentially serve as a means for both VTP induction and real-time VTP monitoring in a theranostic approach.

The unique spectral signatures and biologically inert compositions of surface-enhanced resonance Raman scattering (SERRS) nanoparticles make them promising contrast agents for in vivo cancer imaging. Our SERRS nanoparticles consist of a 60-nm gold nanoparticle core that is encapsulated in a 15-nm-thick silica shell wherein the resonant Raman reporter is embedded. Subtle aspects of their preparation can shift their limit of detection by orders of magnitude. In this protocol, we present the optimized, step-by-step procedure for generating reproducible SERRS nanoparticles with femtomolar (10-15 M) limits of detection. We provide ways of characterizing the optical properties of SERRS nanoparticles using UV/VIS and Raman spectroscopy, and their physicochemical properties using transmission electron microscopy and nanoparticle tracking analysis. We introduce several applications of these nanoprobes for biomedical research, with a focus on intraoperative cancer imaging via Raman imaging. A detailed account is provided for successful i.v. administration of SERRS nanoparticles such that delineation of cancerous lesions can be achieved in vivo and ex vivo on resected tissues without the need for specific biomarker targeting. This straightforward, yet comprehensive, protocol-from initial de novo gold nanoparticle synthesis to SERRS nanoparticle contrast-enhanced preclinical Raman imaging in animal models-takes ∼96 h.

Research activity in medical and cancer nanotechnology has grown dramatically over the past 15 years. The field has become a cradle of multidisciplinary investigations bringing together physicists, chemists, and engineers working with clinicians and biologists to address paramount problems in cancer care and treatment. Some have argued that the explosion in the number of research papers has not been followed by sufficient clinical activity in nanomedicine. However, three new nanodrugs have now been approved by the U.S. Food and Drug Administration (FDA) in the past three years, confirming the validity of nanotechnology approaches in cancer. Excitingly, translational pipelines contain several additional intriguing candidates. In this Nano Focus article, we discuss potential barriers inhibiting further incorporation of nanomedicines into patient care, possible strategies to overcome these barriers, and promising new directions in cancer interventions based on nanotechnology. Insights presented herein are outcomes of discussions held at a recent strategic workshop hosted by the National Cancer Institute (NCI), which brought together research, clinical, and commercial leaders of the nanomedicine field.

Rationale: The goal of imaging tumors at depth with high sensitivity and specificity represents a significant challenge in the field of biomedical optical imaging. 'Surface enhanced Raman scattering' (SERS) nanoparticles (NPs) have been employed as image contrast agents and can be used to specifically target cells in vivo. By tracking their unique “fingerprint” spectra, it becomes possible to determine their precise location. However, while the detection of SERS NPs is very sensitive and specific, conventional Raman spectroscopy imaging devices are limited in their inability to probe through tissue depths of more than a few millimetres, due to scattering and absorption of photons by biological tissues. Here, we combine the use of "Spatially Offset Raman spectroscopy" (SORS) with that of "surface-enhanced resonance Raman spectroscopy" (SERRS) in a technique known as "surface enhanced spatially offset resonance Raman spectroscopy" (SESO(R)RS) to image deep-seated glioblastoma multiforme (GBM) tumors in vivo in mice through the intact skull. Methods: A SORS imaging system was built in-house. Proof of concept SORS imaging was achieved using a PTFE-skull-tissue phantom. Imaging of GBMs in the RCAS-PDGF/N-tva transgenic mouse model was achieved through the use of gold nanostars functionalized with a resonant Raman reporter to create SERRS nanostars. These were then encapsulated in a thin silica shell and functionalized with a cyclic-RGDyK peptide to yield integrin-targeting SERRS nanostars. Non-invasive in vivo SORS image acquisition of the integrin-targeted nanostars was then performed in living mice under general anesthesia. Conventional non-SORS imaging was used as a direct comparison. Results: Using a low power density laser, GBMs were imaged via SESORRS in mice (n = 5) and confirmed using MRI and histopathology. The results demonstrate that via utilization of the SORS approach, it is possible to acquire clear and distinct Raman spectra from deep-seated GBMs in mice in vivo through the skull. SESORRS images generated using classical least squares outlined the tumors with high precision as confirmed via MRI and histology. Unlike SESORRS, conventional Raman imaging of the same areas did not provide a clear delineation of the tumor. Conclusion: To the best of our knowledge this is the first report of in vivo SESO(R)RS imaging. In a relevant brain tumor mouse model we demonstrate that this technique can overcome the limitations of conventional Raman imaging with regards to penetration depth. This work therefore represents a significant step forward in the potential clinical translation of SERRS nanoparticles for high precision cancer imaging.

