Transcriptome profiling and metabolic pathway analysis towards reliable biomarker discovery in early-stage lung cancer.

Earlier diagnosis of lung cancer is crucial for reducing mortality and morbidity in high-risk patients. Liquid biopsy is a critical technique for detecting the cancer earlier and tracking the treatment outcomes. However, noninvasive biomarkers are desperately needed due to the lack of therapeutic sensitivity and early-stage diagnosis. Therefore, we have utilized transcriptomic profiling of early-stage lung cancer patients to discover promising biomarkers and their associated metabolic functions. Initially, PCA highlights the diversity level of gene expression in three stages of lung cancer samples. We have identified two major clusters consisting of highly variant genes among the three stages. Further, a total of 7742, 6611, and 643 genes were identified as DGE for stages I-III respectively. Topological analysis of the protein-protein interaction network resulted in seven candidate biomarkers such as JUN, LYN, PTK2, UBC, HSP90AA1, TP53, and UBB cumulatively for the three stages of lung cancers. Gene enrichment and KEGG pathway analyses aid in the comprehension of pathway mechanisms and regulation of identified hub genes in lung cancer. Importantly, the medial survival rates up to ~ 70 months were identified for hub genes during the Kaplan-Meier survival analysis. Moreover, the hub genes displayed the significance of risk factors during gene expression analysis using TIMER2.0 analysis. Therefore, we have reason that these biomarkers may serve as a prospective targeting candidate with higher treatment efficacy in early-stage lung cancer patients.

Machine learning driven drug repurposing strategy for identification of potential RET inhibitors against non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) remains the leading cause of mortality and morbidity worldwide accounting about 85% of total lung cancer cases. The receptor REarranged during Transfection (RET) plays an important role by ligand independent activation of kinase domain resulting in carcinogenesis. Presently, the treatment for RET driven NSCLC is limited to multiple kinase inhibitors. This situation necessitates the discovery of novel and potent RET specific inhibitors. Thus, we employed high throughput screening strategy to repurpose FDA approved compounds from DrugBank comprising of 2509 molecules. It is worth noting that the initial screening is accomplished with the aid of in-house machine learning model built using IC50 values corresponding to 2854 compounds obtained from BindingDB repository. A total of 497 compounds (19%) were predicted as actives by our generated model. Subsequent in silico validation process such as molecular docking, MMGBSA and density function theory analysis resulted in identification of two lead compounds named DB09313 and DB00471. The simulation study highlights the potency of DB00471 (Montelukast) as potential RET inhibitor among the investigated compounds. In the end, the half-minimal inhibitory activity of montelukast was also predicted against RET protein expressing LC-2/ad cell lines demonstrated significant anticancer activity. Collective analysis from our study highlights that montelukast could be a promising candidate for the management of RET specific NSCLC.

Designing Novel Compounds for the Treatment and Management of RET-Positive Non-Small Cell Lung Cancer-Fragment Based Drug Design Strategy.

Rearranged during transfection (RET) is an oncogenic driver receptor that is overexpressed in several cancer types, including non-small cell lung cancer. To date, only multiple kinase inhibitors are widely used to treat RET-positive cancer patients. These inhibitors exhibit high toxicity, less efficacy, and specificity against RET. The development of drug-resistant mutations in RET protein further deteriorates this situation. Hence, in the present study, we aimed to design novel drug-like compounds using a fragment-based drug designing strategy to overcome these issues. About 18 known inhibitors from diverse chemical classes were fragmented and bred to form novel compounds against RET proteins. The inhibitory activity of the resultant 115 hybrid molecules was evaluated using molecular docking and RF-Score analysis. The binding free energy and chemical reactivity of the compounds were computed using MM-GBSA and density functional theory analysis, respectively. The results from our study revealed that the developed hybrid molecules except for LF21 and LF27 showed higher reactivity and stability than Pralsetinib. Ultimately, the process resulted in three hybrid molecules namely LF1, LF2, and LF88 having potent inhibitory activity against RET proteins. The scrutinized molecules were then subjected to molecular dynamics simulation for 200 ns and MM-PBSA analysis to eliminate a false positive design. The results from our analysis hypothesized that the designed compounds exhibited significant inhibitory activity against multiple RET variants. Thus, these could be considered as potential leads for further experimental studies.

