Immunoglobulin A Nephropathy Presenting in Young Adults as Advanced Chronic Kidney Disease and Posttransplant Early Recurrence: A Case Series.

Immunoglobulin A nephropathy (IgAN) is the most common glomerular disease, leading to chronic kidney disease. The disease is characterized by microscopic hematuria, gross episodic hematuria, hypertension, and subnephrotic proteinuria with or without renal function impairment. It affects individuals of all age groups, commonly seen in 10-40 years of age. It is progressive in nature and leads to chronic kidney disease, necessitating renal replacement therapy. This case series of in a tertiary care hospital in Western India highlights the presentation of this disease in young adults, its aggressive course, its rapid progression, and its early recurrence in the posttransplant period. It also summarizes the treatment recommendations for IgA recurrence in kidney recipients. The disease is known to have a high chance of posttransplant recurrence. Optimizing renin-angiotensin-aldosterone system (RAAS) blockade, blood pressure control, and increasing immunosuppression in rapidly deteriorating cases are the strategies recommended to treat IgA recurrence in kidney transplant recipients.

Molecular basis of conformational changes and mechanics of integrins.

Integrin, as a mechanotransducer, establishes the mechanical reciprocity between the extracellular matrix (ECM) and cells at integrin-mediated adhesion sites. This study used steered molecular dynamics (SMD) simulations to investigate the mechanical responses of integrin αvß3 with and without 10th type III fibronectin (FnIII10) binding for tensile, bending and torsional loading conditions. The ligand-binding integrin confirmed the integrin activation during equilibration and altered the integrin dynamics by changing the interface interaction between ß-tail, hybrid and epidermal growth factor domains during initial tensile loading. The tensile deformation in integrin molecules indicated that fibronectin ligand binding modulates its mechanical responses in the folded and unfolded conformation states. The bending deformation responses of extended integrin models reveal the change in behaviour of integrin molecules in the presence of Mn2+ ion and ligand based on the application of force in the folding and unfolding directions of integrin. Furthermore, these SMD simulation results were used to predict the mechanical properties of integrin underlying the mechanism of integrin-based adhesion. The evaluation of integrin mechanics provides new insights into understanding the mechanotransmission (force transmission) between cells and ECM and contributes to developing an accurate model for integrin-mediated adhesion. This article is part of a discussion meeting issue 'Supercomputing simulations of advanced materials'.

A Coarse-Grained Model for the Mechanical Behavior of Na-Montmorillonite Clay.

Sodium montmorillonite (Na-MMT) is one of the most commonly found swelling clay minerals with diverse engineering and technological applications. The nanomechanical properties of this mineral have been extensively investigated computationally utilizing molecular dynamics (MD) simulations to portray the molecular-level changes at different environmental conditions. As the environmentally found Na-MMT clays are generally sized within hundreds of nanometers, all-atomistic (AA) MD simulations of clays within such size range are particularly challenging due to computational inefficiency. Informed from atomistic modeling, a coarse-grained (CG) modeling technique can be employed to overcome the spatiotemporal limitation. The current study presents a modeling strategy to develop a computationally efficient model of Na-MMT clay with a typical size over ≃100 nm by shrinking the atomistic platelet thickness and reducing the number of center-layer atoms. Using the "strain-energy conservation" approach, the force field parameters for the CG model are obtained and the developed CG model can well preserve in-plane tension, shear, and bending behaviors of atomistic counterparts. Remarkably, the CG tactoid model of Na-MMT, a hierarchical multilayer structure, can reproduce the interlayer shear and adhesion as well as d-spacing among the clay sheets as of atomistic one to a good approximation while gaining significantly improved computational speed. Our study demonstrates the efficacy of the CG modeling framework, paving the way for the bottom-up multiscale prediction of mechanical behaviors of clay and related minerals.

Identification and characterization of a hydrophobin Vmh3 from Pleurotus ostreatus.