Ovarian cancer has a unique pattern of metastatic spread, in that it initially spreads locally within the peritoneal cavity. This is in contrast to most other cancer types, which metastasize early on via the blood stream to distant sites. This unique behavior opens up an opportunity for local application of both therapeutic and imaging agents. Upon initial diagnosis, 75% of patients already present with diffuse peritoneal spread involving abdominal organs. Complete resection of all tumor implants has been shown to be a major factor for improved survival. Unfortunately, it is currently not possible for surgeons to visualize microscopic implants, impeding their removal and leading to tumor recurrences and poor outcomes in most patients. Thus, there is a great need for new intraoperative imaging techniques that can overcome this hurdle. We devised a method that employs folate receptor (FR)-targeted surface-enhanced resonance Raman scattering (SERRS) nanoparticles (NP), as folate receptors are typically overexpressed in ovarian cancer. We report a robust ratiometric imaging approach using anti-FR-SERRS-NPs (αFR-NPs) and non-targeted SERRS-NPs (nt-NPs) multiplexing. We term this method of ‘Topically Applied Surface Enhanced Resonance Raman Ratiometric Spectroscopy’ (“TAS3RS” (\tasers\) for short). TAS3RS successfully enabled the detection of tumor lesions in a murine model of human ovarian adenocarcinoma regardless of their size or localization. Tumors as small as 370 μm were detected as confirmed by bioluminescence imaging and histological staining. TAS3RS holds promise for intraoperative detection of microscopic residual tumors and could reduce recurrence rates in ovarian cancer and other diseases with peritoneal spread.

BACKGROUND Reconstruction after pan-plexus root avulsions often includes gracilis free functioning muscle transfer. For elbow flexion reconstruction, the free functioning muscle transfer distal tendon is inserted into the biceps tendon or more distally (i.e., flexor digitorum profundus/flexor pollicis longus tendons) for combined elbow and finger flexion; the theoretical drawback of the latter approach is weaker elbow flexion. The authors compared elbow flexion strength with a biceps tendon versus a flexor digitorum profundus/flexor pollicis longus tendon attachment to determine which insertion point resulted in better elbow flexion. METHODS Thirty-nine patients underwent free functioning muscle transfer with either a biceps tendon or a distal attachment. Groups were compared on postoperative elbow flexion strength, preoperative and postoperative Disabilities of the Arm, Shoulder, and Hand questionnaire scores, range of motion, and other surgical and demographic characteristics. A biomechanical analysis simulating different tendon attachments determined which reconstruction resulted in optimal elbow flexion mechanics. RESULTS Distal tendon attachment was associated with M3 or M4 elbow flexion and greater range of motion compared with the biceps tendon attachment (p < 0.05). There were no statistically significant improvements in Disabilities of the Arm, Shoulder, and Hand questionnaire scores. Biomechanical analysis demonstrated that all distal tendon attachments studied generated a 15 to 30 percent greater torque compared with the biceps tendon attachment; this was true for attachments either at the flexor digitorum profundus/flexor pollicis longus tendon, or directly at the radius at 10 cm or 15 cm from the elbow axis of rotation. CONCLUSIONS The flexor digitorum profundus/flexor pollicis longus tendon attachment of the gracilis free functioning muscle transfer distal tendon was superior in achieving elbow flexion strength. Patients with only elbow flexion reconstruction may also benefit from a flexor digitorum profundus/flexor pollicis longus tendon attachment or from a more distal attachment to the radius. CLINICAL QUESTION/LEVEL OF EVIDENCE Therapeutic, III.