Manual loop in laparoscopic appendectomy: A retrospective cohort study and literature review.

OBJECTIVES: To determine the incidence of complications [Surgical site infection (SSI), intra-abdominal abscess (IAA), stump leak] related to stump ligation with manual loop of sliding extracorporeal suture knot in laparoscopic appendectomy. METHODS: This cohort study was conducted on patients who underwent laparoscopic appendectomy from June 2014 to November 2020 performed by the same surgeon with almost similar technique. Stump was ligated with manual loops, applied by the surgeon or trainee or both (one by surgeon and other by trainee). SSI and IAA were classified according to Centers for Disease Control and Prevention (CDC) criteria. RESULTS: Total 120 patients were included with median (Interquartile range, IQR) age of 24 (19-35) years and male predominance i.e. 81 (67.5%). Median (IQR) for the duration of symptoms, time from presentation to surgery and duration of surgery was 2(1-4) days, 10 (4-15) hours and 60 (44-70) minutes, respectively. SSI was documented in 9(7.5%) patients, managed by wound hygiene and antibiotics. IAA was observed in one(0.8%) patient who required readmission for antibiotics and radiology guided drain placement. No stump leak was observed. CONCLUSIONS: Manual endo-loop is a safe, reliable and cost effective technique for stump ligation in LA, and can safely be incorporated into teaching of surgical trainees.

Trabeculectomy Augmented with Limited Deep Sclerectomy and Cyclodialysis with Use of Scleral Tissue as a Spacer.

Trabeculectomy remains the most commonly performed surgery for medically uncontrolled glaucoma. Its success in primary open angle glaucoma is approximately 82% in the initial year after surgery and 64% at the end of five years. Lower success rates have been found in secondary glaucomas like neovascular glucoma, uvietic glaucoma, post-traumatic glaucoma, and for repeat surgeries. To illustrate improvement of the efficacy of trabeculectomy, enhancement with cyclodialysis has been introduced. This involves the creation of a cyclodialysis cleft in a controlled manner to allow additional suprachoroidal drainage of the aqueous. Cyclodialysis is the result of the separation of the longitudinal ciliary muscle fibers from the scleral spur, which creates an additional pathway for aqueous humor drainage. However, such a cleft often closes on its own due to associated inflammation caused by the filtration surgery. Deep sclerectomy is a non-penetrating surgery that involves dissection of a scleral patch and excision of a block of scleral tissue, retaining a thin membrane for aqueous drainage. In this study, we introduce a novel surgical technique of combining trabeculectomy with a limited deep sclerectomy and a cyclodialysis in two pseudophakic patients who developed secondary glaucoma after vitreo-retinal surgery with silicone oil insertion. In this technique the excised scleral tissue obtained after deep sclerectomy was utilized as a spacer to maintain the patency of the cyclodialysis cleft.

Flavonoids: Classification, Function, and Molecular Mechanisms Involved in Bone Remodelling.

Flavonoids are polyphenolic compounds spotted in various fruits, vegetables, barks, tea plants, and stems and many more natural commodities. They have a multitude of applications through their anti-inflammatory, anti-oxidative, anti-carcinogenic properties, along with the ability to assist in the stimulation of bone formation. Bone, a rigid connective body tissue made up of cells embedded in a mineralised matrix is maintained by an assemblage of pathways assisting osteoblastogenesis and osteoclastogenesis. These have a significant impact on a plethora of bone diseases. The homeostasis between osteoblast and osteoclast formation decides the integrity and structure of the bone. The flavonoids discussed here are quercetin, kaempferol, icariin, myricetin, naringin, daidzein, luteolin, genistein, hesperidin, apigenin and several other flavonoids. The effects these flavonoids have on the mitogen activated protein kinase (MAPK), nuclear factor kappa ß (NF-kß), Wnt/ß-catenin and bone morphogenetic protein 2/SMAD (BMP2/SMAD) signalling pathways, and apoptotic pathways lead to impacts on bone remodelling. In addition, these polyphenols regulate angiogenesis, decrease the levels of inflammatory cytokines and play a crucial role in scavenging reactive oxygen species (ROS). Considering these important effects of flavonoids, they may be regarded as a promising agent in treating bone-related ailments in the future.

