Proteinaceous Lymphadenopathy Masquerading as Malignancy: a Rare Case Presenting a Diagnostic Dilemma.

Proteinaceous lymphadenopathy (PLD) is a rare poorly defined, underrecognized entity of uncertain etiology, characterized by massive deposition of amorphous, acellular, eosinophilic, PAS-positive material within an enlarged lymph node. We report an unusual case of a 46-year-old female with a large abdominal lump in the left lumbar region with inguinal lymphadenopathy. Contrast-enhanced computed tomography (CECT) showed multiple variable-sized lobulated non-enhancing soft tissue attenuated masses showing multiple peripheral and central calcific foci in the right para-aortic, bilateral iliac region, pelvis on the left side and left inguinal region. No evidence of any abnormal hypermetabolic focus was found in the neck, chest, abdomen, and pelvis on fluorodeoxyglucose positron emission tomography. A large, well-defined, non-FDG avid mass lesion with significant central and peripheral calcification in the left iliac fossa, abutting the descending colon, was seen. A biopsy of left-sided inguinal lymph nodes revealed large masses of an amorphous, acellular, eosinophilic material with areas of mature lymphoid cell aggregates interspersed between the pink amorphous materials. A final impression of proteinaceous lymphadenopathy was given. Proteinaceous lymphadenopathy is a benign condition with often a large mass masquerading as malignancy. It is a major therapeutic challenge for pathologists and clinicians. Histopathologists need to be vigilant in such cases and be aware of the morphological appearances in such cases.

Experimental and computational investigation of the α-amylase catalyzed Friedel-Crafts reaction of isatin to access symmetrical and unsymmetrical 3,3',3''-trisindoles.

Trisindoles are of tremendous interest due to their wide range of biological activities. In this context, a number of methods have been reported in the past to synthesize 3,3',3''-trisindoles. However, most of the methods are only able to produce symmetrical 3,3',3''-trisindoles. Herein, we develop a sustainable and efficient approach to synthesize symmetrical as well as unsymmetrical 3,3',3''-trisindoles in a very selective manner using the α-amylase enzyme as a catalyst. Furthermore, various differently substituted isatin and indoles were used to prove the generality of the protocol and symmetrical or unsymmetrical 3,3',3''-trisindoles were obtained in 43-97% isolated yields. Next, a probable mechanism is proposed and investigated using molecular dynamics (MD) investigation to gain more insight into the role of residues available in the active site of the α-amylase enzyme. These studies revealed that Glu230, Lys209, and Asp206 in the active site of α-amylase play an important role in this catalysis. Moreover, the DFT studies suggested the formation of bisindole and alkylideneindolenine intermediates during the transformation. We synthesized four different biologically important 3,3',3''-trisindoles on a gram scale, which proved the robustness and scalability of this protocol.

Enzymatic Synthesis of Indole-Based Imidazopyridine using α-Amylase.

The imidazo[1,2-a]pyridine scaffold has gained significant attention due to its presence as a lead structure in several commercially available pharmaceuticals like zolimidine, zolpidem, olprinone, soraprazan, etc. Further, indole-based imidazo[1,2-a]pyridine derivatives have been found interesting due to their anticancer and antibacterial activities. However, limited methods have been reported for the synthesis of indole-based imidazo[1,2-a]pyridines. In this study, we have successfully developed a biocatalytic process for synthesizing indole-based imidazo[1,2-a]pyridine derivatives using the α-amylase enzyme catalyzed Groebke-Blackburn-Bienayme (GBB) multicomponent reaction of 2-aminopyridine, indole-3-carboxaldehyde, and isocyanide. The generality and robustness of this protocol were shown by synthesizing differently substituted indole-based imidazo[1,2-a]pyridines in good isolated yields. Furthermore, to make α-amylase a reusable catalyst for GBB multicomponent reaction, it was immobilized onto magnetic metal-organic framework (MOF) materials [Fe3 O4 @MIL-100(Fe)] and found reusable up to four consecutive catalytic cycles without the significant loss in catalytic activity.