Here we demonstrate a novel application of ‘surface enhanced resonance Raman scattering nanoparticles’ (SERRS NPs) for imaging breast cancer (BC) lung metastases with much higher precision than currently feasible. The BC lung metastasis mouse model was established by intravenous injection of LM2 cells. SERRS NPs conjugated with ALT-836, an anti-tissue factor (TF) monoclonal antibody, were administered to these mice intravenously and subjected to Raman imaging to visualize the expression of TF both in vivo and ex vivo. Raman imaging indicated marked uptake of αTF-SERRS NPs by the lung metastases (n=4) compared to isotype (n=4) and blocking controls (n=3). Conversely, little uptake of αTF-SERRS NPs was observed in the lungs of healthy mice. Successful detection and delineation of pulmonary micrometastatic lesions as small as 200 μm, corroborated by histology, immunohistochemistry, and bioluminescence imaging (BLI) confirmed the suitability of both TF as a target and αTF-SERRS NPs as an effective contrast agent for imaging breast cancer lung metastases. Further advancements of this technique in the form of Raman endoscopes coupled with ultrabright SERRS NPs developed in this work could lead to minimally invasive detection and resection of lung metastases.

Magnetic resonance imaging (MRI) is a fundamental tool for the noninvasive evaluation of liver pathologies. The anatomical position, the mobility with respiration, and the peculiar hemodynamics of the liver present several issues that have taken advantage from technological advances in MRI technology in the last decade, in particular regarding hardware, software, and contrast media (CM). We provided an overview of these topics in the first part of the review. MRI pulse sequences of clinical use in liver imaging can be divided into (1) T2-weighted sequences, for detection and characterization of focal and diffuse lesions without contrast, (2) T1-weighted sequences, with or without contrast injection, for further lesion characterization, and (3) additional techniques, such as diffusion-weighted imaging (DWI) and magnetic resonance cholangiopancreatography (MRCP). Most sequences have different names and abbreviations, and the reader is invited to find the respective commercial name specific for each vendor in appropriate references.1 The use of CM adds information on function and vascularity of liver parenchyma and focal lesions. The optimization of the technique for postcontrast studies and the basic concepts for characterization of focal liver lesions are briefly discussed in the following sections. MRI provides several advanced techniques for quantitative evaluation of diffuse liver disease. The complexity of these techniques is beyond the scope of this review, and only a brief description on magnetic resonance elastography (MRE), fat and iron quantification would be provided. At the end of our overview, we would discuss strategies to build basic liver protocols.

Nanoparticles labeled with radiometals enable whole-body nuclear imaging and therapy. Though chelating agents are commonly used to radiolabel biomolecules, nanoparticles offer the advantage of attaching a radiometal directly to the nanoparticle itself without the need of such agents. We previously demonstrated that direct radiolabeling of silica nanoparticles with hard, oxophilic ions, such as the positron emitters zirconium-89 and gallium-68, is remarkably efficient. However, softer radiometals, such as the widely employed copper-64, do not stably bind to the silica matrix and quickly dissociate under physiological conditions. Here, we overcome this limitation through the use of silica nanoparticles functionalized with a soft electron-donating thiol group to allow stable attachment of copper-64. This approach significantly improves the stability of copper-64 labeled thiol-functionalized silica nanoparticles relative to native silica nanoparticles, thereby enabling in vivo PET imaging, and may be translated to other softer radiometals with affinity for sulfur. The presented approach expands the application of silica nanoparticles as a platform for facile radiolabeling with both hard and soft radiometal ions.