Discovery of a Potent Candidate for RET-Specific Non-Small-Cell Lung Cancer-A Combined In Silico and In Vitro Strategy.

Rearranged during transfection (RET) is a tyrosine kinase oncogenic receptor, activated in several cancers including non-small-cell lung cancer (NSCLC). Multiple kinase inhibitors vandetanib and cabozantinib are commonly used in the treatment of RET-positive NSCLC. However, specificity, toxicity, and reduced efficacy limit the usage of multiple kinase inhibitors in targeting RET protein. Thus, in the present investigation, we aimed to figure out novel and potent candidates for the inhibition of RET protein using combined in silico and in vitro strategies. In the present study, screening of 11,808 compounds from the DrugBank repository was accomplished by different hypotheses such as pharmacophore, e-pharmacophore, and receptor cavity-based models in the initial stage. The results from the different hypotheses were then integrated to eliminate the false positive prediction. The inhibitory activities of the screened compounds were tested by the glide docking algorithm. Moreover, RF score, Tanimoto coefficient, prime-MM/GBSA, and density functional theory calculations were utilized to re-score the binding free energy of the docked complexes with high precision. This procedure resulted in three lead molecules, namely DB07194, DB03496, and DB11982, against the RET protein. The screened lead molecules together with reference compounds were then subjected to a long molecular dynamics simulation with a 200 ns time duration to validate the inhibitory activity. Further analysis of compounds using MM-PBSA and mutation studies resulted in the identification of potent compound DB07194. In essence, a cell viability assay with RET-specific lung cancer cell line LC-2/ad was also carried out to confirm the in vitro biological activity of the resultant compound, DB07194. Indeed, the results from our study conclude that DB07194 can be effectively translated for this new therapeutic purpose, in contrast to the properties for which it was originally designed and synthesized.

Prediction of Micronucleus Assay Outcome Using In Vivo Activity Data and Molecular Structure Features.

In vivo micronucleus assay is the widely used genotoxic test to determine the extent of chromosomal aberrations caused by the chemicals in human beings, which plays a significant role in the drug discovery paradigm. To reduce the uncertainties of the in vivo experiments and the expenses, we intended to develop novel machine learning-based tools to predict the toxicity of the compounds with high precision. A total of 372 compounds with known toxicity information were retrieved from the PubChem Bioassay database and literature. The fingerprints and descriptors of the compounds were generated using PaDEL and ChemSAR, respectively, for the analysis. The performance of the models was assessed using the three tires of evaluation strategies such as fivefold, tenfold, and validation by external dataset. Further, structural alerts causing genotoxicity of the compounds were identified using SARpy method. Of note, fingerprint-based random forest model built in our analysis is able to demonstrate the highest accuracy of about 0.97 during tenfold cross-validation. In essence, our study highlights that structural alerts such as chlorocyclohexane and trimethylamine are likely to be the leading cause of toxicity in humans. Indeed, we believe that random forest model generated in this study is appropriate for reduction of test animals and should be considered in the future for the good practice of animal welfare.

Need of additional iridotomies despite lens extraction in spherophakes.

Spherophakes are known to have irregular anterior chamber (AC) depths due to their poorly supported zonules. This irregularity leads to an unstable AC, often resulting in angle closure glaucoma from anterior subluxation of globular lenses. A peripheral iridotomy may be helpful to encourage aqueous drainage in initial stages, however, is not often required once lens is extracted. But, we have observed persistent instability of AC in spherophakic eyes despite lens extraction, leading to frequent iridocorneal contact locally over some quadrants. The presumed aetiology in such scenarios could be constant anteroposterior instability of IOL-bag complex due to generalised zonulopathy and hence localised iris bombe in areas with previous iridocorneal contact. Timely identification and performing additional iridotomies during surgery at such sectors even after lens extraction facilitated symmetric deepening of the AC. Hence, we recommend use of additional iridotomies at areas with persistent iridocorneal contact even after lens extraction.

