Exploring the impact of trifluoromethyl (-CF3) functional group on the anti-cancer activity of isoxazole-based molecules: design, synthesis, biological evaluation and molecular docking analysis.

Herein we report the design and synthesis of a series of fully-substituted 4-(trifluoromethyl)isoxazoles and evaluation of their anti-cancer activities against MCF-7, 4T1 and PC-3 cell lines as a proof of concept study. 4-(Trifluoromethyl)isoxazole is a synthetically challenging class of molecules and very few synthetic methods have been developed so far and all of them suffered from several serious limitations. Recently we developed a novel, metal-free, and general synthetic strategy to access synthetically challenging 4-(trifluoromethyl)isoxazoles starting from readily available chalcones using cheap CF3SO2Na as the source of the -CF3 group and multitasking t BuONO as an oxidant as well as the source of N and O and thus we have overcome the limitations of the previous methods. Based on the structure of an isoxazole-based anti-cancer agent, 3-(3,4-dimethoxyphenyl)-5-(thiophen-2-yl)isoxazole 14, we designed a set of 4-(trifluoromethyl)isoxazoles for synthesis and further anti-cancer evaluation. Among various molecules, 3-(3,4-dimethoxyphenyl)-5-(thiophen-2-yl)-4-(trifluoromethyl)isoxazole 2g (IC50 = 2.63 µM) and 3-(thiophen-2-yl)-5-(4-(thiophen-2-yl)-1H-pyrrol-3-yl)-4-(trifluoromethyl)isoxazole 5 (IC50 = 3.09 µM) exhibited the best anti-cancer activity against the human breast cancer cell-lines (MCF-7), 2g being the lead molecule among all. Interestingly, 2g is found to be almost 8 times more active compared to its non-trifluoromethylated analogue, i.e., 3-(3,4-dimethoxyphenyl)-5-(thiophen-2-yl)isoxazole 14 (IC50 = 19.72 µM) which revealed the importance of a 'CF3' moiety in enhancing the anti-cancer activity of 14. Further studies such as apoptosis induction, cell cycle analysis, and nuclear staining revealed an apoptotic cell death mechanism. The in silico molecular docking, induced fit analysis, and ADME studies further supported the effect of a -CF3 moiety on the enhancement of anti-cancer activity of isoxazole-based anti-cancer molecules. Further exploration of the biodistribution and therapeutic efficacy of lead 2gin vivo holds significant promise, positioning it as a potential candidate for anticancer therapy.

Electricity-driven, oxidative C-H selenylative and tellurylative annulation of N-(2-alkynyl)anilines: sustainable synthesis of 3-selanyl/tellanylquinolines.

A metal- and oxidant-free, radical C-H selenylative and tellurylative annulation of N-(2-alkynyl)anilines with diorganyl dichalcogenides is developed under electrochemical conditions for the sustainable synthesis of valuable 3-selanyl/tellanylquinolines up to 92% yield at room temperature. The developed protocol required only electricity as the green reagent and offers high atom economy, broad substrate scope, and efficient scalability.

meta-Fluorophores: an uncharted ocean of opportunities.

meta-Fluorophores (MFs) are unique ultra-light (in terms of molecular weight (MW)) fluorophores exhibiting luminescence with a wide colour gamut ranging from blue to the NIR. Single benzenic MFs are easy to synthesize, are quite bright (with photoluminescence quantum yield (PLQY) as high as 63%) and exhibit very large Stokes shift (as high as 260 nm (8965 cm-1)), with large solvatochromic shift (as high as 175 nm), and very long excited-state-lifetime (as high as 26 ns) for such ultra-light fluorophores. An emission maximum of ≥600 nm has been achieved with an MF in a polar medium having a MW of only 177 g mol-1 and in a nonpolar medium having MW of only 255 g mol-1; therefore, a large-sized π-conjugated para-fluorophore is no longer a prerequisite for red/NIR emission. Structurally varied MFs pave the way for creating an ocean of opportunities and are thus promising for replacing para-fluorophores for different applications, ranging from bioimaging to LEDs.

Unleashing the power of porphyrin photosensitizers: Illuminating breakthroughs in photodynamic therapy.