Deltoid paralysis following isolated axillary nerve injury can be managed with triceps motor branch transfer or interpositional grafting. No consensus exists on the treatment that results in superior deltoid function. The purpose of this study was to review the authors' experience with axillary nerve injury management and compare functional outcomes following these two treatment options.Twenty-nine adult isolated axillary nerve injury patients that had either interpositional nerve grafting or triceps motor branch transfer with greater than 1 year of follow-up between 2002 and 2013 were reviewed for demographic and clinical factors and functional outcomes of deltoid reinnervation, including clinical examination (shoulder abduction and forward flexion graded by the Medical Research Council system) and electromyographic recovery. Disabilities of the Arm, Shoulder, and Hand scale grades were also compared.Twenty-one patients had a triceps motor transfer and eight had interpositional nerve grafting. At a mean follow-up of 22 months, Medical Research Council scores were greater in the grafting group compared with the nerve transfer group (4.3 versus 3.0), and more graft patients achieved useful deltoid function (Medical Research Council score ≥3) recovery (100 percent versus 62 percent); however, both groups had similar improvement in self-reported disability: change in Disabilities of the Arm, Shoulder, and Hand score of 11 following nerve transfer versus 15 following nerve graft.Although the question of nerve transfer versus grafting for restoration of axillary nerve function is controversial, this study demonstrates that grafting can result in good objective functional outcomes, particularly during an earlier time course after injury. This question requires further investigation in a larger, prospective patient population.Therapeutic, III.

Cancers of the gastrointestinal (GI) tract are among the most frequent and most lethal cancers worldwide. An important reason for this high mortality is that early disease is typically asymptomatic, and patients often present with advanced, incurable disease. Even in high-risk patients who routinely undergo endoscopic screening, lesions can be missed due to their small size or subtle appearance. Thus, current imaging approaches lack the sensitivity and specificity to accurately detect incipient GI tract cancers. Here we report our finding that a single dose of a high-sensitivity surface-enhanced resonance Raman scattering nanoparticle (SERRS-NP) enables reliable detection of precancerous GI lesions in animal models that closely mimic disease development in humans. Some of these animal models have not been used previously to evaluate imaging probes for early cancer detection. The studies were performed using a commercial Raman imaging system, a newly developed mouse Raman endoscope, and finally a clinically applicable Raman endoscope for larger animal studies. We show that this SERRS-NP-based approach enables robust detection of small, premalignant lesions in animal models that faithfully recapitulate human esophageal, gastric, and colorectal tumorigenesis. This method holds promise for much earlier detection of GI cancers than currently possible and could lead therefore to marked reduction of morbidity and mortality of these tumor types.

Purpose To report the clinical outcomes and describe the surgical technique of triceps muscle reinnervation using 2 different distal nerve transfers: the flexor carpi ulnaris (FCU) fascicle of the ulnar nerve and the posterior branch of the axillary nerve (PBAN) to the triceps nerve branch. Methods A retrospective review of patients undergoing FCU fascicle of ulnar nerve or PBAN to triceps nerve branch transfer was performed. Outcome measures included preoperative and postoperative modified British Medical Research Council (MRC) score, EMG results, and complications. Results Between September 2003 and April 2017, 6 patients were identified. Four patients with a traumatic upper trunk and posterior cord palsy underwent ulnar nerve fascicle to triceps nerve transfer. Two patients with a recovering upper trunk following a pan-brachial plexus palsy underwent PBAN to triceps nerve branch transfer. The median age was 30.0 years (range, 18–68 years). Surgery was performed at a median of 6.9 months (range, 5.0–8.9 months) postinjury, with a median follow-up of 18.4 months (range, 7.6–176.3) months. Before surgery, 4 patients exhibited grade M0 and 2 patients exhibited grade M1 triceps strength. Four patients had M5 donor muscle strength and 2 had grade M4. Postoperatively, 4 patients regained MRC grade M4 triceps muscle strength, 1 regained M3, and 1 regained M2. There was no noticeable donor muscle weakness. Conclusions Nerve fascicles to the FCU and PBAN are viable options for obtaining meaningful triceps muscle recovery in a select group of patients. Type of study/level of evidence Therapeutic V.