Divergent and Selective Synthesis of 3-Alkylidene Oxindoles Using Pd-Catalyzed Multicomponent Reaction.

Herein, a divergent and selective synthesis of (E)-3-alkylidene oxindole, which is a highly valuable framework due to its presence in biologically important molecules, via palladium-catalyzed multicomponent reaction of 3-diazo oxindole, isocyanide, and aniline has been developed. Further, the feasibility of the reaction was demonstrated by employing differently substituted 3-diazo oxindoles, isocyanides, and anilines as starting material and obtaining the corresponding products in 31-83% isolated yields. Besides, a plausible mechanism has been presented and further investigated using DFT calculations which suggest the formation of the Pd-carbene complex and ketenimine intermediate as the key step during the catalytic cycle.

Sclerosing angiomatoid nodular transformation of spleen: A rare case report.

ABSTRACT: Sclerosing angiomatoid nodular transformation (SANT) is a reactive non-neoplastic, rare vascular lesion of the spleen. The histology shows multiple angiomatoid nodules surrounded by proliferative stroma. A 31-year-old lady presented with an abdominal mass for 6 months. Contrast-enhanced computed tomography (CECT) abdomen was suggestive of hemangiopericytoma/hemangioendothelioma. An open splenectomy was performed, and the resected specimen was sent for histopathology examination. The gross examination showed a bosselated mass present at the lower pole of the spleen measuring 8 × 8 cm with peripherally located coalescing red-brown nodules embedded in a dense fibrous stroma on the cut surface. On microscopy, multiple circumscribed angiomatoid nodules comprising irregular slit-like vascular channels lined by plump endothelial cells were seen embedded in dense sclerotic stroma. Because of the lack of specific diagnostic features, it is difficult to diagnose SANT clinically and radiologically. However, the typical histopathological findings are a clue in clinching the diagnosis.

Production of a recyclable nanobiocatalyst to synthesize quinazolinone derivatives.

Nanobiocatalysts (NBCs) are an emerging innovation that paves the way toward sustainable and eco-friendly endeavors. In the quest for a robust and reusable nanobiocatalyst, herein, we report a nanobiocatalyst, namely CALB@MrGO, developed via immobilizing Candida antarctica lipase B onto the surface of Fe3O4-decorated reduced graphene oxide (MrGO). Next, the enormous potential of the NBC (CALB@MrGO) was checked by employing it to synthesize clinically important quinazolinone derivatives in good to excellent yield (70-95%) using differently substituted aryl aldehydes with 2-aminobenzamide. Further, the synthetic utility and generality of this protocol was proved by setting up a gram-scale reaction, which afforded the product in 87% yield. The green chemistry metrics calculated for the gram-scale reaction those prove the greenness of this protocol.

A Pd-catalyzed one-pot cascade consisting of C-C/C-O/N-N bond formation to access benzoxazine fused 1,2,3-triazoles.

A Pd-catalyzed one-pot cascade consisting of C-C/C-O/N-N bond formation to access clinically important fused 1,2,3-triazoles using N-aryl-α-(tosylhydrazone)acetamides with isocyanide has been developed. Besides, various substitutions on the N-aryl part of acetamides along with different isocyanides show good compatibility in this protocol. Next, two plausible mechanistic routes were proposed; however, one of the routes was more favourable which involved the formation of a benzoxazine ring first followed by the realization of a triazole ring. Additionally, the more favourable mechanistic route was investigated using DFT studies which suggests that the formations of a Pd(II)-isocyanide complex and α-diazoimino intermediates were key steps in the catalytic cycle.

Agriculture, dairy and fishery farming practices and greenhouse gas emission footprint: a strategic appraisal for mitigation.