This comprehensive review provides the current trends and recent developments of porphyrin-based photosensitizers. We discuss their evolution from first-generation to third-generation compounds, including cutting-edge nanoparticle-integrated derivatives, and explores their pivotal role in advancing photodynamic therapy (PDT) for enhanced cancer treatment. Integrating porphyrins with nanoparticles represents a promising avenue, offering improved selectivity, reduced toxicity, and heightened biocompatibility. By elucidating recent breakthroughs, innovative methodologies, and emerging applications, this review provides a panoramic snapshot of the dynamic field, addressing challenges and charting prospects. With a focus on harnessing reactive oxygen species (ROS) through light activation, PDT serves as a minimally invasive therapeutic approach. This article offers a valuable resource for researchers, clinicians, and PDT enthusiasts, highlighting the potential of porphyrin photosensitizers to improve the future of cancer therapy.

Critical Analysis and Optimization of Stoichiometric Ratio of Drug-Coformer on Cocrystal Design: Molecular Docking, In Vitro and In Vivo Assessment.

In this present research, an attempt has been made to address the influence of drug-coformer stoichiometric ratio on cocrystal design and its impact on improvement of solubility and dissolution, as well as bioavailability of poorly soluble telmisartan. The chemistry behind cocrystallization and the optimization of drug-coformer molar ratio were explored by the molecular docking approach, and theoretical were implemented practically to solve the solubility as well as bioavailability related issues of telmisartan. A new multicomponent solid form, i.e., cocrystal, was fabricated using different molar ratios of telmisartan and maleic acid, and characterized by SEM, DSC and XRD studies. The molecular docking study suggested that specific molar ratios of drug-coformer can successfully cluster with each other and form a specific geometry with favourable energy conformation to form cocrystals. Synthesized telmisartan-maleic acid cocrystals showed remarkable improvement in solubility and dissolution of telmisartan by 9.08-fold and 3.11-fold, respectively. A SEM study revealed the formation of cocrystals of telmisartan when treated with maleic acid. DSC and XRD studies also confirmed the conversion of crystalline telmisartan into its cocrystal state upon treating with maleic acid. Preclinical investigation revealed significant improvement in the efficacy of optimized cocrystals in terms of plasma drug concentration, indicating enhanced bioavailability through improved solubility as well as dissolution of telmisartan cocrystals. The present research concluded that molecular docking is an important path in selecting an appropriate stoichiometric ratio of telmisartan: maleic acid to form cocrystals and improve the solubility, dissolution, and bioavailability of poorly soluble telmisartan.

Synthesis of (4-Trifluoromethyl)isoxazoles through a Tandem Trifluoromethyloximation/Cyclization/Elimination Reaction of α,ß-Unsaturated Carbonyls.

We disclose a metal-free, cascade regio- and stereoselective trifluormethyloximation, cyclization, and elimination strategy with readily available α,ß-unsaturated carbonyl compounds to access a wide variety of pharmaceutically potential heteroaromatics, i.e., 4-(trifluoromethyl)isoxazoles including a trifluoromethyl analogue of an anticancer agent. The transformation requires only a couple of commercially available and cheap reagents i.e., CF3SO2Na as the trifluoromethyl source, and tBuONO as an oxidant as well as a source of N and O. Notably, 5-alkenyl-4-(trifluoromethyl)isoxazoles were further synthetically diversified to a new class of biheteroaryls, i.e., 5-(3-pyrrolyl)-4-(trifluoromethyl)isoxazoles. Mechanistic studies revealed a radical pathway for the reaction.

Highly sensitive protein detection by aptamer-based single-molecule kinetic fingerprinting.

Sensitive assays of protein biomarkers play critical roles in clinical diagnostics and biomedical research. Such assays typically employ immunoreagents such as monoclonal antibodies that suffer from several drawbacks, including relatively tedious production, significant batch-to-batch variability, and challenges in site-specific, stoichiometric modification with fluorophores or other labels. One proposed alternative to such immunoreagents, nucleic acid aptamers generated by systematic evolution of ligand by exponential enrichment (SELEX), can be chemically synthesized with much greater ease, precision, and reproducibility than antibodies. However, most aptamers exhibit relatively poor affinity, yielding low sensitivity in the assays employing them. Recently, single molecule recognition through equilibrium Poisson sampling (SiMREPS) has emerged as a platform for detecting proteins and other biomarkers with high sensitivity without requiring high-affinity detection probes. In this manuscript, we demonstrate the applicability and advantages of aptamers as detection probes in SiMREPS as applied to two clinically relevant biomarkers, VEGF165 and IL-8, using a wash-free protocol with limits of detection in the low femtomolar range (3-9 fM). We show that the kinetics of existing RNA aptamers can be rationally optimized for use as SiMREPS detection probes by mutating a single nucleotide in the conserved binding region or by shortening the aptamer sequence. Finally, we demonstrate the detection of endogenous IL-8 from human serum at a concentration below the detection limit of commercial ELISAs.

