NFSI mediated C3-ether oxidation of glycals for the synthesis of hex-3-enuloses.

Hex-3-enuloses constitute a vital carbohydrate synthetic intermediate that provide access to wide range of chiral molecules through diverse derivatizations. Herein we report synthesis of these fascinating scaffolds by oxidation of C3-ether protections on glycals in presence of N-fluorobenzenesulfonimide (NFSI) under Cu(I) catalysed conditions. Benzyl, methyl and silyl ethers have been efficiently oxidized to the carbonyl group. The oxidation has been found to be highly regioselective where an array of protecting groups were tolerant to the reaction conditions. Pyranosyl glycals from various commercially available sugars have been studied in this work to evaluate the broad substrate scope.

Super-resolution imaging of antibody-conjugated biodegradable periodic mesoporous organosilica nanoparticles for targeted chemotherapy of prostate cancer.

Biodegradable periodic mesoporous organosilica nanoparticles (nanoPMOs) are widely used as responsive drug delivery platforms for targeted chemotherapy of cancer. However, the evaluation of their properties such as surface functionality and biodegradability is still challenging, which has a significant impact on the efficiency of chemotherapy. In this study, we have applied direct stochastic optical reconstruction microscopy (dSTORM), a single-molecule super-resolution microscopy technique, to quantify the degradation of nanoPMOs triggered by glutathione and the multivalency of antibody-conjugated nanoPMOs. Subsequently, the effect of these properties on cancer cell targeting, drug loading and release capability, and anticancer activity is also studied. Due to the higher spatial resolution at the nanoscale, dSTORM imaging is able to reveal the structural properties (i.e., size and shape) of fluorescent and biodegradable nanoPMOs. The quantification of nanoPMOs' biodegradation using dSTORM imaging demonstrates their excellent structure-dependent degradation behavior at a higher glutathione concentration. The surface functionality of anti-M6PR antibody-conjugated nanoPMOs as quantified by dSTORM imaging plays a key role in prostate cancer cell labeling: oriented antibody is more effective than random ones, while high multivalency is also effective. The higher biodegradability and cancer cell-targeting properties of nanorods conjugated with oriented antibody (EAB4H) effectively deliver the anticancer drug doxorubicin to cancer cells, exhibiting potent anticancer effects.

Effect of dietary oregano essential oil and milk replacer on physiological status and immunological responses of pre- and post-weaned Ghoongroo piglets.

Ghoongroo pigs have good adaptability in a low input production system with high prolificacy. The present study was conducted on pre-and post-weaned Ghoongroo piglets from 2-3 days to 12 weeks of age to evaluate the effect of a milk replacer and oregano essential oil (EO) on growth, physiological and immunological responses. Thirty six piglets were randomly divided into three groups. The control group (n = 12) was allowed to suck mother's milk. Second group piglets were provided milk replacer (MR) and piglets of the third group were provided milk replacer along with oregano EO at 500 mg/kg diet. After weaning, piglets were provided standard concentrate diets. The results showed that the body weight in MR and MR + EO groups were significantly higher compared with the control. The MR + EO group had better intestinal microbiota, greater nonspecific innate immunity with the phagocytosis efficacy of neutrophils, lower cortisol concentration and more stable thyroid hormones than the other groups. The better haematological status supported the rapid organ development and improved intestinal health status in both the experimental groups. In conclusion, milk replacer, especially with the inclusion of oregano EO, can lower weaning stress, enhance nonspecific immunity and improve growth and health status of piglets.

Magnetic Nanostructures as Emerging Therapeutic Tools to Boost Anti-Tumour Immunity.

Cancer immunotherapy has shown remarkable results in various cancer types through a range of immunotherapeutic approaches, including chimeric antigen receptor-T cell (CAR-T) therapy, immune checkpoint blockade (ICB), and therapeutic vaccines. Despite the enormous potential of cancer immunotherapy, its application in various clinical settings has been limited by immune evasion and immune suppressive mechanisms occurring locally or systemically, low durable response rates, and severe side effects. In the last decades, the rapid advancement of nanotechnology has been aiming at the development of novel synthetic nanocarriers enabling precise and enhanced delivery of immunotherapeutics, while improving drug stability and effectiveness. Magnetic nanostructured formulations are particularly intriguing because of their easy surface functionalization, low cost, and robust manufacturing procedures, together with their suitability for the implementation of magnetically-guided and heat-based therapeutic strategies. Here, we summarize and discuss the unique features of magnetic-based nanostructures, which can be opportunely designed to potentiate classic immunotherapies, such as therapeutic vaccines, ICB, adoptive cell therapy (ACT), and in situ vaccination. Finally, we focus on how multifunctional magnetic delivery systems can facilitate the anti-tumour therapies relying on multiple immunotherapies and/or other therapeutic modalities. Combinatorial magnetic-based therapies are indeed offering the possibility to overcome current challenges in cancer immunotherapy.