Rising global population would force farmers to amplify food production substantially in upcoming 3-4 decades. The easiest way to increase grain production is through expanding cropping area by clearing uncultivated land. This is attained by permitting deadly loss of carbon (C) stocks, jeopardizing ecosystem biodiversity and deteriorating environmental quality. We aim to propose key agronomical tactics, livestock management strategy and advance approaches for aquaculture to increase productivity and simultaneously reduce the environmental impacts of farming sector. For this, we considered three major sectors of farming, i.e. agriculture, fishery and dairy. We collected literatures stating approaches or technologies that could reduce GHG emission from these sectors. Thereafter, we synthesized strategies or options that are more feasible and accessible for inclusion in farm sector to reduce GHG emission. Having comprehensively reviewed several publications, we propose potential strategies to reduce GHG emission. Agronomic practices like crop diversification, reducing summer fallow, soil organic carbon sequestration, tillage and crop residue management and inclusion of N2-fixing pulses in crop rotations are some of those. Livestock management through changing animals' diets, optimal use of the gas produced from manures, frequent and complete manure removal from animal housing and aquaculture management strategies to improve fish health and improve feed conversion efficiency could reduce their GHG emission footprint too. Adapting of effective and economic practices GHG emission footprint reduction potential of farming sector could make farming sector a C neutral enterprise. To overcome the ecological, technological and institutional barriers, policy on trade, tax, grazing practice and GHG pricing should be implemented properly.

Application of a MYC degradation screen identifies sensitivity to CDK9 inhibitors in KRAS-mutant pancreatic cancer.

Stabilization of the MYC oncoprotein by KRAS signaling critically promotes the growth of pancreatic ductal adenocarcinoma (PDAC). Thus, understanding how MYC protein stability is regulated may lead to effective therapies. Here, we used a previously developed, flow cytometry-based assay that screened a library of >800 protein kinase inhibitors and identified compounds that promoted either the stability or degradation of MYC in a KRAS-mutant PDAC cell line. We validated compounds that stabilized or destabilized MYC and then focused on one compound, UNC10112785, that induced the substantial loss of MYC protein in both two-dimensional (2D) and 3D cell cultures. We determined that this compound is a potent CDK9 inhibitor with a previously uncharacterized scaffold, caused MYC loss through both transcriptional and posttranslational mechanisms, and suppresses PDAC anchorage-dependent and anchorage-independent growth. We discovered that CDK9 enhanced MYC protein stability through a previously unknown, KRAS-independent mechanism involving direct phosphorylation of MYC at Ser62 Our study thus not only identifies a potential therapeutic target for patients with KRAS-mutant PDAC but also presents the application of a screening strategy that can be more broadly adapted to identify regulators of protein stability.

Myoglobin-Catalyzed C-H Functionalization of Unprotected Indoles.

Functionalized indoles are recurrent motifs in bioactive natural products and pharmaceuticals. While transition metal-catalyzed carbene transfer has provided an attractive route to afford C3-functionalized indoles, these protocols are viable only in the presence of N-protected indoles, owing to competition from the more facile N-H insertion reaction. Herein, a biocatalytic strategy for enabling the direct C-H functionalization of unprotected indoles is reported. Engineered variants of myoglobin provide efficient biocatalysts for this reaction, which has no precedents in the biological world, enabling the transformation of a broad range of indoles in the presence of ethyl α-diazoacetate to give the corresponding C3-functionalized derivatives in high conversion yields and excellent chemoselectivity. This strategy could be exploited to develop a concise chemoenzymatic route to afford the nonsteroidal anti-inflammatory drug indomethacin.

Use of Protein Kinase-Focused Compound Libraries for the Discovery of New Inositol Phosphate Kinase Inhibitors.