Predominance of genomically defined A lineage of HPV16 over D lineage in Indian patients from eastern India with squamous cell carcinoma of the cervix in association with distinct oncogenic phenotypes.

Human papillomavirus type-16 (HPV16) is classified into lineages, A, B, C and D and 10 sub-lineages portraying variable infectivity, persistence, and cytological outcomes, however, with geographical variations. Our objective was to delineate the distinctive features of lineages among cervical squamous cell carcinoma (SCC) in the eastern region of India. A total of 145 SCC cases and 24 non-malignant specimens, harboring episomal HPV16, were included. The presence of higher proportion of lineage A over D was observed among SCC cases (86.89% A1, 8.97% D1 and 4.14% D2), while only A1 sub-lineage viruses were found among control specimens. Among the A1 viruses, an association of variants in the E5 (Y44L, I65V), E6 (L83V) genes and LCR: C7577T with SCC, with combined Odd's ratio (95% CI) of 20.5(4.61-91.25) was observed. Network analyses revealed the presence of 10 clades of lineage A viruses comprising of 64 HPV16 genomes harboring the risk alleles. High episomal HPV16 DNA copy numbers (adjusted p-value= 0.0271) and E7 mRNA expression (p-value=0.000017) predominated in SCC with lineage A, over D. Our study highlights the distinctive modalities of oncogenicity among different HPV16 lineages.

Synthesis of Organosulfur and Related Heterocycles under Mechanochemical Conditions.

In the last few decades, ball-milling has received tremendous attention as a "green tool" for conducting various challenging organic transformations under transition-metal-free and solvent-free conditions. Organosulfur and related heterocycles are ubiquitous in numerous biologically active molecules with potential applications, and those molecules could be synthesized from readily available starting materials under mechanochemical conditions without using any hazardous chemical or solvent. This synopsis highlights the green strategies developed in recent times to synthesize organosulfur and related heterocycles under ball-milling conditions.

Iodine-Catalyzed Methylthiolative Annulation of 2-Alkynyl Biaryls with DMSO: A Metal-Free Approach to 9-Sulfenylphenanthrenes.

An iodine-catalyzed sustainable, cost-effective, and atom-economic synthetic methodology is developed to synthesize a wide variety of valuable sulfenylphenanthrenes and polycyclic heteroaromatics in moderate to high yield through electrophilic thiolative annulation of 2-alkynyl biaryls (6-endo-dig cyclization) using methyl sulfoxides such as dimethyl sulfoxide (DMSO) as the sulfur source under transition-metal-free conditions. The transformation requires only iodine in a catalytic amount and trifluoroacetic anhydride. Notably, DMSO played multiple roles such as methylthiolating reagent, oxidant, and solvent in this reaction.

Direct Kinetic Fingerprinting for High-Accuracy Single-Molecule Counting of Diverse Disease Biomarkers.