Ambient inhalable particulate matter (PM) is a serious health concern worldwide, but especially so in China where high PM concentrations affect huge populations. Atmospheric processes and emission sources cause spatial and temporal variations in PM concentration and chemical composition, but their influence on the toxicological characteristics of PM are still inadequately understood.In this study, we report an extensive chemical and toxicological characterization of size-segregated urban air inhalable PM collected in August and October 2013 from Nanjing, and assess the effects of atmospheric processes and likely emission sources. A549 human alveolar epithelial cells were exposed to day- and nighttime PM samples (25, 75, 150, 200, 300 mu g/ml) followed by analyses of cytotoxicity, genotoxicity, cell cycle, and inflammatory response.PM10-2.5 and PM0.2 caused the greatest toxicological responses for different endpoints, illustrating that particles with differing size and chemical composition activate distinct toxicological pathways in A549 cells. PM10-2.5 displayed the greatest oxidative stress and genotoxic responses; both were higher for the August samples compared with October. In contrast, PM0.2 and PM2.5-1.0 samples displayed high cytotoxicity and substantially disrupted cell cycle; August samples were more cytotoxic whereas October samples displayed higher cell cycle disruption. Several components associated with combustion, traffic, and industrial emissions displayed strong correlations with these toxicological responses. The lower responses for PM1.0-0.2 compared to PM0.2 and PM2.5-1.0 indicate diminished toxicological effects likely due to aerosol aging and lower proportion of fresh emission particles rich in highly reactive chemical components in the PM1.0-0.2 fraction.Different emission sources and atmospheric processes caused variations in the chemical composition and toxicological responses between PM fractions, sampling campaigns, and day and night. The results indicate different toxicological pathways for coarse-mode particles compared to the smaller particle fractions with typically higher content of combustion-derived components. The variable responses inside PM fractions demonstrate that differences in chemical composition influence the induced toxicological responses.

The dismal prognosis of patients with malignant brain tumors such as glioblastoma multiforme (GBM) is attributed mostly to their diffuse growth pattern and early microscopic tumor spread to distant regions of the brain. Because the microscopic tumor foci cannot be visualized with current imaging modalities, it remains impossible to direct treatments optimally. Here we explored the ability of integrin-targeted surface-enhanced resonance Raman spectroscopy (SERRS) nanoparticles to depict the true tumor extent in a GBM mouse model that closely mimics the pathology in humans. The recently developed SERRS-nanoparticles have a sensitivity of detection in the femtomolar range. An RGD-peptide-conjugated version for integrin-targeting (RGD-SERRS) was compared directly to its non-targeted RAD-SERRS control in the same mice via Raman multiplexing. Pre-blocking with RGD peptide before injection of RGD-SERRS nanoparticles was used to verify the specificity of integrin-targeting. In contrast to the current belief that the enhanced permeability and retention (EPR) effect results in a baseline uptake of nanoparticles regardless of their surface chemistry, integrin-targeting was shown to be highly specific, with markedly lower accumulation after pre-blocking. While the non-targeted SERRS particles enabled delineation of the main tumor, the RGD-SERRS nanoparticles afforded a major improvement in visualization of the true extent and the diffuse margins of the main tumor. This included the detection of unexpected tumor areas distant to the main tumor, tracks of migrating cells of 2-3 cells in diameter, and even isolated distant tumor cell clusters of less than 5 cells. This Raman spectroscopy-based nanoparticle-imaging technology holds promise to allow high precision visualization of the true extent of malignant brain tumors.

