Correlation of Narrative Evaluations to Clerkship Grades Using Statistical Sentiment Analysis.

Introduction: Narrative evaluations are essential components of medical student assessment. This study evaluated how well narrative clerkship evaluation word choice correlated with an assigned letter grade. Methods: One hundred clerkship evaluations, 50 from family medicine (FM) and 50 from internal medicine (IM), with even distribution of "Honors" and "Near-Honors" among medical students that graduated in 2020 from the Oregon Health and Science University (OHSU) were examined. A textual sentiment analysis, which evaluates positive and negative word choice, was used to determine each evaluation's collective sentiment. An average sentiment score and character count were calculated for Honors and Near-Honors evaluations from both clerkship disciplines. Sentiment word totals were used to form "word clouds" that highlight the most frequent word selections. Results: While sentiment scores positively correlated with the assigned grade, there was no statistically significant difference between the average sentiment scores among Honors and Near Honors graded evaluations within the FM or IM clerkship evaluation sets. There was no significant difference in evaluation character length among the assigned grades. Among FM evaluations, "outstanding" and "excellent" were the two most common sentiment words used in both Honors and Near-Honors. Among IM evaluations, outstanding and excellent were most commonly used in Honors evaluations, while "excellent" and "good" were most common in Near-Honors. Conclusion: This study outlines a novel text analysis method for analyzing narrative evaluation association with assigned grade that other institutions can utilize. Sentiment word choices are not significantly different among Honors and Near Honors clerkship narrative evaluations. Supplementary Information: The online version contains supplementary material available at 10.1007/s40670-022-01654-2.

Antiplasmodial activity of targeted zinc(II)-dipicolylamine complexes.

This study measured the antiplasmodial activity of nine zinc-dipicolylamine (ZnDPA) complexes against three strains of Plasmodium falciparum, the causative parasite of malaria. Growth inhibition assays showed significant activity against all tested strains, with 50% inhibitory concentrations between 5 and 600nM and almost no toxic effect against host cells including healthy red blood cells. Fluorescence microscopy studies with a green-fluorescent ZnDPA probe showed selective targeting of infected red blood cells. The results suggest that ZnDPA coordination complexes are promising antiplasmodial agents with potential for targeted malaria treatment.

Preassembled Fluorescent Multivalent Probes for the Imaging of Anionic Membranes.

A new self-assembly process known as Synthavidin (synthetic avidin) technology was used to prepare targeted probes for near-infrared fluorescence imaging of anionic membranes and cell surfaces, a hallmark of many different types of disease. The probes were preassembled by threading a tetralactam macrocycle with six appended zinc-dipicolylamine (ZnDPA) targeting units onto a linear scaffold with one or two squaraine docking stations to produce hexavalent or dodecavalent fluorescent probes. A series of liposome titration experiments showed that multivalency promoted stronger membrane binding by the dodecavalent probe. In addition, the dodecavalent probe exhibited turn-on fluorescence due to probe unfolding during fluorescence microscopy at the membrane surface. However, the dodecavalent probe also had a higher tendency to self-aggregate after membrane binding, leading to probe self-quenching under certain conditions. This self-quenching effect was apparent during fluorescence microscopy experiments that recorded low fluorescence intensity from anionic dead and dying mammalian cells that were saturated with the dodecavalent probe. Conversely, probe self-quenching was not a factor with anionic microbial surfaces, where there was intense fluorescence staining by the dodecavalent probe. A successful set of rat tumor imaging experiments confirmed that the preassembled probes have sufficient mechanical stability for effective in vivo imaging. The results demonstrate the feasibility of this general class of preassembled fluorescent probes for multivalent targeting, but fluorescence imaging performance depends on the specific physical attributes of the biomarker target, such as the spatial distance between different copies of the biomarker and the propensity of the probe-biomarker complex to self-aggregate.

Imaging and therapeutic applications of zinc(ii)-dipicolylamine molecular probes for anionic biomembranes.

This feature article describes the development of synthetic zinc(ii)-dipicolylamine (ZnDPA) receptors as selective targeting agents for anionic membranes in cell culture and living subjects. There is a strong connection between anionic cell surface charge and disease, and ZnDPA probes have been employed extensively for molecular imaging and targeted therapeutics. Fluorescence and nuclear imaging applications include detection of diseases such as cancer, neurodegeneration, arthritis, and microbial infection, and also quantification of cell death caused by therapy. Therapeutic applications include selective targeting of cytotoxic agents and drug delivery systems, photodynamic inactivation, and modulation of the immune system. The article concludes with a summary of expected future directions.

Pre-Assembly of Near-Infrared Fluorescent Multivalent Molecular Probes for Biological Imaging.