Methods for detecting and quantifying disease biomarkers in biofluids with high specificity and sensitivity play a pivotal role in enabling clinical diagnostics, including point-of-care tests. The most widely used molecular biomarkers include proteins, nucleic acids, hormones, metabolites, and other small molecules. While numerous methods have been developed for analyzing biomarkers, most techniques are challenging to implement for clinical use due to insufficient analytical performance, high cost, and/or other practical shortcomings. For instance, the detection of cell-free nucleic acid (cfNA) biomarkers by digital PCR and next-generation sequencing (NGS) requires time-consuming nucleic acid extraction steps, often introduces enzymatic amplification bias, and can be costly when high specificity is required. While several amplification-free methods for detecting cfNAs have been reported, these techniques generally suffer from low specificity and sensitivity. Meanwhile, the quantification of protein biomarkers is generally performed using immunoassays such as enzyme-linked immunosorbent assay (ELISA); the analytical performance of these methods is often limited by the availability of antibodies with high affinity and specificity as well as the significant nonspecific binding of antibodies to assay surfaces. To address the drawbacks of existing biomarker detection methods and establish a universal diagnostics platform capable of detecting different types of analytes, we have developed an amplification-free approach, named single-molecule recognition through equilibrium Poisson sampling (SiMREPS), for the detection of diverse biomarkers with arbitrarily high specificity and single-molecule sensitivity. SiMREPS utilizes the transient, reversible binding of fluorescent detection probes to immobilized target molecules to generate kinetic fingerprints that are detected by single-molecule fluorescence microscopy. The analysis of these kinetic fingerprints enables nearly perfect discrimination between specific binding to target molecules and any nonspecific binding. Early proof-of-concept studies demonstrated the in vitro detection of miRNAs with a limit of detection (LOD) of approximately 1 fM and >500-fold selectivity for single-nucleotide polymorphisms. The SiMREPS approach was subsequently expanded to the detection of rare mutant DNA alleles from biofluids at mutant allele fractions of as low as 1 in 1 million, corresponding to a specificity of >99.99999%. Recently, SiMREPS was generalized to protein quantification using dynamically binding antibody probes, permitting LODs in the low-femtomolar to attomolar range. Finally, SiMREPS has been demonstrated to be suitable for the in situ detection of miRNAs in cultured cells, the quantification of small-molecule toxins and drugs, and the monitoring of telomerase activity at the single-molecule level. In this Account, we discuss the principles of SiMREPS for the highly specific and sensitive detection of molecular analytes, including considerations for assay design. We discuss the generality of SiMREPS for the detection of very disparate analytes and provide an overview of data processing methods, including the expansion of the dynamic range using super-resolution analysis and the improvement of performance using deep learning algorithms. Finally, we describe current challenges, opportunities, and future directions for the SiMREPS approach.

Ultraspecific analyte detection by direct kinetic fingerprinting of single molecules.

The detection and quantification of biomarkers have numerous applications in biological research and medicine. The most widely used methods to detect nucleic acids require amplification via the polymerase chain reaction (PCR). However, errors arising from the imperfect copying fidelity of DNA polymerases, limited specificity of primers, and heat-induced damage reduce the specificity of PCR-based methods, particularly for single-nucleotide variants. Furthermore, not all analytes can be amplified efficiently. While amplification-free methods avoid these pitfalls, the specificity of most such methods is strictly constrained by probe binding thermodynamics, which for example hampers detection of rare somatic mutations. In contrast, single-molecule recognition through equilibrium Poisson sampling (SiMREPS) provides ultraspecific detection with single-molecule and single-nucleotide sensitivity by monitoring the repetitive interactions of a fluorescent probe with surface-immobilized targets. In this review, we discuss SiMREPS in comparison with other analytical approaches, and describe its utility in quantifying a range of nucleic acids and other analytes.

Direct kinetic fingerprinting and digital counting of single protein molecules.

The sensitive and accurate quantification of protein biomarkers plays important roles in clinical diagnostics and biomedical research. Sandwich ELISA and its variants accomplish the capture and detection of a target protein via two antibodies that tightly bind at least two distinct epitopes of the same antigen and have been the gold standard for sensitive protein quantitation for decades. However, existing antibody-based assays cannot distinguish between signal arising from specific binding to the protein of interest and nonspecific binding to assay surfaces or matrix components, resulting in significant background signal even in the absence of the analyte. As a result, they generally do not achieve single-molecule sensitivity, and they require two high-affinity antibodies as well as stringent washing to maximize sensitivity and reproducibility. Here, we show that surface capture with a high-affinity antibody combined with kinetic fingerprinting using a dynamically binding, low-affinity fluorescent antibody fragment differentiates between specific and nonspecific binding at the single-molecule level, permitting the direct, digital counting of single protein molecules with femtomolar-to-attomolar limits of detection (LODs). We apply this approach to four exemplary antigens spiked into serum, demonstrating LODs 55- to 383-fold lower than commercially available ELISA. As a real-world application, we establish that endogenous interleukin-6 (IL-6) can be quantified in 2-µL serum samples from chimeric antigen receptor T cell (CAR-T cell) therapy patients without washing away excess serum or detection probes, as is required in ELISA-based approaches. This kinetic fingerprinting thus exhibits great potential for the ultrasensitive, rapid, and streamlined detection of many clinically relevant proteins.