A PKCß-LYN-PYK2 Signaling Axis Is Critical for MCP-1-Dependent Migration and Adhesion of Monocytes.

MCP-1-induced monocyte chemotaxis is a crucial event in inflammation and atherogenesis. Identifying the important signal transduction pathways that control monocyte chemotaxis can unravel potential targets for preventive therapies in inflammatory disease conditions. Previous studies have shown that the focal adhesion kinase Pyk2 plays a critical role in monocyte motility. In this study, we investigated the MCP-1-mediated activation of Pyk2 (particularly by the phosphorylation of Tyr402) in primary human peripheral blood monocytes. We showed that MCP-1 induces Src phosphorylation in a similar time frame and that the MCP-1-induced Pyk2 tyrosine phosphorylation is controlled by the Src family kinase. We also report, in this study, that PKCß, an isoform of PKC, is required for both Src and Pyk2 activation/phosphorylation in response to MCP-1 stimulation. We identified Lyn as the specific Src kinase isoform that is activated by MCP-1 and acts upstream of Pyk2 in primary monocytes. Furthermore, Lyn is found to be indispensable for monocyte migration in response to MCP-1 stimulation. Moreover, our coimmunoprecipitation studies in monocytes revealed that PKCß, Pyk2, and Lyn exist constitutively in a molecular complex. To our knowledge, our study has uncovered a novel PKCß-Lyn-Pyk2 signaling cascade in primary monocytes that regulates MCP-1-induced monocyte adhesion and migration.

Cyanomethyl Ether as an Orthogonal Participating Group for Stereoselective Synthesis of 1,2-trans-ß-O-Glycosides.

Stereoselective formation of glycosidic linkages has been the prime focus for contemporary carbohydrate chemistry. Herein, we report cyanomethyl (CNMe) ether as an efficient and effective participating orthogonal protecting group for the stereoselective synthesis of 1,2-trans-ß-O-glycosides. The participating group facilitated good to high ß-selective glycosylation with a broad range of electron-rich and electron-deficient glycosyl acceptors. Detailed experimental and theoretical studies reveal the involvement of CNMe ether in the formation of a six-membered imine-type cyclic intermediate for the observed stereoselectivity. Rapid incorporation and selective removal of the CNMe ether group in the presence of benzyl ether and isopropylidene acetal protection have also been reported here. The nitrile group provided an opportunity for the glycodiversification through further derivatizations.

Colloidal polymer-coated Zn-doped iron oxide nanoparticles with high relaxivity and specific absorption rate for efficient magnetic resonance imaging and magnetic hyperthermia.

Colloidally stable nanoparticles-based magnetic agents endowed with very high relaxivity and specific absorption rate are extremely desirable for efficient magnetic resonance imaging and magnetic hyperthermia, respectively. Here, we report a water dispersible magnetic agent consisting of zinc-doped superparamagnetic iron oxide nanoparticles (i.e., Zn-SPIONs) of 15 nm size with high saturation magnetization coated with an amphiphilic polymer for effective magnetic resonance imaging and magnetic hyperthermia of glioblastoma cells. These biocompatible polymer-coated Zn-SPIONs had 24 nm hydrodynamic diameter and exhibited high colloidal stability in various aqueous media, very high transverse relaxivity of 471 mM-1 s-1, and specific absorption rate up to 743.8 W g-1, which perform better than most iron oxide nanoparticles reported in the literature, including commercially available agents. Therefore, using these polymer-coated Zn-SPIONs even at low concentrations, T2-weighted magnetic resonance imaging and moderate magnetic hyperthermia of glioblastoma cells under clinically relevant magnetic field were successfully implemented. In addition, the results of this in vitro study suggest the superior potential of Zn-SPIONs as a theranostic nanosystem for brain cancer treatment, simultaneously acting as a contrast agent for magnetic resonance imaging and a heat mediator for localized magnetic hyperthermia.