Hydrophobins (HPs) are relatively small surface-active proteins of fungal origin. Being an industrially important protein, isolation of new molecules from GRAS (Generally Regarded as Safe) strains like mushrooms is the need of the time. In the present work, hydrophobin Vmh3-1 is isolated, purified, and identified from a culture broth and vegetative mycelia of Pleurotus ostreatus grown in a Potato dextrose broth (PDB) in static culture conditions. Purified proteins from the broth and the cell wall showed bands of 11 kDa and 17 kDa when analyzed on SDS-PAGE. Hydrophobin Vmh3-1 was identified in purified protein samples by the Orbitrap-HR-LC-MS/MS analysis with a maximum of 66% sequence coverage. The amphipathic nature of the protein was revealed by an increase in the water contact angle (WCA) of the hydrophilic surface of glass by 87% as well as a decrease in the WCA of the hydrophobic surface of Teflon by 19%. The emulsification property was tested with food-grade oils and Hexane. A maximum activity (EI 24) of 87.64% was recorded for Sunflower oil. In CD (Circular dichroism) spectra, Vmh3-1 showed the typical spectra of hydrophobin with a dominance of ß-sheets (51%) in the secondary structure and a minimum percentage of the α-helix (2%). The protein did not show a self-aggregating property on vigorous shaking making it suitable for numerous industrial applications. The identification of Vmh3-1 with detailed amino acid sequencing and the characterization of the protein to evaluate its potential in surface modifications for various industrial applications is demonstrated herein for the first time.

Dissociation Mechanisms of G-actin Subunits Govern Deformation Response of Actin Filament.

Actin molecules are essential structural components of the cellular cytoskeleton. Here, we report a comprehensive analysis of F-actin's deformation behavior and highlight underlying mechanisms using steered molecular dynamics simulations (SMD). The investigation of F-actin was done under tension, compression, bending, and torsion. We report that the dissociation pattern of conformational locks at intrastrand and interstrand G-actin interfaces regulates the deformation response of F-actin. The conformational locks at the G-actin interfaces are portrayed by a spheroidal joint, interlocking serrated plates' analogy. Further, the SMD simulation approach was utilized to evaluate Young's modulus, flexural rigidity, persistent length, and torsional rigidity of F-actin, and the values obtained were found to be consistent with available experimental data. The evaluation of the mechanical properties of actin and the insight into the fundamental mechanisms contributing to its resilience described here are necessary for developing accurate models of eukaryotic cells and for assessing cellular viability and mobility.

Binding of SARS-COV-2 (COVID-19) and SARS-COV to human ACE2: Identifying binding sites and consequences on ACE2 stiffness.

The SARS-CoV-2 coronavirus (COVID-19) that is causing the massive global pandemic exhibits similar human cell invasion mechanism as the coronavirus SARS-CoV, which had significantly lower fatalities. The cell membrane protein Angiotensin-converting enzyme 2 (ACE2) is the initiation point for both the coronavirus infections in humans. Here, we model the molecular interactions and mechanical properties of ACE2 with both SARS-CoV and COVID-19 spike protein receptor-binding domains (RBD). We report that the COVID-19 spike RBD interacts with ACE2 more strongly and at only two protein residues, as compared to multi-residue interaction of the SARS-CoV. Although both coronaviruses stiffen the ACE2, the impact of COVID-19 is six times larger, which points towards differences in the severity of the reported respiratory distress. The recognition of specific residues of ACE2 attachments to coronaviruses is important as the residues suggest potential sites of intervention to inhibit attachment and subsequent entry of the COVID-19 into human host cells.

Differences in Interactions Within Viral Replication Complexes of SARS-CoV-2 (COVID-19) and SARS-CoV Coronaviruses Control RNA Replication Ability.

COVID-19 has become a global pandemic caused by the SARS-CoV-2 coronavirus. SARS-CoV-2 shares many similarities with SARS coronavirus (SARS-CoV). A viral replication complex containing non-structural proteins (nsps) is the toolbox for RNA replication and transcription of both coronaviruses. In both cases, the RNA-dependent RNA polymerase (RdRp) domain of the coronaviral replication complex dictates the primary polymerase activity by cooperating with cofactors. The higher transmissibility and mortality due to SARS-CoV-2 are related to its higher RNA replication activity compared to SARS-CoV. The discrepancy between the RNA replication efficiency of SARS-CoV and SARS-CoV-2 can be understood by exploring interactions within their viral replication complexes. Our modeling of molecular interactions within the viral replication complexes of SARS-CoV and SARS-CoV-2 using molecular dynamics simulations suggests that in contrast to SARS-CoVnsp12, SARS-CoV2nsp12 prefers helices as the dominant interacting secondary motifs. The relative differences in nonbonded interactions between nsps could suggest viral RNA replication ability in coronaviruses. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s11837-021-04662-6.