Cervical cancer subtypes harbouring integrated and/or episomal HPV16 portray distinct molecular phenotypes based on transcriptome profiling of mRNAs and miRNAs.

Heterogeneity in cervical cancers (CaCx) in terms of HPV16 physical status prompted us to investigate the mRNA and miRNA signatures among the different categories of CaCx samples. We performed microarray-based mRNA expression profiling and quantitative real-time PCR-based expression analysis of some prioritised miRNAs implicated in cancer-related pathways among various categories of cervical samples. Such samples included HPV16-positive CaCx cases that harboured either purely integrated HPV16 genomes (integrated) and those that harboured episomal viral genomes, either pure or concomitant with integrated viral genomes (episomal), which were compared with normal cervical samples that were either HPV negative or positive for HPV16. The mRNA expression profile differed characteristically between integrated and episomal CaCx cases for enriched biological pathways. miRNA expression profiles also differed among CaCx cases compared with controls (upregulation-miR-21, miR-16, miR-205, miR-323; downregulation-miR-143, miR-196b, miR-203, miR-34a; progressive upregulation-miR-21 and progressive downregulation-miR-143, miR-34a, miR-196b and miR-203) in the order of HPV-negative controls, HPV16-positive non-malignant samples and HPV16-positive CaCx cases. miR-200a was upregulated in HPV16-positive cervical tissues irrespective of histopathological status. Expression of majority of the predicted target genes was negatively correlated with their corresponding miRNAs, irrespective of the CaCx subtypes. E7 mRNA expression correlated positively with miR-323 expression among episomal cases and miR-203, among integrated cases. miR-181c expression was downregulated only among the episomal CaCx cases and negatively correlated with protein coding transcript of the proliferative target gene, CKS1B of the significantly enriched "G2/M DNA Damage Checkpoint Regulation" pathway among CaCx cases. Thus, the two CaCx subtypes are distinct entities at the molecular level, which could be differentially targeted for therapy. In fact, availability of a small molecule inhibitor of CKS1B, suggests that drugging CKS1B could be a potential avenue of treating the large majority of CaCx cases harbouring episomal HPV16.

Base-Promoted Synthesis of 2-Aryl Quinazolines from 2-Aminobenzylamines in Water.

A transition-metal-free procedure for the synthesis of a highly valuable class of heteroaromatics, quinazolines, was developed by using easily available 2-aminobenzylamines and α,α,α-trihalotoluenes. The transformation proceeded smoothly in the presence of only sodium hydroxide and molecular oxygen in water at 100 °C, furnishing a variety of 2-aryl quinazolines. The crystallization process of the crude reaction mixture for the purification of the solid products circumvents huge solvent-consuming workup and column chromatographic techniques, which make the overall process more sustainable and economical.

Cyanopyrimidine-Carbazole Hybrid Host Materials for High-Efficiency and Low-Efficiency Roll-Off TADF OLEDs.

Two isomeric host materials (Sy and Asy) comprising carbazole (donor) and CN-substituted pyrimidine (acceptor) were synthesized, characterized, and utilized as host materials for green and blue thermally activated delayed fluorescence (TADF) organic light emitting diodes (OLEDs). Both molecules have high triplet energy and small energy difference between singlet and triplet states, leading to feasible TADF. The different linking topologies of carbazole and CN groups on the pyrimidine core provide distinct photophysical properties and molecular packing manners, which further influence the efficiency as they served as hosts in TADF OLEDs. As compared to Asy-based cases, the Sy-hosted TADF OLED device gave higher maximum external quantum efficiencies (EQE) of 24.0% (vs 22.5%) for green (4CzIPN as a dopant) and 20.4% (vs 15.0%) for blue (2CzTPN as a dopant) and low efficiency roll-off. The high horizontal dipole ratio (Θ ≈ 88%) for both emitters dispersed in Sy and Asy hosts accounts for the high device efficiency. A clear molecular structure-physical property-device performance relationship has been established to highlight the importance of symmetrical structure in TADF host material design.

Impact of viral and host DNA methylations on HPV16-related cervical cancer pathogenesis.

