Increased [18F]FDG uptake of radiation-induced giant cells: a single-cell study in lung cancer models.

Positron emission tomography (PET), a cornerstone in cancer diagnosis and treatment monitoring, relies on the enhanced uptake of fluorodeoxyglucose ([18F]FDG) by cancer cells to highlight tumors and other malignancies. While instrumental in the clinical setting, the accuracy of [18F]FDG-PET is susceptible to metabolic changes introduced by radiation therapy. Specifically, radiation induces the formation of giant cells, whose metabolic characteristics and [18F]FDG uptake patterns are not fully understood. Through a novel single-cell gamma counting methodology, we characterized the [18F]FDG uptake of giant A549 and H1299 lung cancer cells that were induced by radiation, and found it to be considerably higher than that of their non-giant counterparts. This observation was further validated in tumor-bearing mice, which similarly demonstrated increased [18F]FDG uptake in radiation-induced giant cells. These findings underscore the metabolic implications of radiation-induced giant cells, as their enhanced [18F]FDG uptake could potentially obfuscate the interpretation of [18F]FDG-PET scans in patients who have recently undergone radiation therapy.

Ultrasensitive and multiplexed tracking of single cells using whole-body PET/CT.

In vivo molecular imaging tools are crucially important for elucidating how cells move through complex biological systems; however, achieving single-cell sensitivity over the entire body remains challenging. Here, we report a highly sensitive and multiplexed approach for tracking upward of 20 single cells simultaneously in the same subject using positron emission tomography (PET). The method relies on a statistical tracking algorithm (PEPT-EM) to achieve a sensitivity of 4 becquerel per cell and a streamlined workflow to reliably label single cells with over 50 becquerel per cell of 18F-fluorodeoxyglucose (FDG). To demonstrate the potential of the method, we tracked the fate of more than 70 melanoma cells after intracardiac injection and found they primarily arrested in the small capillaries of the pulmonary, musculoskeletal, and digestive organ systems. This study bolsters the evolving potential of PET in offering unmatched insights into the earliest phases of cell trafficking in physiological and pathological processes and in cell-based therapies.

Efficient and multiplexed tracking of single cells using whole-body PET/CT.

In vivo molecular imaging tools are crucially important for elucidating how cells move through complex biological systems, however, achieving single-cell sensitivity over the entire body remains challenging. Here, we report a highly sensitive and multiplexed approach for tracking upwards of 20 single cells simultaneously in the same subject using positron emission tomography (PET). The method relies on a new tracking algorithm (PEPT-EM) to push the cellular detection threshold to below 4 Bq/cell, and a streamlined workflow to reliably label single cells with over 50 Bq/cell of 18F-fluorodeoxyglucose (FDG). To demonstrate the potential of method, we tracked the fate of over 70 melanoma cells after intracardiac injection and found they primarily arrested in the small capillaries of the pulmonary, musculoskeletal, and digestive organ systems. This study bolsters the evolving potential of PET in offering unmatched insights into the earliest phases of cell trafficking in physiological and pathological processes and in cell-based therapies.

Characterization of a Class A ß-Lactamase from Francisella tularensis (Ftu-1) Belonging to a Unique Subclass toward Understanding AMR.

ß-lactamase production with vast catalytic divergence in the pathogenic strain limits the antibiotic spectrum in the clinical environment. Class A carbapenemase shares significant sequence similarities, structural features, and common catalytic mechanisms although their resistance spectrum differs from class A ß-lactamase in carbapenem and monobactam hydrolysis. In other words, it limited the antibiotic treatment option against infection, causing carbapenemase-producing superbugs. Ftu-1 is a class A ß-lactamase expressed by the Francisella tularensis strain, a potent causative organism of tularemia. The chromosomally encoded class A ß-lactamase shares two conserved cysteine residues, a common characteristic of a carbapenemase, and a distinctive class in the phylogenetic tree. Complete biochemical and biophysical characterization of the enzyme was performed to understand the overall stability and environmental requirements to perform optimally. To comprehend the enzyme-drug interaction and its profile toward various chemistries of ß-lactam and ß-lactamase inhibitors, comprehensive kinetic and thermodynamic analyses were conducted using various ß-lactam drugs. The dynamic property of Ftu-1 ß-lactamase was also predicted using molecular dynamics (MD) simulation to compare its loop flexibility and ligand binding with other related class A ß-lactamases. Overall, this study fosters a comprehensive understanding of Ftu-1, proposed to be an intermediate class by characterizing its kinetic profiling, stability by biochemical and biophysical methodologies, and susceptibility profiling. This understanding would be beneficial for the design of new-generation therapeutics.