Highlights • Both the 2D and 3D handheld MSOT systems hold promise for clinical translation. • The 2D MSOT handheld probe provides high resolution, sensitivity, accuracy and reliability. • The 3D MSOT handheld probe adds unique 3D capabilities and shortens imaging time., A handheld approach to optoacoustic imaging is essential for the clinical translation. The first 2- and 3-dimensional handheld multispectral optoacoustic tomography (MSOT) probes featuring real-time unmixing have recently been developed. Imaging performance of both probes was determined in vitro and in a brain melanoma metastasis mouse model in vivo. T1-weighted MR images were acquired for anatomical reference. The limit of detection of melanoma cells in vitro was significantly lower using the 2D than the 3D probe. The signal decrease was more profound in relation to depth with the 3D versus the 2D probe. Both approaches were capable of imaging the melanoma tumors qualitatively at all time points. Quantitatively, the 2D approach enabled closer anatomical resemblance of the tumor compared to the 3D probe, particularly at depths beyond 3 mm. The 3D probe was shown to be superior for rapid 3D imaging and, thus, holds promise for more superficial target structures.

OBJECTIVEReconstructive options for brachial plexus lesions continue to expand and improve. The purpose of this study was to evaluate the prevalence and quality of restored elbow extension in patients with brachial plexus injuries who underwent transfer of the spinal accessory nerve to the motor branch of the radial nerve to the long head of the triceps muscle with an intervening autologous nerve graft and to identify patient and injury factors that influence functional triceps outcome.METHODSA total of 42 patients were included in this retrospective review. All patients underwent transfer of the spinal accessory nerve to the motor branch of the radial nerve to the long head of the triceps muscle as part of their reconstruction plan after brachial plexus injury. The primary outcome was elbow extension strength according to the modified Medical Research Council muscle grading scale, and signs of triceps muscle recovery were recorded using electromyography.RESULTSWhen evaluating the entire study population (follow-up range 12–45 months, mean 24.3 months), 52.4% of patients achieved meaningful recovery. More specifically, 45.2% reached Grade 0 or 1 recovery, 19.1% obtained Grade 2, and 35.7% improved to Grade 3 or better. The presence of a vascular injury impaired functional outcome. In the subgroup with a minimum follow-up of 20 months (n = 26), meaningful recovery was obtained by 69.5%. In this subgroup, 7.7% had no recovery (Grade 0), 19.2% had recovery to Grade 1, and 23.1% had recovery to Grade 2. Grade 3 or better was reached by 50% of patients, of whom 34.5% obtained Grade 4 elbow extension.CONCLUSIONSTransfer of the spinal accessory nerve to the radial nerve branch to the long head of the triceps muscle with an interposition nerve graft is an adequate option for restoration of elbow extension, despite the relatively long time required for reinnervation. The presence of vascular injury impairs functional recovery of the triceps muscle, and the use of shorter nerve grafts is recommended when and if possible.

Chelator-free nanoparticles for intrinsic radiolabeling are highly desirable for whole-body imaging and therapeutic applications. Several reports have successfully demonstrated the principle of intrinsic radiolabeling. However, the work done to date has suffered from much of the same specificity issues as conventional molecular chelators, insofar as there is no singular nanoparticle substrate that has proven effective in binding a wide library of radiosotopes. Here we present amorphous silica nanoparticles as general substrates for chelator-free radiolabeling and demonstrate their ability to bind six medically relevant isotopes of various oxidation states with high radiochemical yield. We provide strong evidence that the stability of the binding correlates with the hardness of the radioisotope, corroborating the proposed operating principle. Intrinsically labeled silica nanoparticles prepared by this approach demonstrate excellent in vivo stability and efficacy in lymph node imaging.