Multifunctional Magnetic Gold Nanomaterials for Cancer.

The integration of multiple imaging and therapeutic agents into a customizable nanoplatform for accurate identification and rapid prevention of cancer is attracting great attention. Among the available theranostic nanosystems, magnetic gold nanoparticles are particularly promising as they exhibit unique physicochemical properties that can support multiple functions, including cancer diagnosis by magnetic resonance imaging, X-ray computed tomography, Raman and photoacoustic imaging, drug delivery, and plasmonic photothermal and photodynamic therapies. This review gives an overview of recent advances in the fabrication of multifunctional gold nanohybrids with magnetic and optical properties and their successful demonstration in multimodal imaging and therapy of cancer. Concerns around toxicity of these nanomaterials are also discussed in view of an imminent transition to clinical practice.

Recent advances in magnetic fluid hyperthermia for cancer therapy.

Recently, magnetic fluid hyperthermia using biocompatible magnetic nanoparticles as heat mediators for cancer therapy has been extensively investigated due to its high efficiency and limited side effects. However, the development of more efficient heat nanomediators that exhibit very high specific absorption rate (SAR) value is essential for clinical application to overcome the several restrictions previously encountered due to the large quantity of nanomaterial required for effective treatment. In this review, we focus on the current progress in the development of magnetic nanoparticles based hyperthermia therapy as well as combined therapy harnessing hyperthermia with heat-mediated drug delivery for cancer treatment. We also address the fundamental principles of magnetic hyperthermia, basics of magnetism including the effect of several parameters on heating capacity, synthetic methods and nanoparticle surface chemistry needed to design and develop an ideal magnetic nanoparticle heat mediator suitable for clinical translation in cancer therapy.

Novel decellularized animal conchal cartilage graft for application in human patient.

Restoration of the external ear and nose in human patients, in either congenital deformity or acquired defects, is a challenge in reconstructive surgery. Optimization of the currently available materials is necessary for rhinoplasty and microtia correction to avoid intraoperative manoeuvring and early rejection. The aim of this study was to develop cross-linked decellularized caprine conchal cartilages as biocompatible, robust, and non-toxic matrix template. The characterization of the decellularized tissue encompasses in vitro lymphoproliferation assay, cytotoxicity test, agar gel precipitation test, in vivo immunocompatibility study, histology, and determination of pro-inflammatory cytokines in animal model. Decellularized cartilage was implanted in human volunteer at R. G. Kar Medical College and Hospital, Kolkata, India, and samples were assessed histologically by retrieving those after 4 months. The processed cartilages were implanted in rhinoplasty (nine) and microtia patients (six) keeping autogenous cartilage graft as control up to 18 months after surgery. Primary outcomes were viability and safety of the material, both in animal model and human pre-application in actual site. Secondary outcomes included self-assessed clinical findings on gross examination. This study is under the ethical approval no. RKC/14 dated January 27, 2012. The in vitro cellular reactivity was less in processed cartilage protein than control. Histology of retrieved tissues in animal model and human volunteer showed no adverse reactions. Production of IL-2, IL-6, and TNF-α cytokines was lower at 4 weeks. The rhinoplasty and microtia operation in clinical patients utilizing the processed cartilage showed satisfactory recovery with improved facial look. These low cost, easily available, biocompatible, safe xenocartilage biomatrices of caprine conchal cartilage origin are very flexible in shape and size, enabling them as potential bioimplant for repair of nasal and auricular structure without any rejection or diverse biomedical applications.

Cancer biomarker determination by resonance energy transfer using functional fluorescent nanoprobes.

The development of bioanalytical methods that provide early detection of the presence of cancer by sensitive and specific determination of biomarkers such as small biomolecules, nucleic acids, proteins, enzymes, and even whole cells are essential to improve opportunity for improved patient treatment and to diminish the rate of cancer mortality. Förster resonance energy transfer (FRET) methods have been increasingly used to develop bioassays that offer speed, selectivity and low detection levels with practicality that is appropriate for providing point-of-care measurements for screening. The unique optical and photophysical properties of fluorescent nanoparticles such as semiconductor quantum dots (QDs), upconversion nanoparticles (UCNPs), graphene quantum dots (GQDs) and other materials have been reported to operate as efficient donors and/or acceptors for replacement of fluorescent organic dye molecules in various FRET-based assays. This review is focused on the recent progress that has been made in the development of nanoparticle-based FRET bioassays, and considers nanoparticle synthesis, design of optical properties, conjugation chemistry and approaches to fluorescence detection that provide for selective and sensitive quantification of cancer biomarkers.