Inositol hexakisphosphate kinases (IP6Ks) regulate a myriad of cellular processes, not only through their catalytic activity (which synthesizes InsP7, a multifunctional inositol pyrophosphate signaling molecule) but also through protein-protein interactions. To further study the enzymatic function and distinguish between these different mechanisms, specific inhibitors that target IP6K catalytic activity are required. Only one IP6K inhibitor is commonly used: N2-( m-(trifluoromethyl)benzyl) N6-( p-nitrobenzyl)purine (TNP). TNP is, however, compromised by weak potency, inability to distinguish between IP6K isoenzymes, off-target activities, and poor pharmacokinetic properties. Herein, we describe a new inhibitor discovery strategy, based on the high degree of structural conservation of the nucleotide-binding sites of IP6Ks and protein kinases; we screened for novel IP6K2 inhibitors using a focused set of compounds with features known, or computationally predicted, to target nucleotide binding by protein kinases. We developed a time-resolved fluorescence resonance energy transfer (TR-FRET) assay of adenosine diphosphate (ADP) formation from adenosine triphosphate (ATP). Novel hit compounds for IP6K2 were identified and validated with dose-response curves and an orthogonal assay. None of these inhibitors affected another inositol pyrophosphate kinase, PPIP5K. Our screening strategy offers multiple IP6K2 inhibitors for future development and optimization. This approach will be applicable to inhibitor discovery campaigns for other inositol phosphate kinases.

Highly Diastereo- and Enantioselective Synthesis of Trifluoromethyl-Substituted Cyclopropanes via Myoglobin-Catalyzed Transfer of Trifluoromethylcarbene.

We report an efficient strategy for the asymmetric synthesis of trifluoromethyl-substituted cyclopropanes by means of myoglobin-catalyzed olefin cyclopropanation reactions in the presence of 2-diazo-1,1,1-trifluoroethane (CF3CHN2) as the carbene donor. These transformations were realized using a two-compartment setup in which ex situ generated gaseous CF3CHN2 is processed by engineered myoglobin catalysts expressed in bacterial cells. This approach was successfully applied to afford a variety of trans-1-trifluoromethyl-2-arylcyclopropanes in high yields (61-99%) and excellent diastereo- and enantioselectivity (97-99.9% de and ee). Furthermore, mirror-image forms of these products could be obtained using myoglobin variants featuring stereodivergent selectivity. These reactions provide a convenient and effective biocatalytic route to the stereoselective synthesis of key fluorinated building blocks of high value for medicinal chemistry and drug discovery. This work expands the range of carbene-mediated transformations accessible via metalloprotein catalysts and introduces a potentially general strategy for exploiting gaseous and/or hard-to-handle carbene donor reagents in biocatalytic carbene transfer reactions.

Gram-Scale Synthesis of Chiral Cyclopropane-Containing Drugs and Drug Precursors with Engineered Myoglobin Catalysts Featuring Complementary Stereoselectivity.

Engineered hemoproteins have recently emerged as promising systems for promoting asymmetric cyclopropanations, but variants featuring predictable, complementary stereoselectivity in these reactions have remained elusive. In this study, a rationally driven strategy was implemented and applied to engineer myoglobin variants capable of providing access to 1-carboxy-2-aryl-cyclopropanes with high trans-(1R,2R) selectivity and catalytic activity. The stereoselectivity of these cyclopropanation biocatalysts complements that of trans-(1S,2S)-selective variants developed here and previously. In combination with whole-cell biotransformations, these stereocomplementary biocatalysts enabled the multigram synthesis of the chiral cyclopropane core of four drugs (Tranylcypromine, Tasimelteon, Ticagrelor, and a TRPV1 inhibitor) in high yield and with excellent diastereo- and enantioselectivity (98-99.9% de; 96-99.9% ee). These biocatalytic strategies outperform currently available methods to produce these drugs.

Biocatalytic Synthesis of Allylic and Allenyl Sulfides through a Myoglobin-Catalyzed Doyle-Kirmse Reaction.