A programmable pre-assembly method is described and shown to produce near-infrared fluorescent molecular probes with tunable multivalent binding properties. The modular assembly process threads one or two copies of a tetralactam macrocycle onto a fluorescent PEGylated squaraine scaffold containing a complementary number of docking stations. Appended to the macrocycle periphery are multiple copies of a ligand that is known to target a biomarker. The structure and high purity of each threaded complex was determined by independent spectrometric methods and also by gel electrophoresis. Especially helpful were diagnostic red-shift and energy transfer features in the absorption and fluorescence spectra. The threaded complexes were found to be effective multivalent molecular probes for fluorescence microscopy and in vivo fluorescence imaging of living subjects. Two multivalent probes were prepared and tested for targeting of bone in mice. A pre-assembled probe with 12 bone-targeting iminodiacetate ligands produced more bone accumulation than an analogous pre-assembled probe with six iminodiacetate ligands. Notably, there was no loss in probe fluorescence at the bone target site after 24 h in the living animal, indicating that the pre-assembled fluorescent probe maintained very high mechanical and chemical stability on the skeletal surface. The study shows how this versatile pre-assembly method can be used in a parallel combinatorial manner to produce libraries of near-infrared fluorescent multivalent molecular probes for different types of imaging and diagnostic applications, with incremental structural changes in the number of targeting groups, linker lengths, linker flexibility, and degree of PEGylation.

Zinc(II)-Dipicolylamine Coordination Complexes as Targeting and Chemotherapeutic Agents for Leishmania major.

Cutaneous leishmaniasis is a neglected tropical disease that causes painful lesions and severe disfigurement. Modern treatment relies on a few chemotherapeutics with serious limitations, and there is a need for more effective alternatives. This study describes the selective targeting of zinc(II)-dipicolylamine (ZnDPA) coordination complexes toward Leishmania major, one of the species responsible for cutaneous leishmaniasis. Fluorescence microscopy of L. major promastigotes treated with a fluorescently labeled ZnDPA probe indicated rapid accumulation of the probe within the axenic promastigote cytosol. The antileishmanial activities of eight ZnDPA complexes were measured using an in vitro assay. All tested complexes exhibited selective toxicity against L. major axenic promastigotes, with 50% effective concentration values in the range of 12.7 to 0.3 µM. Similar toxicity was observed against intracellular amastigotes, but there was almost no effect on the viability of mammalian cells, including mouse peritoneal macrophages. In vivo treatment efficacy studies used fluorescence imaging to noninvasively monitor changes in the red fluorescence produced by an infection of mCherry-L. major in a mouse model. A ZnDPA treatment regimen reduced the parasite burden nearly as well as the reference care agent, potassium antimony(III) tartrate, and with less necrosis in the local host tissue. The results demonstrate that ZnDPA coordination complexes are a promising new class of antileishmanial agents with potential for clinical translation.

Phenoxide-Bridged Zinc(II)-Bis(dipicolylamine) Probes for Molecular Imaging of Cell Death.

Cell death is involved in many pathological conditions, and there is a need for clinical and preclinical imaging agents that can target and report cell death. One of the best known biomarkers of cell death is exposure of the anionic phospholipid phosphatidylserine (PS) on the surface of dead and dying cells. Synthetic zinc(II)-bis(dipicolylamine) (Zn2BDPA) coordination complexes are known to selectively recognize PS-rich membranes and act as cell death molecular imaging agents. However, there is a need to improve in vivo imaging performance by selectively increasing target affinity and decreasing off-target accumulation. This present study compared the cell death targeting ability of two new deep-red fluorescent probes containing phenoxide-bridged Zn2BDPA complexes. One probe was a bivalent version of the other and associated more strongly with PS-rich liposome membranes. However, the bivalent probe exhibited self-quenching on the membrane surface, so the monovalent version produced brighter micrographs of dead and dying cells in cell culture and also better fluorescence imaging contrast in two living animal models of cell death (rat implanted tumor with necrotic core and mouse thymus atrophy). An (111)In-labeled radiotracer version of the monovalent probe also exhibited selective cell death targeting ability in the mouse thymus atrophy model, with relatively high amounts detected in dead and dying tissue and low off-target accumulation in nonclearance organs. The in vivo biodistribution profile is the most favorable yet reported for a Zn2BDPA complex; thus, the monovalent phenoxide-bridged Zn2BDPA scaffold is a promising candidate for further development as a cell death imaging agent in living subjects.

Bacterial imaging and photodynamic inactivation using zinc(II)-dipicolylamine BODIPY conjugates.

Targeted imaging and antimicrobial photodynamic inactivation (PDI) are emerging methods for detecting and eradicating pathogenic microorganisms. This study describes two structurally related optical probes that are conjugates of a zinc(II)-dipicolylamine targeting unit and a BODIPY chromophore. One probe is a microbial targeted fluorescent imaging agent, mSeek, and the other is an oxygen photosensitizing analogue, mDestroy. The conjugates exhibited high fluorescence quantum yield and singlet oxygen production, respectively. Fluorescence imaging and detection studies examined four bacterial strains: E. coli, S. aureus, K. pneumonia, and B. thuringiensis vegetative cells and purified spores. The fluorescent probe, mSeek, is not phototoxic and enabled detection of all tested bacteria at concentrations of ∼100 CFU mL(-1) for B. thuringiensis spores, ∼1000 CFU mL(-1) for S. aureus and ∼10,000 CFU mL(-1) for E. coli. The photosensitizer analogue, mDestroy, inactivated 99-99.99% of bacterial samples and selectively killed bacterial cells in the presence of mammalian cells. However, mDestroy was ineffective against B. thuringiensis spores. Together, the results demonstrate a new two-probe strategy to optimize PDI of bacterial infection/contamination.