Regulation of monoamine oxidase A (MAO-A) expression, activity, and function in IL-13-stimulated monocytes and A549 lung carcinoma cells.

Monoamine oxidase A (MAO-A) is a mitochondrial flavoenzyme implicated in the pathogenesis of atherosclerosis and inflammation and also in many neurological disorders. MAO-A also has been reported as a potential therapeutic target in prostate cancer. However, the regulatory mechanisms controlling cytokine-induced MAO-A expression in immune or cancer cells remain to be identified. Here, we show that MAO-A expression is co-induced with 15-lipoxygenase (15-LO) in interleukin 13 (IL-13)-activated primary human monocytes and A549 non-small cell lung carcinoma cells. We present evidence that MAO-A gene expression and activity are regulated by signal transducer and activator of transcription 1, 3, and 6 (STAT1, STAT3, and STAT6), early growth response 1 (EGR1), and cAMP-responsive element-binding protein (CREB), the same transcription factors that control IL-13-dependent 15-LO expression. We further established that in both primary monocytes and in A549 cells, IL-13-stimulated MAO-A expression, activity, and function are directly governed by 15-LO. In contrast, IL-13-driven expression and activity of MAO-A was 15-LO-independent in U937 promonocytic cells. Furthermore, we demonstrate that the 15-LO-dependent transcriptional regulation of MAO-A in response to IL-13 stimulation in monocytes and in A549 cells is mediated by peroxisome proliferator-activated receptor γ (PPARγ) and that signal transducer and activator of transcription 6 (STAT6) plays a crucial role in facilitating the transcriptional activity of PPARγ. We further report that the IL-13-STAT6-15-LO-PPARγ axis is critical for MAO-A expression, activity, and function, including migration and reactive oxygen species generation. Altogether, these results have major implications for the resolution of inflammation and indicate that MAO-A may promote metastatic potential in lung cancer cells.

Deregulation of LIMD1-VHL-HIF-1α-VEGF pathway is associated with different stages of cervical cancer.

To understand the mechanism of cellular stress in basal-parabasal layers of normal cervical epithelium and during different stages of cervical carcinoma, we analyzed the alterations (expression/methylation/copy number variation/mutation) of HIF-1α and its associated genes LIMD1, VHL and VEGF in disease-free normal cervix (n = 9), adjacent normal cervix of tumors (n = 70), cervical intraepithelial neoplasia (CIN; n = 32), cancer of uterine cervix (CACX; n = 174) samples and two CACX cell lines. In basal-parabasal layers of normal cervical epithelium, LIMD1 showed high protein expression, while low protein expression of VHL was concordant with high expression of HIF-1α and VEGF irrespective of HPV-16 (human papillomavirus 16) infection. This was in concordance with the low promoter methylation of LIMD1 and high in VHL in the basal-parabasal layers of normal cervix. LIMD1 expression was significantly reduced while VHL expression was unchanged during different stages of cervical carcinoma. This was in concordance with their frequent methylation during different stages of this tumor. In different stages of cervical carcinoma, the expression pattern of HIF-1α and VEGF was high as seen in basal-parabasal layers and inversely correlated with the expression of LIMD1 and VHL. This was validated by demethylation experiments using 5-aza-2'-deoxycytidine in CACX cell lines. Additional deletion of LIMD1 and VHL in CIN/CACX provided an additional growth advantage during cervical carcinogenesis through reduced expression of genes and associated with poor prognosis of patients. Our data showed that overexpression of HIF-1α and its target gene VEGF in the basal-parabasal layers of normal cervix was due to frequent inactivation of VHL by its promoter methylation. This profile was maintained during different stages of cervical carcinoma with additional methylation/deletion of VHL and LIMD1.

Activation of Wnt-ß-catenin pathway in basal-parabasal layers of normal cervical epithelium comparable during development of uterine cervical carcinoma.