The first example of a biocatalytic [2,3]-sigmatropic rearrangement reaction involving allylic sulfides and diazo reagents (Doyle-Kirmse reaction) is reported. Engineered variants of sperm whale myoglobin catalyze this synthetically valuable C-C bond-forming transformation with high efficiency and product conversions across a variety of sulfide substrates (e.g., aryl-, benzyl-, and alkyl-substituted allylic sulfides) and α-diazo esters. Moreover, the scope of this myoglobin-mediated transformation could be extended to the conversion of propargylic sulfides to give substituted allenes. Active-site mutations proved effective in enhancing the catalytic efficiency of the hemoprotein in these reactions as well as modulating the enantioselectivity, resulting in the identification of the myoglobin variant Mb(L29S,H64V,V68F), which is capable of mediating asymmetric Doyle-Kirmse reactions with an enantiomeric excess up to 71 %. This work extends the toolbox of currently available biocatalytic strategies for the asymmetric formation of carbon-carbon bonds.

Chemoenzymatic synthesis and antileukemic activity of novel C9- and C14-functionalized parthenolide analogs.

Parthenolide is a naturally occurring terpene with promising anticancer properties, particularly in the context of acute myeloid leukemia (AML). Optimization of this natural product has been challenged by limited opportunities for the late-stage functionalization of this molecule without affecting the pharmacologically important α-methylene-γ-lactone moiety. Here, we report the further development and application of a chemoenzymatic strategy to afford a series of new analogs of parthenolide functionalized at the aliphatic positions C9 and C14. Several of these compounds were determined to be able to kill leukemia cells and patient-derived primary AML specimens with improved activity compared to parthenolide, exhibiting LC50 values in the low micromolar range. These studies demonstrate that different O-H functionalization chemistries can be applied to elaborate the parthenolide scaffold and that modifications at the C9 or C14 position can effectively enhance the antileukemic properties of this natural product. The C9-functionalized analogs 22a and 25b were identified as the most interesting compounds in terms of antileukemic potency and selectivity toward AML versus healthy blood cells.

Elevated CXC chemokines in urine noninvasively discriminate OAB from UTI.

Overlapping symptoms of overactive bladder (OAB) and urinary tract infection (UTI) often complicate the diagnosis and contribute to overprescription of antibiotics. Inflammatory response is a shared characteristic of both UTI and OAB and here we hypothesized that molecular differences in inflammatory response seen in urine can help discriminate OAB from UTI. Subjects in the age range of (20-88 yr) of either sex were recruited for this urine analysis study. Urine specimens were available from 62 UTI patients with positive dipstick test before antibiotic treatment. Six of these patients also provided urine after completion of antibiotic treatment. Subjects in cohorts of OAB (n = 59) and asymptomatic controls (n = 26) were negative for dipstick test. Urinary chemokines were measured by MILLIPLEX MAP Human Cytokine/Chemokine Immunoassay and their association with UTI and OAB was determined by univariate and multivariate statistics. Significant elevation of CXCL-1, CXCL-8 (IL-8), and CXCL-10 together with reduced levels for a receptor antagonist of IL-1A (sIL-1RA) were seen in UTI relative to OAB and asymptomatic controls. Elevated CXCL-1 urine levels predicted UTI with odds ratio of 1.018 and showed a specificity of 80.77% and sensitivity of 59.68%. Postantibiotic treatment, reduction was seen in all CXC chemokines with a significant reduction for CXCL-10. Strong association of CXCL-1 and CXCL-10 for UTI over OAB indicates mechanistic differences in signaling pathways driving inflammation secondary of infection in UTI compared with a lack of infection in OAB. Urinary chemokines highlight molecular differences in the paracrine signaling driving the overlapping symptoms of UTI and OAB.

Myoglobin-Catalyzed Olefination of Aldehydes.

The olefination of aldehydes constitutes a most valuable and widely adopted strategy for constructing carbon-carbon double bonds in organic chemistry. While various synthetic methods have been made available for this purpose, no biocatalysts are known to mediate this transformation. Reported herein is that engineered myoglobin variants can catalyze the olefination of aldehydes in the presence of α-diazoesters with high catalytic efficiency (up to 4,900 turnovers) and excellent E diastereoselectivity (92-99.9 % de). This transformation could be applied to the olefination of a variety of substituted benzaldehydes and heteroaromatic aldehydes, also in combination with different alkyl α-diazoacetate reagents. This work provides a first example of biocatalytic aldehyde olefination and extends the spectrum of synthetically valuable chemical transformations accessible using metalloprotein-based catalysts.