OBJECT Transfer of the triceps motor branch has been used for treatment of isolated axillary nerve palsy in the adult population. However, there are no published data on the effectiveness of this procedure in the pediatric population with traumatic injuries. The authors reviewed demographics and outcomes in their series of pediatric patients who underwent this procedure. METHODS Six patients ranging in age from 10 to 17 years underwent triceps motor branch transfer for the treatment of isolated axillary nerve injuries between 4 and 8 months after the inciting injury. Deltoid muscle strength was evaluated using the modified British Medical Research Council (MRC) grading system. Shoulder abduction at last follow-up was measured. RESULTS The mean duration of follow-up was 38 months. The average postoperative MRC grading of deltoid muscle strength was 3.6 ± 1.3. The median MRC grade was 4. One patient who did not achieve an MRC grade of 3 suffered multiple injuries from high-velocity trauma. Unlike in the adult population, age, body mass index of the patient, and delay from injury to surgery were not significant factors affecting the outcome of the procedure. CONCLUSIONS In the pediatric population with traumatic injuries, isolated axillary nerve injury treated with triceps motor branch transfer can result in good outcomes.

Recently, surface-enhanced Raman scattering (SERS) nanoprobes (NPs) have shown promise in the field of cancer imaging due to their unparalleled signal specificity and high sensitivity. Here we report the development of a DNA aptamer targeted SERS NP. Recently, aptamers are being investigated as a viable alternative to more traditional antibody targeting due to their low immunogenicity and low cost of production. We developed a strategy to functionalize SERS NPs with DNA aptamers, which target Mucin1 (MUC1) in human breast cancer (BC). Thorough in vitro characterization studies demonstrated excellent serum stability and specific binding of the targeted NPs to MUC1. In order to test their in vivo targeting capability, we co-injected MUC1-targeted SERS NPs, and as controls non-targeted and blocked MUC1-targeted SERS NPs in BC xenograft mouse models. A two-tumor mouse model with differential expression of MUC1 (MDA-MB-468 and MDA-MB-453) was used to control for active versus passive targeting in the same animals. The results showed that the targeted SERS NPs home to the tumors via active targeting of MUC1, with low levels of passive targeting. We expect this strategy to be an advantageous alternative to antibody-based targeting and useful for targeted imaging of tumor extent, progression, and therapeutic response.

Gold nanoparticles have unique properties that are highly dependent on their shape and size. Synthetic methods that enable precise control over nanoparticle morphology currently require shape-directing agents such as surfactants or polymers that force growth in a particular direction by adsorbing to specific crystal facets. These auxiliary reagents passivate the nanoparticles' surface, and thus decrease their performance in applications like catalysis and surface-enhanced Raman scattering. Here, a surfactant- and polymer-free approach to achieving high-performance gold nanoparticles is reported. A theoretical framework to elucidate the growth mechanism of nanoparticles in surfactant-free media is developed and it is applied to identify strategies for shape-controlled syntheses. Using the results of the analyses, a simple, green-chemistry synthesis of the four most commonly used morphologies: nanostars, nanospheres, nanorods, and nanoplates is designed. The nanoparticles synthesized by this method outperform analogous particles with surfactant and polymer coatings in both catalysis and surface-enhanced Raman scattering.

Purpose To report our technique and experience with use of free functioning muscle transfer (FFMT) in reconstruction of traumatic brachial plexus injuries (BPIs) in children as well as its complications and outcomes. Methods Twelve patients with complete BPI underwent FFMT for reconstruction between 2000 and 2012. Eight had single-stage gracilis transfer for restoration of elbow flexion, and 4 children had double free gracilis muscle transfer for restoration of elbow flexion and prehension. Mean duration of follow-up was 27 months (range, 14–55 mo). Results Eleven out of 12 patients achieved at least M3 elbow flexion, with 8 patients achieving M4 or greater elbow flexion. Eight of 12 patients had nerve transfers to the musculocutaneous nerve. Mean active elbow arc of motion was 79° (range, 30°–130°). Two patients aged 8 and 11 years with open growth plates developed elbow joint contractures, which limited range of motion, but they recovered M4 and M5 elbow flexion strength. Conclusions FFMTs can result in good outcomes following reconstruction for traumatic BPI. The use of FFMT should be carefully considered in children prior to skeletal maturity because of the risk of the development of an elbow flexion contracture. Type of study/level of evidence Therapeutic IV.