In this study, importance of Wnt-ß-catenin pathway in the development of uterine cervical carcinoma was evaluated. For this purpose, the profiles (expression/methylation/deletion) of ß-catenin, p-ß-catenin (Y654), Wnt3a, and APC were studied in disease free normal cervical epithelium (n = 9), adjacent normal cervical epithelium of primary tumors (n = 70), CIN (n = 28), CACX (n = 102) samples, and two CACX cell lines (HeLa and SiHa). Immunohistochemical analysis revealed high/medium (74-95%) expression of ß-catenin/p-ß-catenin (Y654) and Wnt3a and low expression (23-26%) of APC in proliferating basal-parabasal layers contrary to differentiated spinous layer in normal cervix irrespective of HPV16 infection. The expression profile of the genes in the basal-parabasal layers did not change significantly during development of CACX. High (66%) promoter methylation of APC was seen in basal-parabasal layers and the cervical lesions (42-69%), unlike in spinous layers (25%). The promoter methylation status of APC was validated by in vitro demethylation experiments using 5-aza-dC in CACX cell lines. However, additional deletion of APC was significantly increased from CIN (12%) to stage I/II (40%) and became comparable in stage III/IV (48%) of the tumor. Patients with alterations (deletion/methylation) of APC and high/medium expression of Wnt3a/ß-catenin/p-ß-catenin (Y654) showed significantly poor survival. Thus our data indicate that cumulative effect of Wnt3a overexpression and APC inactivation are needed for overexpression of ß-catenin during the development of CACX.

Detection of a cancer biomarker protein on modified cellulose paper by fluorescence using aptamer-linked quantum dots.

The development of point-of-care bioassays for sensitive screening of protein-based cancer biomarkers would improve the opportunity for early stage diagnosis. A strategy for a fluorescence resonance energy transfer (FRET)-based bioassay has been investigated that makes use of modified cellulose paper for the detection of an epithelial cell adhesion molecule (EpCAM), which is a transmembrane glycoprotein that is overexpressed in several tumors of epithelial origin. The paper matrix was a substrate for immobilized aptamer-linked quantum dots (QDs-Apt) and Cy3 labeled complementary DNA (cDNA), which served as a donor and an acceptor, respectively. Competitive binding of EpCAM displaced the cDNA, resulting in the reduction of FRET. The paper-based bioassay was able to detect EpCAM in buffer solution as well as in 10% bovine serum solution using a reaction time of no more than 60 minutes. The dynamic range was 1-100 nM in buffer with a precision better than 4%, and the limit of detection was 250 pM in buffer and 600 pM in 10% serum.

Risk of high-grade precancerous lesions and invasive cancers in high-risk HPV-positive women with normal cervix or CIN 1 at baseline-A population-based cohort study.

Infection with high-risk human papillomavirus (HR-HPV) is transient and clears on its own in majority of the women. Only a few women who have persistent infection may finally develop cervical intraepithelial neoplasia (CIN) or cervical cancer in later years. The risk of progression in the HR-HPV-positive women with normal cervix or low-grade lesion on colposcopy and histopathology at baseline is less studied. We performed a longitudinal study on 650 HR-HPV-positive women with colposcopy and/or histopathology-proved normal or CIN1 diagnosis at baseline to assess the cumulative risk of development of high-grade CIN. After a mean follow-up of 2.1 person years of observation (PYO) (range 0.1-5.1), the cumulative incidence of CIN2+ (6.4%; 3.0/100 PYO) was significantly higher in women who had persistent HR-HPV infection compared to those who cleared the infection (adjusted HR 6.28; 95% CI 2.87-13.73). The risk of viral persistence in women aged 50-60 years was two times higher compared to women aged 40-49 years and three times higher compared to women aged 30-39 years. The probability of having persistent infection increased progressively with higher viral load at baseline (adjusted HR 3.29, 95% CI 2.21-4.90 for RLU ≥100; adjusted HR 2.69, 95% CI 1.71-4.22 for RLU 10-100). Women with increasing viral load at follow-up had four times higher risk of developing CIN2 or worse lesions as compared to those with decreasing load (20.9% vs 4.8%; p < 0.001). In the context of developing countries where cytology or genotyping triaging is not feasible, colposcopy referral of HR-HPV-positive women with advancing age, viral persistence, and increasing viral load may be considered.