Epigenetic alterations within human papillomavirus (HPV) and host cellular genomes are known to occur during cervical carcinogenesis. Our objective was to analyse the influence of (1) methylation within two immunostimulatory CpG motifs within HPV16 E6 and E7 genes around the viral late promoter and their correlation, if any, with expression deregulation of host receptor (TLR9) and DNA methyltransferases (DNMT1, DNMT3A and DNMT3B) and (2) global DNA methylation levels within CpGs of the repetitive Alu sequences, on cervical cancer (CaCx) pathogenesis. Significantly higher proportions of CaCx samples portrayed methylation in immunostimulatory CpG motifs, compared to HPV16-positive non-malignant samples, with cases harbouring episomal HPV16 showing decreased methylation compared to those with viral integration. A significant linear trend of TLR9 upregulation was recorded in the order of HPV-negative controls < HPV16-positive non-malignant samples < HPV16-positive CaCx cases. TLR9 upregulation in cases with episomal HPV16 was again higher among those with non-methylated immunostimulatory CpG motifs. Comparison of cases with HPV-negative controls revealed that DNMT3A was significantly downregulated only among integrated cases, DNMT3B was significantly overexpressed among both categories of cases, although at variable levels, while DNMT1 failed to show any deregulated expression among the cases. Global host DNA hypomethylation, also showed a significant linear increasing trend through the progressive CaCx development stages mentioned above and was most prominently higher among cases with episomal HPV16 as opposed to viral integration. Thus, HPV16 and host methylations appear to influence CaCx pathogenesis, with differential molecular signatures among CaCx cases with episomal and integrated HPV16.

Visible-Light-Induced Synthesis of Carbazoles by in Situ Formation of Photosensitizing Intermediate.

A visible-light-induced synthesis of N-H carbazoles from easily accessible 2,2'-diaminobiaryls in the absence of any external photosensitizer is reported. The process only requires tBuONO and natural resources, visible light, and molecular oxygen for the synthesis of N-H carbazoles. Experimental and computational studies support that the in situ formation of a visible-light-absorbing photosensitizing intermediate, benzocinnoline N-imide, is responsible for the activation of triplet molecular oxygen to singlet oxygen that, in turn, promotes the synthesis of carbazole.

Extended Study of Visible-Light-Induced Photocatalytic [4 + 2] Benzannulation: Synthesis of Polycyclic (Hetero)Aromatics.

Herein we report an extended study of [4 + 2] benzannulation reactions of 2-(hetero)aryl-substituted anilines with alkynes by visible light photocatalysis. The method requires the use of tBuONO as a diazotizing agent and 0.3 mol % of fac-Ir(ppy)3 as a photocatalyst at room temperature. The reaction proceeded in a chemo- and regioselective manner with high functional group tolerance under mild conditions allowing the preparation of a wide variety of polycyclic (hetero)aromatic compounds, including phenanthrenes, in moderate to high yields. This procedure is amenable to gram-scale synthesis of 9-phenylphenanthrene.

Molecular origin of photoluminescence of carbon dots: aggregation-induced orange-red emission.

The molecular origin of the photoluminescence of carbon dots (CDs) is not known. This restricts the design of CDs with desired optical properties. We have synthesized CDs starting from carbohydrates by employing a simple synthesis method. We were able to demonstrate that the CDs are composed of aggregated hydroxymethylfurfural (HMF) derivatives. The optical properties of these CDs are quite unique. These CDs exhibit an excitation-independent PL emission maximum in the orange-red region (λ ∼ 590 nm). These CDs also exhibit excitation as well as monitoring wavelength-independent single exponential PL decay. These observations indicate that only one type of chromophore (HMF derivative) is present within the CDs. Several HMF derivatives are aggregated within the CDs; therefore, the aggregated structure cause a large Stokes shift (∼150 nm). By several control experiments, we showed that the same aggregated chromophore unit (HMF derivative), and not the individual fluorophores, is the fluorescing unit. The emission maximum and the single exponential PL lifetime are independent of the polarity of the medium. The existence of a low-lying trap state could be reduced quite significantly. A model has been proposed to explain the interesting steady state and dynamical photoluminescence behaviour of the CDs. As the molecular origin of their photoluminescence is known, CDs with desired optical properties can be designed.