Compression of Na-Montmorillonite Swelling Clay Interlayer Is Influenced by Fluid Polarity: A Steered Molecular Dynamics Study.

The compressive responses of an interlayer of the dry sodium-montmorillonite (Na-MMT) swelling clay as well as the clay intercalated with organic fluids of a wide range of dielectric constants from 110 (formamide) to 20 (acetone) are quantitatively evaluated using steered molecular dynamics simulations. Representative dry clay and clay with fluid (clay-fluid) molecular models are constructed, and the stress-strain relationships upon compression of these models are studied using constant force steered molecular dynamics (SMD) simulations. Our results show that the polarity of the fluids and the amount of the fluid molecules in the clay interlayer play a significant role in the interlayer spacing, interlayer volume, interlayer strain, interlayer modulus, nonbonded interactions, and conformation of the fluid molecules upon externally applied stresses. The clay interlayer responses upon compression are essential for the development of multiscale modeling of swelling clays and prediction of the reliable compressive behavior, which are critical for the accurate analysis and economical design of the infrastructures in swelling clay areas and the densification of clays for ceramics manufacturing.

Electrical behavior and enhanced photocatalytic activity of (Ag, Ni) co-doped ZnO nanoparticles synthesized from co-precipitation technique.

Methylene blue (MB) dye is the most common harmful, toxic, and non-biodegradable effluent produced by the textile industries. The present study investigates the effect of zinc oxide (ZnO) nanoparticles (NPs) and Ag-Ni doped ZnO NPs on the performance of photocatalytic degradation of MB dye. Pure ZnO and Ag-Ni doped ZnO NPs are synthesized using the co-precipitation method. The crystalline nature and surface morphology of the synthesized pure ZnO and Ag-Ni doped ZnO NPs was characterized by powder X-ray diffraction, scanning electron microscopy (SEM), and high resolution transmission electron microscopy (HRTEM) analysis. The presence of spherical-like morphologies was confirmed from SEM and HRTEM analysis. The presence of Ni-O and Zn-O bands in the synthesized materials was found by Fourier transform infrared (FTIR) spectroscopy analysis. The MB dye was degraded under UV-light exposure in various pH conditions. The Ag (0.02%)-Ni doped ZnO NPs exhibits highest photocatalytic activity of 77% under pH 4.

Positive Selection on a Regulatory Insertion-Deletion Polymorphism in FADS2 Influences Apparent Endogenous Synthesis of Arachidonic Acid.

Long chain polyunsaturated fatty acids (LCPUFA) are bioactive components of membrane phospholipids and serve as substrates for signaling molecules. LCPUFA can be obtained directly from animal foods or synthesized endogenously from 18 carbon precursors via the FADS2 coded enzyme. Vegans rely almost exclusively on endogenous synthesis to generate LCPUFA and we hypothesized that an adaptive genetic polymorphism would confer advantage. The rs66698963 polymorphism, a 22-bp insertion-deletion within FADS2, is associated with basal FADS1 expression, and coordinated induction of FADS1 and FADS2 in vitro. Here, we determined rs66698963 genotype frequencies from 234 individuals of a primarily vegetarian Indian population and 311 individuals from the US. A much higher I/I genotype frequency was found in Indians (68%) than in the US (18%). Analysis using 1000 Genomes Project data confirmed our observation, revealing a global I/I genotype of 70% in South Asians, 53% in Africans, 29% in East Asians, and 17% in Europeans. Tests based on population divergence, site frequency spectrum, and long-range haplotype consistently point to positive selection encompassing rs66698963 in South Asian, African, and some East Asian populations. Basal plasma phospholipid arachidonic acid (ARA) status was 8% greater in I/I compared with D/D individuals. The biochemical pathway product-precursor difference, ARA minus linoleic acid, was 31% and 13% greater for I/I and I/D compared with D/D, respectively. This study is consistent with previous in vitro data suggesting that the insertion allele enhances n-6 LCPUFA synthesis and may confer an adaptive advantage in South Asians because of the traditional plant-based diet practice.