Evaluation of a compact, rechargeable, magnifying device to triage VIA and HPV positive women in a cervical cancer screening program in rural India.

PURPOSE: Many limited-resourced countries have either introduced cervical cancer screening programs or are contemplating to do so using visual inspection after acetic acid application (VIA) or human papillomavirus (HPV) detection tests. Both tests have high false-positivity and a suitable triaging strategy is required. Colposcopy triaging is not practicable in most resource-limited settings due to several reasons. We evaluated a portable, battery-operated, magnifying device (GynocularTM) to triage screen positive women in community setting in India. METHODS: Women positive on VIA or oncogenic HPV test were examined with Gynocular by clinicians in primary health clinics. Findings were documented using the International Federation for Cervical Pathology and Colposcopy (IFCPC) terminology. Swede score was also calculated. Biopsy was performed irrespective of Gynocular findings. The accuracy of Gynocular to detect high-grade lesions or cancer (HSIL+) was estimated. The suitability of Gynocular to correctly triage screen positive cases for immediate ablative treatment was also evaluated by creating simulated scenarios. RESULTS: Sensitivity and specificity of Gynocular were 96.4 and 47.1 %, respectively, to detect HSIL + at the threshold of IFCPC grade 1 findings. Increasing threshold to grade 2 changed sensitivity and specificity to 92.9 and 94.1 %, respectively. Optimum combination of sensitivity and specificity as determined by the receiver operating curve analysis was at the cut-off Swede score of 5. Triaging of VIA/HPV positive women to treatment using grade 2 criteria would have resulted in modest overtreatment and missing of very few high-grade lesions. CONCLUSION: Gynocular can be used as an effective triaging device for VIA/HPV positive women.

Valence tautomerism in synthetic models of cytochrome P450.

CytP450s have a cysteine-bound heme cofactor that, in its as-isolated resting (oxidized) form, can be conclusively described as a ferric thiolate species. Unlike the native enzyme, most synthetic thiolate-bound ferric porphyrins are unstable in air unless the axial thiolate ligand is sterically protected. Spectroscopic investigations on a series of synthetic mimics of cytP450 indicate that a thiolate-bound ferric porphyrin coexists in organic solutions at room temperature (RT) with a thiyl-radical bound ferrous porphyrin, i.e., its valence tautomer. The ferric thiolate state is favored by greater enthalpy and is air stable. The ferrous thiyl state is favored by entropy, populates at RT, and degrades in air. These ground states can be reversibly interchanged at RT by the addition or removal of water to the apolar medium. It is concluded that hydrogen bonding and local electrostatics protect the resting oxidized cytP450 active site from degradation in air by stabilizing the ferric thiolate ground state in contrast to its synthetic analogs.

HPV detection-based cervical cancer screening program in low-resource setting: lessons learnt from a community-based demonstration project in India.

PURPOSE: A demonstration project was conducted to assess feasibility of implementing HPV detection-based cervical cancer screening in primary care settings in India and to generate local evidence on feasibility and effectiveness of HPV detection in primary screening. METHODS: The project was implemented by setting up screening clinics at primary health centers. Eligible women were screened by HPV DNA test (Hybrid capture 2). All samples were processed and tested in a single laboratory. Colposcopy services were provided to screen-positive women at the same community clinics. Project utilized services of community health workers for community mobilization, recall of screen-positive and disease-positive women. Women with ≥CIN2 diagnosis were treated at tertiary hospital. RESULTS: Totally, 44,110 women were screened and HPV positivity was 4.7 %. Compliance to recall of HC2-positive women for colposcopy was 78 %. Detection rate of CIN3+ by HPV test was 3.9/1,000 women. Compliance of women to treatment was 80.1 %. However, compliance of HPV-positive women for follow-up at 1 year was poor (23.2 %). Concurrent use of VIA to screen the women did not have any advantage but increased number of unnecessary colposcopies and biopsies. CONCLUSIONS: Our project demonstrated that it was possible to implement HPV detection-based screening using existing primary healthcare infrastructure. Performing colposcopy at primary setting is feasible, improves compliance and reduces over-treatment. In settings with low to moderately high HPV prevalence, direct referral of HPV-positive women is advisable. Community health workers can be effectively used for recalling the positive women.