Preclinical Evaluation of 89Zr-Panitumumab for Biology-Guided Radiation Therapy.

PURPOSE: Biology-guided radiation therapy (BgRT) uses real-time line-of-response data from on-board positron emission tomography (PET) detectors to guide beamlet delivery during therapeutic radiation. The current workflow requires 18F-fluorodeoxyglucose (FDG) administration daily before each treatment fraction. However, there are advantages to reducing the number of tracer injections by using a PET tracer with a longer decay time. In this context, we investigated 89Zr-panitumumab (89Zr-Pan), an antibody PET tracer with a half-life of 78 hours that can be imaged for up to 9 days using PET. METHODS AND MATERIALS: The BgRT workflow was evaluated preclinically in mouse colorectal cancer xenografts (HCT116) using small-animal positron emission tomography/computed tomography (PET/CT) for imaging and image-guided kilovoltage conformal irradiation for therapy. Mice (n = 5 per group) received 7 MBq of 89Zr-Pan as a single dose 2 weeks after tumor induction, with or without fractionated radiation therapy (RT; 6 × 6.6 Gy) to the tumor region. The mice were imaged longitudinally to assess the kinetics of the tracer over 9 days. PET images were then analyzed to determine the stability of the PET signal in irradiated tumors over time. RESULTS: Mice in the treatment group experienced complete tumor regression, whereas those in the control group were killed because of tumor burden. PET imaging of 89Zr-Pan showed well-delineated tumors with minimal background in both groups. On day 9 postinjection, tumor uptake of 89Zr-Pan was 7.2 ± 1.7 in the control group versus 5.2 ± 0.5 in the treatment group (mean percentage of injected dose per gram of tissue [%ID/g] ± SD; P = .07), both significantly higher than FDG uptake (1.1 ± 0.5 %ID/g) 1 hour postinjection. To assess BgRT feasibility, the clinical eligibility criteria was computed using human-equivalent uptake values that were extrapolated from preclinical PET data. Based on this semiquantitative analysis, BgRT may be feasible for 5 consecutive days after a single 740-MBq injection of 89Zr-Pan. CONCLUSIONS: This study indicates the potential of long-lived antibody-based PET tracers for guiding clinical BgRT.

Real-time optical oximetry during FLASH radiotherapy using a phosphorescent nanoprobe.

The rapid depletion of oxygen during irradiation at ultra-high dose rate calls for tissue oximeters capable of high temporal resolution. This study demonstrates a water-soluble phosphorescent nanoprobe and fiber-coupled instrument, which together are used to measure the kinetics of oxygen depletion at 200 Hz during irradiation of in vitro solutions.

Inhibitory effect of selected Indian honey on colon cancer cell growth by inducing apoptosis and targeting the ß-catenin/Wnt pathway.