Nuclear Localizing Peptide-Conjugated, Redox-Sensitive Polymersomes for Delivering Curcumin and Doxorubicin to Pancreatic Cancer Microtumors.

Improving the therapeutic index of anticancer agents is an enormous challenge. Targeting decreases the side effects of the therapeutic agents by delivering the drugs to the intended destination. Nanocarriers containing the nuclear localizing peptide sequences (NLS) translocate to the cell nuclei. However, the nuclear localization peptides are nonselective and cannot distinguish the malignant cells from the healthy counterparts. In this study, we designed a "masked" NLS peptide which is activated only in the presence of overexpressed matrix metalloproteinase-7 (MMP-7) enzyme in the pancreatic cancer microenvironment. This peptide is conjugated to the surface of redox responsive polymersomes to deliver doxorubicin and curcumin to the pancreatic cancer cell nucleus. We have tested the formulation in both two- and three-dimensional cultures of pancreatic cancer and normal cells. Our studies revealed that the drug-encapsulated polymeric vesicles are significantly more toxic toward the cancer cells (shrinking the spheroids up to 49%) compared to the normal cells (shrinking the spheroids up to 24%). This study can lead to the development of other organelle targeted drug delivery systems for various human malignancies.

A Prospective Study of Acute Kidney Injury According to KDIGO Definition and its Mortality Predictors.

BACKGROUND: Acute kidney injury is no longer considered to be an innocent bystander merely reflecting co-existent pathologies but an independent risk factor for mortality in the ICU. AIMS AND OBJECTIVES: To study clinical profile and correlation of patients with acute kidney injury (AKI) according to KDIGO definition with respect to incidence, outcome and different causes of AKI in critical care unit. STUDY DESIGN AND SETTING: It is a prospective observational study; and was carried out in the ICU of a tertiary care, teaching, public hospital. MATERIAL AND METHODS: We studied 316 patients developing AKI in ICU over a period of 1 year. RESULTS AND CONCLUSION: Incidence of AKI in our ICU was 37.71% and mortality rate was 51.9%. Tropical Acute febrile illnesses followed by sepsis were the most common causes of AKI in ICU. Most common cause of AKI among tropical acute febrile illnesses (AFI) was malaria and among sepsis group was lung infection. In our study KDIGO staging could not predict outcome because majority of patients had multisystem failure. Pre-existing co-morbidities, multi-organ system failure were associated with high mortality. APACHE II scoring system under- predicted the mortality in patients with AKI.

The roles of cellular nanomechanics in cancer.

The biomechanical properties of cells and tissues may be instrumental in increasing our understanding of cellular behavior and cellular manifestations of diseases such as cancer. Nanomechanical properties can offer clinical translation of therapies beyond what are currently employed. Nanomechanical properties, often measured by nanoindentation methods using atomic force microscopy, may identify morphological variations, cellular binding forces, and surface adhesion behaviors that efficiently differentiate normal cells and cancer cells. The aim of this review is to examine current research involving the general use of atomic force microscopy/nanoindentation in measuring cellular nanomechanics; various factors and instrumental conditions that influence the nanomechanical properties of cells; and implementation of nanoindentation methods to distinguish cancer cells from normal cells or tissues. Applying these fundamental nanomechanical properties to current discoveries in clinical treatment may result in greater efficiency in diagnosis, treatment, and prevention of cancer, which ultimately can change the lives of patients.

DNA methylation analysis of phenotype specific stratified Indian population.