The ability to control the movement of nanoparticles remotely and with high precision would have far-reaching implications in many areas of nanotechnology. We have designed a unique dynamic magnetic field (DMF) generator that can induce rotational movements of superparamagnetic iron oxide nanoparticles (SPIONs). We examined whether the rotational nanoparticle movement could be used for remote induction of cell death by injuring lysosomal membrane structures. We further hypothesized that the shear forces created by the generation of oscillatory torques (incomplete rotation) of SPIONs bound to lysosomal membranes would cause membrane permeabilization, lead to extravasation of lysosomal contents into the cytoplasm, and induce apoptosis. To this end, we covalently conjugated SPIONs with antibodies targeting the lysosomal protein marker LAMP1 (LAMP1-SPION). Remote activation of slow rotation of LAMP1-SPIONs significantly improved the efficacy of cellular internalization of the nanoparticles. LAMP1-SPIONs then preferentially accumulated along the membrane in lysosomes in both rat insulinoma tumor cells and human pancreatic beta cells due to binding of LAMP1-SPIONs to endogenous LAMP1. Further activation of torques by the LAMP1-SPIONs bound to lysosomes resulted in rapid decrease in size and number of lysosomes, attributable to tearing of the lysosomal membrane by the shear force of the rotationally activated LAMP1-SPIONs. This remote activation resulted in an increased expression of early and late apoptotic markers and impaired cell growth. Our findings suggest that DMF treatment of lysosome-targeted nanoparticles offers a noninvasive tool to induce apoptosis remotely and could serve as an important platform technology for a wide range of biomedical applications.

Introduction: The current literature indicates that neurologic injuries during shoulder surgery occur infrequently and result in little if any morbidity. The purpose of this study was to review one institution’s experience treating patients with iatrogenic nerve injuries after shoulder surgery. Methods: A retrospective review of the records of patients evaluated in a brachial plexus specialty clinic from 2000 to 2010 identified twenty-six patients with iatrogenic nerve injury secondary to shoulder surgery. The records were reviewed to determine the operative procedure, time to presentation, findings on physical examination, treatment, and outcome. Results: The average age was forty-three years (range, seventeen to seventy-two years), and the average delay prior to referral was 5.4 months (range, one to fifteen months). Seven nerve injuries resulted from open procedures done to treat instability; nine, from arthroscopic surgery; four, from total shoulder arthroplasty; and six, from a combined open and arthroscopic operation. The injury occurred at the level of the brachial plexus in thirteen patients and at a terminal nerve branch in thirteen. Fifteen patients (58%) did not recover nerve function after observation and required surgical management. A structural nerve injury (laceration or suture entrapment) occurred in nine patients (35%), including eight of the thirteen who presented with a terminal nerve branch injury and one of the thirteen who presented with an injury at the level of the brachial plexus. Conclusions: Nerve injuries occurring during shoulder surgery can produce severe morbidity and may require surgical management. Injuries at the level of a peripheral nerve are more likely to be surgically treatable than injuries of the brachial plexus. A high index of suspicion and early referral and evaluation should be considered when evaluating patients with iatrogenic neurologic deficits after shoulder surgery.

We report a nerve transfer to the triceps using the posterior branch of the axillary nerve to restore elbow extension in an 18-year-old woman with a C7-T1 injury. Elbow extension strength improved from M0 to M4, whereas deltoid strength was minimally affected. Her Disabilities of the Arm, Shoulder and Hand score improved 14 points. This method may be considered for restoring triceps function in lower pattern brachial plexus injury.