Comparative Evaluation of Rebound and Perkins Tonometers in Pediatric Glaucoma With Varied Corneal Characteristics.

PRECIS: Icare tonometer overestimated intraocular pressure (IOP) as compared with Perkins and this variation was higher in IOP>19 mm Hg and corneal opacity in patients with pediatric glaucoma. PURPOSE: To compare the IOP measured by Icare ic200 with Perkins tonometer in pediatric glaucoma with different corneal characteristics. METHODS: Patients of pediatric glaucoma posted for routine examination under anesthesia, age below 12 years were enrolled. All patients underwent IOP measurement with Perkins and Icare ic200 tonometer by the same observer. Basic demographic data and other relevant clinical data were recorded. Central corneal thickness (CCT), horizontal corneal diameter, and corneal characteristics such as cornea clarity was recorded. RESULTS: A total of 194 eyes of 105 patients were analyzed. The difference between Perkins and Icare IOP was -0.816 mm Hg with the Bland-Altman plot 95% limits of agreement (LoA) from -11.194 to 9.562 mm Hg and 5.1% (10) values lying outside LoA. At IOP <19 mm Hg, the difference was -0.65 mm Hg and IOP ≥19 mm Hg, the difference was higher, -1.12 mm Hg. In the clear cornea group (123 eyes), the difference in IOP by 2 tonometers was -0.776 mm Hg with the Bland-Altman plot 95% LoA between -10.679 and 9.128 mm Hg. In hazy corneas (36 eyes), the difference in IOP was 0.531 mm Hg. The Bland-Altman plot showed 95% LoA between -6.242 and 7.303 mm Hg. In the scarred cornea group (35 eyes), the difference in IOP between the 2 was -2.343 mm Hg and the Bland-Altman plot showed wide 95% LoA from -16.302 to 11.616 mm Hg. CONCLUSION: Icare tonometer overestimated IOP as compared with Perkins and this variation was higher in eyes with IOP≥19 mm Hg, CCT >615 µm, and scarred corneas. A moderate correlation between IOP and CCT for both tonometers was noted.

Gene expression profiling of corona virus microarray datasets to identify crucial targets in COVID-19 patients.

The current outbreak of coronavirus disease (COVID-19) has been affecting millions of people and has caused devastating mortality worldwide. Moreover, it is to be noted that cytokine storm has become an important cause for the rising mortality. However, the efforts for the development of drugs, vaccines and treatment has also been intervened due to poor understanding of host's defense mechanism and also due to the development of cytokine storm against this viral infection. Thus, a deeper understanding of the mechanism behind the immune dysregulation and cytokine storm development might give us clues for the clinical management of the severe cases. Hence, we have implemented differential gene expression analysis together with protein-protein interaction and Gene Ontology (GO) studies with the help of Severe Acute respiratory syndrome coronavirus (SARS-CoV) data sets such as GSE1739 and GSE33267 to give us more knowledge on the host immune response for the pathogenic coronavirus which in turn reduces the mortality. A total of 79 differentially-expressed genes (DEGs) were identified in our data set using the filters such as P-value and log2 fold change values of less than 0.05 and 1.5 respectively. Further, network analysis and GO studies showed that differential expression of two hub genes namely ELANE and LTF which could induce higher levels of pro-inflammatory cytokines in the lungs. We are certain that differential expression of ELANE and LTF results in an excessive inflammatory reaction known as the cytokine storm and ultimately leading to death. Therefore, targeting these key drivers of cytokine storm genes appears to be the potential therapeutic targets for combating the Severe Acute respiratory syndrome coronavirus - 2 (SARS-CoV-2) infection ultimately resulting in reduced mortality. Indeed, this predictive view may open new insights for designing an immune intervention for COVID-19 in the near future resulting in the mitigation of mortality rate.