BACKGROUND: DNA methylation and its perturbations are an established attribute to a wide spectrum of phenotypic variations and disease conditions. Indian traditional system practices personalized medicine through indigenous concept of distinctly descriptive physiological, psychological and anatomical features known as prakriti. Here we attempted to establish DNA methylation differences in these three prakriti phenotypes. METHODS: Following structured and objective measurement of 3416 subjects, whole blood DNA of 147 healthy male individuals belonging to defined prakriti (Vata, Pitta and Kapha) between the age group of 20-30years were subjected to methylated DNA immunoprecipitation (MeDIP) and microarray analysis. After data analysis, prakriti specific signatures were validated through bisulfite DNA sequencing. RESULTS: Differentially methylated regions in CpG islands and shores were significantly enriched in promoters/UTRs and gene body regions. Phenotypes characterized by higher metabolism (Pitta prakriti) in individuals showed distinct promoter (34) and gene body methylation (204), followed by Vata prakriti which correlates to motion showed DNA methylation in 52 promoters and 139 CpG islands and finally individuals with structural attributes (Kapha prakriti) with 23 and 19 promoters and CpG islands respectively. Bisulfite DNA sequencing of prakriti specific multiple CpG sites in promoters and 5'-UTR such as; LHX1 (Vata prakriti), SOX11 (Pitta prakriti) and CDH22 (Kapha prakriti) were validated. Kapha prakriti specific CDH22 5'-UTR CpG methylation was also found to be associated with higher body mass index (BMI). CONCLUSION: Differential DNA methylation signatures in three distinct prakriti phenotypes demonstrate the epigenetic basis of Indian traditional human classification which may have relevance to personalized medicine.

pH-triggered echogenicity and contents release from liposomes.

Liposomes are representative lipid nanoparticles widely used for delivering anticancer drugs, DNA fragments, or siRNA to cancer cells. Upon targeting, various internal and external triggers have been used to increase the rate for contents release from the liposomes. Among the internal triggers, decreased pH within the cellular lysosomes has been successfully used to enhance the rate for releasing contents. However, imparting pH sensitivity to liposomes requires the synthesis of specialized lipids with structures that are substantially modified at a reduced pH. Herein, we report an alternative strategy to render liposomes pH sensitive by encapsulating a precursor which generates gas bubbles in situ in response to acidic pH. The disturbance created by the escaping gas bubbles leads to the rapid release of the encapsulated contents from the liposomes. Atomic force microscopic studies indicate that the liposomal structure is destroyed at a reduced pH. The gas bubbles also render the liposomes echogenic, allowing ultrasound imaging. To demonstrate the applicability of this strategy, we have successfully targeted doxorubicin-encapsulated liposomes to the pancreatic ductal carcinoma cells that overexpress the folate receptor on the surface. In response to the decreased pH in the lysosomes, the encapsulated anticancer drug is efficiently released. Contents released from these liposomes are further enhanced by the application of continuous wave ultrasound (1 MHz), resulting in substantially reduced viability for the pancreatic cancer cells (14%).

Mmp-9 responsive PEG cleavable nanovesicles for efficient delivery of chemotherapeutics to pancreatic cancer.

Significant differences in biochemical parameters between normal and tumor tissues offer an opportunity to chemically design drug carriers which respond to these changes and deliver the drugs at the desired site. For example, overexpression of the matrix metalloproteinase-9 (MMP-9) enzyme in the extracellular matrix of tumor tissues can act as a trigger to chemically modulate the drug delivery from the carriers. In this study, we have synthesized an MMP-9-cleavable, collagen mimetic lipopeptide which forms nanosized vesicles with the POPC, POPE-SS-PEG, and cholesteryl-hemisuccinate lipids. The lipopeptide retains the triple-helical conformation when incorporated into these nanovesicles. The PEG groups shield the substrate lipopeptides from hydrolysis by MMP-9. However, in the presence of elevated glutathione levels, the PEG groups are reductively removed, exposing the lipopeptides to MMP-9. The resultant peptide-bond cleavage disturbs the vesicles' lipid bilayer, leading to the release of encapsulated contents. These PEGylated nanovesicles are capable of encapsulating the anticancer drug gemcitabine with 50% efficiency. They were stable in physiological conditions and in human serum. Effective drug release was demonstrated using the pancreatic ductal carcinoma cells (PANC-1 and MIAPaCa-2) in two-dimensional and three-dimensional "tumor-like" spheroid cultures. A reduction in tumor growth was observed after intravenous administration of the gemcitabine-encapsulated nanovesicles in the xenograft model of athymic, female nude mice.

Anaesthetic consideration during laparoscopic bilateral simultaneous nephrectomy.