Colon cancer is the most prevalent cause of death from cancer across the globe. Although chemotherapy drugs are predominantly used, their toxicity always remains a cause of concern. As an alternative to synthetic drugs, natural compounds or nutraceuticals are comparatively less toxic. Honey is widely used across different cultures as an alternative form of medicine. It represents a prominent source of plant-phenolic compounds and there is demonstrable evidence of its anti-oxidant and anti-microbial activities. The aim of the present work was to investigate the anti-proliferative effect of some Indian honeys and analyze their mechanism of action in colon cancer. In order to establish the composition-activity relationship, we evaluated the bioactive components present in selected honey samples by GC-MS and HPLC analysis. Indian honey samples showed a significant inhibitory impact on cell growth by restricting cell proliferation, causing apoptosis, and restricting the cell cycle in the G2/M phase specifically for colon cancer cells. The apoptotic activities, as imparted by the honey samples, were established by Annexin V/PI staining, real-time PCR, and immunoblot analyses. The treated cells showed increased expressions of p53 and caspases 3, 8, and 9, thus indicating the involvement of both extrinsic and intrinsic apoptotic pathways. The honey samples were also found to inhibit the ß-catenin/Wnt pathway. In the next phase of the study, the efficacy of these honey samples was evaluated in colon carcinoma induced SD-rats. Overall, these findings demonstrated that selected Indian honeys could be established as effective nutraceuticals for the prevention as well as cure of colon cancer.

A highly transparent tri-polymer complexin situhydrogel of HA, collagen and four-arm-PEG as potential vitreous substitute.

There is a requirement of removal and replacement of vitreous for various ophthalmic diseases, e.g. retinopathy and retinal detachment. Clinical tamponades, e.g. silicone oil and fluorinated gases are used but limited due to their toxicity and some complications. A lot of polymer-based materials have been tested and proposed as vitreous substitute, but till date, there is no ideal vitreous substitute available. Thus, it requires to develop an improved vitreous substitute which will be highly suitable for vitreous replacement. We have developed tri-polymer complexin situhydrogels by crosslinking among hyaluronic acid (HA), collagen (Coll) and four-arm-polyethylene glycol (PEG). All the developed hydrogels are biocompatible with NIH 3T3 mouse fibroblast cells, having pH in the range 7-7.44 and refractive index in the range 1.333-1.345. The developed hydrogels are highly transparent, showing transmittance >97%. FTIR study shows that the hydrogel was crosslinked by amide bond formation between HA and PEG, and between Coll and PEG. The rheological study shows that all the developed hydrogels exhibit viscoelastic behavior and all the hydrogels have storage modulus values (>100 pa) which is greater than loss modulus values-indicating sufficient elasticity for vitreous application. The elastic nature of the hydrogel increases with the increase in PEG concentration. The gel is formed in between 2 and 3 min-indicating its applicationin situ. The viscosity of the developed hydrogels shows shear thinning behavior. The pre-gel solution of the hydrogel is injectable through a 22 G needle-indicating its applicationin situthrough vitrectomy surgery. All the hydrogels are hydrophilic and have water content of 96% approximately. Thus, the results show the positive properties for its application as a potential vitreous substitute.

Betel leaf extract and its major component hydroxychavicol promote osteogenesis and alleviate glucocorticoid-induced osteoporosis in rats.

Piper betle leaves possess several ethnomedicinal properties and are immensely used in traditional medicinal practices in regions of Asian and African subcontinents. However, their effects in treating skeletal complications are least known. In this study, we evaluated cellular and molecular effects of betel leaf extract (BLE) and its major phytoconstituent, hydroxychavicol (HCV) in promoting osteogenesis in vitro and alleviating glucocorticoid induced osteoporosis (GIO) in vivo. Both BLE and HCV markedly stimulated osteoblast differentiation of C3H10T1/2 cells with increased expression of RUNX2 and osteopontin through the GSK-3ß/ß-catenin-signaling pathway. Also, oral administration of BLE and HCV in GIO rats resulted in restoration of bone mass and tissue microarchitecture. Thus, with our findings we conclude that BLE and HCV promote osteogenesis of C3H10T1/2 cells via the GSK-3ß/ß-catenin pathway and alleviate GIO in rats.

Robust expression of LINE-1 retrotransposon encoded proteins in oral squamous cell carcinoma.