Intermolecular carbene S-H insertion catalysed by engineered myoglobin-based catalysts†.

The first example of a biocatalytic strategy for the synthesis of thioethers via an intermolecular carbene S-H insertion reaction is reported. Engineered variants of sperm whale myoglobin were found to efficiently catalyze this C-S bond forming transformation across a diverse set of aryl and alkyl mercaptan substrates and α-diazoester carbene donors, providing high conversions (60-99%) and high numbers of catalytic turnovers (1,100-5,400). Furthermore, the enantioselectivity of these biocatalysts could be tuned through mutation of amino acid residues within the distal pocket of the hemoprotein, leading to myoglobin variants capable of supporting asymmetric S-H insertions with up to 49% ee. Rearrangement experiments support a mechanism involving the formation of a sulfonium ylide generated upon attack of the thiol substrate to a heme-bound carbene intermediate.

Highly diastereoselective and enantioselective olefin cyclopropanation using engineered myoglobin-based catalysts.

Using rational design, an engineered myoglobin-based catalyst capable of catalyzing the cyclopropanation of aryl-substituted olefins with catalytic proficiency (up to 46,800 turnovers) and excellent diastereo- and enantioselectivity (98-99.9%) was developed. This transformation could be carried out in the presence of up to 20 g L(-1) olefin substrate with no loss in diastereo- and/or enantioselectivity. Mutagenesis and mechanistic studies support a cyclopropanation mechanism mediated by an electrophilic, heme-bound carbene species and a model is provided to rationalize the stereopreference of the protein catalyst. This work shows that myoglobin constitutes a promising and robust scaffold for the development of biocatalysts with carbene-transfer reactivity.

Association of inflammaging (inflammation + aging) with higher prevalence of OAB in elderly population.

INTRODUCTION: Although epidemiology studies consistently report increased prevalence of overactive bladder (OAB) with age, an accurate deciphering of causative links between the two entities remains elusive. Studies on aged rodent bladder have so far yielded contradictory results on age-associated changes in muscarinic receptors, which highlight the challenge posed by species differences in understanding OAB pathology. We hypothesized that age-related biochemical changes in bladder leading to altered bladder function will be reflected in altered urinary proteome of elderly OAB patients. METHODS: Single time point urine specimens were obtained from 140 OAB patients in the age range of 25-90 years of either sex coming routinely to the urology clinics. Eight chemokines in urine were measured by MILLIPLEX MAP human cytokine/chemokine multiplex immunoassay and ELISA. Multivariate and univariate statistical analyses were done to determine association of age with urinary chemokines in OAB patients. RESULTS: In agreement with age-dependent higher prevalence of OAB, the logistic regression of the data also revealed the significant association of OAB symptoms with age [odds ratio (OR) 1.12; 95 % CI, (1.072, 1.187), p = 0.0001]. Univariate analysis of 8 urinary proteins revealed an age-associated elevation of NGF (nerve growth factor) in 137 out of 140 OAB patients [Pearson r = 0.274; 95 %CI (0.112-0.422); p = 0.001]. Modest correlation with age was also noted for MCP-1 (monocyte chemoattractant protein-1), which was detected in 115 OAB patients, and the remaining chemokines were undetectable in nearly two-third of OAB patients included in our cohort. CONCLUSIONS: Based on our findings, we postulate that age-associated biochemical changes may accentuate the inflammation associated with OAB. Urinary NGF elevation in elderly OAB patients may be a homeostatic response to counter the senescence of bladder nerves and arrest the progression of OAB into detrusor hyperactivity with impaired contractility. Likewise, elevation of MCP-1 may be related to decreased muscle mass and increased content of adipose tissue in bladder of elderly OAB patients. Urinary NGF and MCP-1 can serve as surrogate markers for monitoring age-associated biochemical changes and the effect of therapeutic interventions in OAB patients.