Attention in visually typical and amblyopic children.

Amblyopia is a cortical visual disorder caused by unequal visual input to the brain from the two eyes during development. Amblyopes show reduced visual acuity and contrast sensitivity and abnormal binocularity, as well as more "global" perceptual losses, such as figure-ground segregation and global form integration. Currently, there is no consensus on the neural basis for these higher-order perceptual losses. One contributing factor could be that amblyopes have deficiencies in attention, such that the attentional processes that control the selection of information favor the better eye. Previous studies in amblyopic adults are conflicting as to whether attentional deficits exist. However, studies where intact attentional ability has been shown to exist were conducted in adults; it is possible that it was acquired through experience. To test this hypothesis, we studied attentional processing in amblyopic children. We examined covert endogenous attention using a classical spatial cueing paradigm in amblyopic and visually typical 5- to 10-year old children. We found that all children, like adults, independently of visual condition, benefited from attentional cueing: They performed significantly better on trials with an informative (valid) cue than with the uninformative (neutral) cue. Response latencies were also significantly shorter for the valid cue condition. No statistically significant difference was found between the performance of the amblyopic and the visually typical children or between dominant and nondominant eyes of all children. The results showed that covert spatial attention is intact in amblyopic and visually typical children and is therefore not likely to account for higher-order perceptual losses in amblyopic children.

A review on recent advancements in diagnosis and classification of cancers using artificial intelligence.

Artificial intelligence has illustrated drastic changes in radiology and medical imaging techniques which in turn led to tremendous changes in screening patterns. In particular, advancements in these techniques led to the development of computer aided detection (CAD) strategy. These approaches provided highly accurate diagnostic reports which served as a "second-opinion" to the radiologists. However, with significant advancements in artificial intelligence strategy, the diagnostic and classifying capabilities of CAD system are meeting the levels of radiologists and clinicians. Thus, it shifts the CAD system from second opinion approach to a high utility tool. This article reviews the strategies and algorithms developed using artificial intelligence for the foremost cancer diagnosis and classification which overcomes the challenges in the traditional method. In addition, the possible direction of AI in medical aspects is also discussed in this study.

Ethics of Glaucoma Widgets.

How to cite this article: Dada T, Ramesh P, Sethi A, et al. Ethics of Glaucoma Widgets. J Curr Glaucoma Pract 2020;14(3):77-80.

Meditation: A Polypill for Comprehensive Management of Glaucoma Patients.

Meditation is an ancient behavioral intervention, however, its benefits for achieving holistic health have been highlighted in recent times with rigorous scientific studies revealing its benefits in many chronic diseases. It has been specially found useful in neurodegenerative diseases and recent evidence points to the positive effects of meditation in preserving gray and white matter in the adult brain. It is also a potential therapy to downregulate processes implicated in brain aging and confer "neuroprotection"-something we all look forward to for our glaucoma patients. In the current review, we evaluate the benefits of meditation practice for the glaucoma patient and support for its candidature as adjunctive therapy for glaucoma patients. It has multiple potential benefits for normal-pressure and high-pressure glaucoma patients including a reduction in intraocular pressure, increasing cerebral blood flow and oxygenation, and decreasing action of the sympathetic nervous system with a corresponding increase in parasympathetic nervous system activity. Meditation leads to a "relaxation response" mediated by nitric oxide with decrease in the stress hormone cortisol, increase in neurotrophins and mitochondrial energy production, and improves the overall quality of life of glaucoma patients. It can also benefit caregivers of glaucoma patients and health care providers. It appears that meditation can function as a multifaceted management approach for glaucoma using the natural potential of the human body and target not only the eye but the patient behind the eye to ameliorate this "sick eye in a sick body" condition.

Generation and characterization of a bivalent protein boost for future clinical trials: HIV-1 subtypes CR01_AE and B gp120 antigens with a potent adjuvant.