BACKGROUND: Oral Squamous Cell Carcinoma (OSCC) results from a series of genetic alteration in squamous cells. This particular type of cancer considers one of the most aggressive malignancies to control because of its frequent local invasions to the regional lymph node. Although several biomarkers have been reported, the key marker used to predict the behavior of the disease is largely unknown. Here we report Long INterpersed Element-1 (LINE1 or L1) retrotransposon activity in post-operative oral cancer samples. L1 is the only active retrotransposon occupying around 17% of the human genome with an estimated 500,000 copies. An active L1 encodes two proteins (L1ORF1p and L1ORF2p); both of which are critical in the process of retrotransposition. Several studies report that the L1 retrotransposon is highly active in many cancers. L1 activity is generally determined by assaying L1ORF1p because of its high expression and availability of the antibody. However, due to its lower expression and unavailability of a robust antibody, detection of L1ORF2p has been limited. L1ORF2p is the crucial protein in the process of retrotransposition as it provides endonuclease and reverse transcriptase (RT) activity. METHODS: Immunohistochemistry and Western blotting were performed on the post-operative oral cancer samples and murine tissues. RESULTS: Using in house novel antibodies against both the L1 proteins (L1ORF1p and L1ORF2p), we found L1 retrotransposon is extremely active in post-operative oral cancer tissues. Here, we report a novel human L1ORF2p antibody generated using an 80-amino-acid stretch from the RT domain, which is highly conserved among different species. The antibody detects significant L1ORF2p expression in human oral squamous cell carcinoma (OSCC) samples and murine germ tissues. CONCLUSIONS: We report exceptionally high L1ORF1p and L1ORF2p expression in post-operative oral cancer samples. The novel L1ORF2p antibody reported in this study will serve as a useful tool to understand why L1 activity is deregulated in OSCC and how it contributes to the progression of this particular cancer. Cross-species reactivity of L1ORF2p antibody due to the conserved epitope will be useful to study the retrotransposon biology in mice and rat germ tissues.

Synthesis and cytotoxicity of novel 14α-O-(andrographolide-3-subsitutedisoxazole-5-carboxylate) derivatives.

Simple and efficient method was established for the synthesis of a new family of 14α-O-(andrographolide-3-subsitutedisoxazole-5-carboxylate) derivatives (10a-j) from naturally occurring andrographolide (1) by selective esterification with propiolic acid at C-14 using protection and deprotection strategy followed by metal free 1,3-dipolar cycloaddition with aryl nitrile oxides. All the synthesised derivatives were tested for their cytotoxicity against HCT-15, HeLa and DU145 cell lines. Most of the compounds exhibited improved cytotoxic activity compared to the parent molecule andrographolide (1), as the compounds 10b, 10c, 10i, 10j, 11d and 11f showed significant cytotoxicity against three cancer cell lines. Except the compound 10b and 11d, all the compounds did not inhibit the normal cell line (VERO). Based on these studies isoxazole ester derivatives at C-14 of andrographolide with various substitutions promoting anticancer activities and better safety profiles. Further studies in this direction with improved water solubility and oral bioavailability are in progress in future.

Ligand substituent effect on the cytotoxicity activity of two new copper(ii) complexes bearing 8-hydroxyquinoline derivatives: validated by MTT assay and apoptosis in MCF-7 cancer cell line (human breast cancer).

In this study, we have examined the effect of ligand substituent on the structure-cytotoxicity relationships of the MCF-7 cancer cell line (human breast cancer), by two copper(ii) complexes {[Cu(qmbn)(Hqmba)(q)]·NO3·2H2O} (1) and {[Cu(Hqmba)2(q)]·NO3·2H2O} (2) (where, qmbn = 2-(quinolin-8-yloxy)(methyl) benzonitrile (L1); Hqmba = 2-((quinolin-8-yloxy)methyl)benzoic acid (L2) and q = quinolin-8-olate). The structural analysis reveals that both the complexes exhibit distorted octahedral (CuN3O3) configuration which is further corroborated by density functional theory (DFT) calculations. The cytotoxicity impact of ligands (L1 and L2) and complexes (1 and 2) was screened against the MCF-7 cell line (human breast cancer). The MTT assay uptake indicated that the presence of -COOH functionality in complex 2 leads to higher cytotoxicity (lower IC50) than that observed for complex 1 containing a -CN group. This could be due to the strong H-bonding forming propensity of the carboxylic acids. Incubation of MCF-7 cancer cells with IC50 concentrations of 1 and 2 promoted cellular detachments via nuclear condensation and membrane destabilization followed by apoptosis as a result of metal-assisted generation of reactive oxygen species. Flow cytometry analysis showed that 1 and 2 might prompt early apoptosis in MCF-7 cells as the maximum percentage of cells appeared in the LR quadrant. Furthermore, mRNA expression analysis confirmed that both the complexes induced apoptosis in MCF-7 cells. Comparative mRNA expression analysis of complexes with their respective ligands also confirmed the enhanced apoptotic behavior of complexes. Furthermore, molecular docking studies of the complexes have also been performed with the active site of EGFR kinase receptors (major target for any cancer causing agent) due to similar analogues with FDA-approved EGFR inhibitors in order to rationalize its promising cytotoxicity activity.