Fluorescence Imaging of Interscapular Brown Adipose Tissue in Living Mice.

Brown adipose tissue (BAT) plays a key role in energy expenditure and heat generation and is a promising target for diagnosing and treating obesity, diabetes and related metabolism disorders. While several nuclear and magnetic resonance imaging methods are established for detecting human BAT, there are no convenient protocols for high throughput imaging of BAT in small animal models. Here we disclose a simple but effective method for non-invasive optical imaging of interscapular BAT in mice using a micellar formulation of the commercially available deep-red fluorescent probe, SRFluor680. Whole-body fluorescence imaging of living mice shows extensive accumulation of the fluorescent probe in the interscapular BAT and ex vivo analysis shows 3.5-fold selectivity for interscapular BAT over interscapular WAT. Additional imaging studies indicate that SRFluor680 uptake is independent of mouse species and BAT metabolic state. The results are consistent with an unusual pharmacokinetic process that involves irreversible translocation of the lipophilic SRFluor680 from the micelle nanocarrier into the adipocytes within the BAT. Multimodal PET/CT and planar fluorescence/X-ray imaging of the same living animal shows co-localization of BAT mass signal reported by the fluorescent probe and BAT metabolism signal reported by the PET agent, 18F-FDG. The results indicate a path towards a new, dual probe molecular imaging paradigm that allows separate and independent non-invasive visualization of BAT mass and BAT metabolism in a living subject.

Evaluation of [¹¹¹In]-labeled zinc-dipicolylamine tracers for SPECT imaging of bacterial infection.

PURPOSE: This study prepared three structurally related zinc-dipicolylamine (ZnDPA) tracers with [(111)In] labels and conducted biodistribution and single-photon emission computed tomography/computed tomography (SPECT/CT) imaging studies of a mouse leg infection model. PROCEDURES: Two monovalent tracers, ZnDPA-[(111)In]DTPA and ZnDPA-[(111)In]DOTA, each with a single zinc-dipicolylamine targeting unit, and a divalent tracer, Bis(ZnDPA)-[(111)In]DTPA, with two zinc-dipicolylamine units were prepared. Organ biodistribution and SPECT and CT imaging studies were performed on living mice with a leg infection created by injection of clinically relevant Gram positive Streptococcus pyogenes. Fluorescent and luminescent Eu(3+)-labeled versions of these tracers were also prepared and used to measure relative affinity for the exterior membrane surface of bacterial cells and mimics of healthy mammalian cells. RESULTS: All three (111)In-labeled radiotracers were prepared with a radiopurity of >90 %. The biodistribution studies showed that the two monovalent tracers were cleared from the body through the liver and kidney, with retained percentage injected dose for all organs of <8 % at 20 h and infected leg target to non-target ratio (T/NT) ratio of ≤3.0. Clearance of the divalent tracer from the bloodstream was slower and primarily through the liver, with a retained percentage injected dose for all organs <37 % at 20 h and T/NT ratio rising to 6.2 after 20 h. The SPECT/CT imaging indicated the same large difference in tracer pharmacokinetics and higher accumulation of the divalent tracer at the site of infection. CONCLUSIONS: All three [(111)In]-ZnDPA tracers selectively targeted the site of a clinically relevant mouse infection model that could not be discerned by visual external inspection of the living animal. The highest target selectivity, observed with a divalent tracer equipped with two zinc-dipicolylamine targeting units, compares quite favorably with the imaging selectivities previously reported for other nuclear tracers that target bacterial cell surfaces. The tracer pharmacokinetics depended heavily on tracer molecular structure suggesting that it may be possible to rapidly fine tune the structural properties for optimized in vivo imaging performance and clinical translation.

Selective recognition of anionic cell membranes using targeted liposomes coated with zinc(ii)-bis(dipicolylamine) affinity units.

Zinc(ii)-bis(dipicolylamine) (Zn2BDPA) coated liposomes are shown to have high recognition selectivity towards vesicle and cell membranes with anionic surfaces. Robust synthetic methods were developed to produce Zn2BDPA-PEG-lipid conjugates with varying PEG linker chain length. One conjugate (Zn2BDPA-PEG2000-DSPE) was used in liposome formulations doped with the lipophilic near-infrared fluorophore DiR. Fluorescence cell microscopy studies demonstrated that the multivalent liposomes selectively and efficiently target bacteria in the presence of healthy mammalian cells and cause bacterial cell agglutination. The liposomes also exhibited selective staining of the surfaces of dead or dying human cancer cells that had been treated with a chemotherapeutic agent.

In vivo imaging using polymeric nanoparticles stained with near-infrared chemiluminescent and fluorescent squaraine catenane endoperoxide.

Polystyrene nanoparticles stained with squaraine catenane endoperoxide exhibit remarkably high chemiluminescence and enable optical imaging of biodistribution in living mice. Whole-body chemiluminescence imaging was much more effective than fluorescence at identifying lung accumulation of the nanoparticles.