The RV144 Phase III clinical trial with ALVAC-HIV prime and AIDSVAX B/E subtypes CRF01_AE (A244) and B (MN) gp120 boost vaccine regime in Thailand provided a foundation for the future development of improved vaccine strategies that may afford protection against the human immunodeficiency virus type 1 (HIV-1). Results from this trial showed that immune responses directed against specific regions V1V2 of the viral envelope (Env) glycoprotein gp120 of HIV-1, were inversely correlated to the risk of HIV-1 infection. Due to the low production of gp120 proteins in CHO cells (2-20 mg/L), cleavage sites in V1V2 loops (A244) and V3 loop (MN) causing heterogeneous antigen products, it was an urgent need to generate CHO cells harboring A244 gp120 with high production yields and an additional, homogenous and uncleaved subtype B gp120 protein to replace MN used in RV144 for the future clinical trials. Here we describe the generation of Chinese Hamster Ovary (CHO) cell lines stably expressing vaccine HIV-1 Env antigens for these purposes: one expressing an HIV-1 subtype CRF01_AE A244 Env gp120 protein (A244.AE) and one expressing an HIV-1 subtype B 6240 Env gp120 protein (6240.B) suitable for possible future manufacturing of Phase I clinical trial materials with cell culture expression levels of over 100 mg/L. The antigenic profiles of the molecules were elucidated by comprehensive approaches including analysis with a panel of well-characterized monoclonal antibodies recognizing critical epitopes using Biacore and ELISA, and glycosylation analysis by mass spectrometry, which confirmed previously identified glycosylation sites and revealed unknown sites of O-linked and N-linked glycosylations at non-consensus motifs. Overall, the vaccines given with MF59 adjuvant induced higher and more rapid antibody (Ab) responses as well as higher Ab avidity than groups given with aluminum hydroxide. Also, bivalent proteins (A244.AE and 6240.B) formulated with MF59 elicited distinct V2-specific Abs to the epitope previously shown to correlate with decreased risk of HIV-1 infection in the RV144 trial. All together, these results provide critical information allowing the consideration of these candidate gp120 proteins for future clinical evaluations in combination with a potent adjuvant.

Impact of Clear Corneal Incision Morphology on Incision-Site Descemet Membrane Detachment in Conventional and Femtosecond Laser-Assisted Phacoemulsification.

PURPOSE: To assess intraoperative morphology of clear corneal incisions (CCI) and its impact on incision-site descemet membrane detachment (DMD) in conventional phacoemulsification and femtosecond laser-assisted cataract surgery (FLACS). METHODS: Prospective comparative study of 129 eyes that underwent either conventional phacoemulsification (Group I, n = 77) or FLACS (Group II, n = 52) was undertaken at an apex tertiary care ophthalmic setup. In group I, a 2.2-mm metal keratome was used to create a biplanar CCI. In group II, femtosecond laser-assisted biplanar CCI was created with 2.2 mm diameter. Incision architecture and incision-site DMD were assessed using microscope-integrated intraoperative OCT (iOCT) and anterior segment OCT on postoperative day (POD) 1 and 30. Visual acuity was assessed on POD 1 and 30. RESULTS: Smooth slit (SS) or ragged slit (RS) morphology of the proximal opening of CCI was observed immediately after creation [Group I: 68.8% SS, 31.2% RS; Group II: 86.5% SS, 13.5% RS]. DMD was observed in 87.1% cases with RS and 16.3% cases with SS morphology (p < 0.001). DMD was more frequent in group I (Group I = 38/77, Group II = 5/52; p < 0.001) and most commonly observed during the step of stromal hydration (83.7%). DMD was self-resolving and did not persist in any group at 1 month. Visual acuity was comparable in both groups on POD 1 and 30. CONCLUSION: Ragged morphology of proximal opening of CCI is the most important predictive factor for incision-site DMD. Femtosecond-laser CCIs have less incision-site DMD as compared to keratome-assisted CCIs. iOCT provides real-time assessment of CCI morphology and DMD.