An Assessment on Impact of COVID-19 Infection in a Gender Specific Manner.

Coronavirus disease 2019 (COVID-19) is caused by novel coronavirus Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). It was first time reported in December 2019 in Wuhan, China and thereafter quickly spread across the globe. Till September 19, 2020, COVID-19 has spread to 216 countries and territories. Severe infection of SARS-CoV-2 cause extreme increase in inflammatory chemokines and cytokines that may lead to multi-organ damage and respiratory failure. Currently, no specific treatment and authorized vaccines are available for its treatment. Renin angiotensin system holds a promising role in human physiological system specifically in regulation of blood pressure and electrolyte and fluid balance. SARS-CoV-2 interacts with Renin angiotensin system by utilizing angiotensin-converting enzyme 2 (ACE2) as a receptor for its cellular entry. This interaction hampers the protective action of ACE2 in the cells and causes injuries to organs due to persistent angiotensin II (Ang-II) level. Patients with certain comorbidities like hypertension, diabetes, and cardiovascular disease are under the high risk of COVID-19 infection and mortality. Moreover, evidence obtained from several reports also suggests higher susceptibility of male patients for COVID-19 mortality and other acute viral infections compared to females. Analysis of severe acute respiratory syndrome coronavirus (SARS) and Middle East respiratory syndrome coronavirus (MERS) epidemiological data also indicate a gender-based preference in disease consequences. The current review addresses the possible mechanisms responsible for higher COVID-19 mortality among male patients. The major underlying aspects that was looked into includes smoking, genetic factors, and the impact of reproductive hormones on immune systems and inflammatory responses. Detailed investigations of this gender disparity could provide insight into the development of patient tailored therapeutic approach which would be helpful in improving the poor outcomes of COVID-19. Graphical abstract.

Targeted bioimaging and sensing of folate receptor-positive cancer cells using folic acid-conjugated sulfur-doped graphene quantum dots.

For the first time is reported a facile in situ synthesis of folic acid-conjugated sulfur-doped graphene quantum dots (FA-SGQDs) through simple pyrolysis of citric acid (CA), 3-mercaptopropionic acid (MPA), and FA. The as-prepared FA-SGQDs were extensively characterized to confirm the synthesis and incidence of FA molecule on the surface of SGQDs through advanced characterization techniques. Upon excitation at 370-nm wavelength, FA-SGQDs exhibited blue fluorescence with an emission band at 455 nm. While exhibiting relatively high quantum yield (~ 78%), favorable biocompatibility, excellent photostability, and desirable optical properties, the FA-SGQDs showed suitability as a fluorescent nanoprobe to distinguish the folate receptor (FR)-positive and FR-negative cancer cells. The experimental studies revealed that FA-SGQDs aptly entered into FR-positive cancer cells via a non-immunogenic FR-mediated endocytosis process. Additionally, the FA-SGQDs exhibited excellent free radical scavenging activity. Hence, these FA-SGQDs hold high promise to serve as efficient fluorescent nanoprobes for the pre-diagnosis of cancer through targeted bioimaging and other pertinent biological studies. Graphical abstract.