STUDY OBJECTIVE: To assess outcome from anaesthesia during laparoscopic bilateral simultaneous nephrectomy. DESIGN: Retrospective study. MEASUREMENTS: Preoperative Hb%, serum potassium, coagulation profile electrocardiography (ECG) changes, 2D Echography, x-ray chest, haemodynamic changes, end-tidal carbon dioxide (EtCO2), fluid management and postoperative analgesia. RESULTS: The mean age was 24.75 ± 14.35 years. The mean duration of surgery was 120 ± 80 minutes. The Hb%, serum creatinine and serum potassium were 9.4 ± 1.04%, 6.79 ± 4.91 meq/L and 3.61 ± 0.51 meq/L, respectively. Pulse rate mean blood pressure and EtCO2 were recorded after creation of pneumoperitoneum and at 15, 30, 45 and after exsufflation of pneumoperitoneum. After pneumoperitoneum, there was increase in pulse rate, systolic blood pressure, diastolic blood pressure and EtCO2. After 30 minutes and throughout the surgery, these variables remained stable. Four patients required nitroglycerine infusion for intraoperative hypertention. Only one patient required packed cell volume (PCV) transfusion and total intravenous fluid was 1 ± 0.5 L. At the time of exsufflation, there was decrease in pulse rate, systolic and diastolic blood pressure and EtCO2. CONCLUSION: Because of advancement in anaesthetic agents and muscle relaxant, there is safe outcome from anaesthesia during laparoscopic bilateral simultaneous nephrectomy.

Mechanoresponsive ETS1 causes endothelial dysfunction and arterialization in varicose veins via NOTCH4/DLL4 signaling.

Varicose veins are the most common venous disorder in humans and are characterized by hemodynamic instability due to valvular insufficiency and orthostatic lifestyle factors. It is unclear how changes in biomechanical signals cause aberrant remodeling of the vein wall. Our previous studies suggest that Notch signaling is implicated in varicose vein arterialization. In the arterial system, mechanoresponsive ETS1 is a transcriptional activator of the endothelial Notch, but its involvement in sensing disrupted venous flow and varicose vein formation has not been investigated. Here, we use human varicose veins and cultured human venous endothelial cells to show that disturbed venous shear stress activates ETS1-NOTCH4/DLL4 signaling. Notch components were highly expressed in the neointima, whereas ETS1 was upregulated in all histological layers of varicose veins. In vitro microfluidic flow-based studies demonstrate that even minute changes in venous flow patterns enhance ETS1-NOTCH4/DLL4 signaling. Uniform venous shear stress, albeit an inherently low-flow system, does not induce ETS1 and Notch proteins. ETS1 activation under altered flow was mediated primarily by MEK1/2 and, to a lesser extent, by MEK5 but was independent of p38 MAP kinase. Endothelial cell-specific ETS1 knockdown prevented disturbed flow-induced NOTCH4/DLL4 expression. TK216, an inhibitor of ETS-family, prevented the acquisition of arterial molecular identity and loss of endothelial integrity in cells exposed to the ensuing altered shear stress. We conclude that ETS1 senses blood flow disturbances and may promote venous remodeling by inducing endothelial dysfunction. Targeting ETS1 rather than downstream Notch proteins could be an effective and safe strategy to develop varicose vein therapies.

Molecular mechanisms of anti-tumor properties of P276-00 in head and neck squamous cell carcinoma.

BACKGROUND: Tumors of the head and neck present aggressive pathological behavior in patients due to high expression of CDK/CCND1 proteins. P276-00, a novel CDK inhibitor currently being tested in clinic, inhibits growth of several cancers in vitro and in vivo. The pre clinical activity of P276-00 in head and neck cancer and its potential mechanisms of action at molecular level are the focus of the current studies. METHOD: We have investigated the anti-cancer activity of P276-00 in head and neck tumors in vitro and in vivo. Candidate gene expression profiling and cell based proteomic approaches were taken to understand the pathways affected by P276-00 treatment. RESULTS: It was observed that P276-00 is cytotoxic across various HNSCC cell lines with an IC50 ranging from 1.0-1.5 µmoles/L and culminated in significant cell-cycle arrest in G1/S phase followed by apoptosis. P276-00 treatment suppressed cell proliferation through inhibition of CCND1 expression, reduced phosphorylation of retinoblastoma protein and abrogative transcription of E2F1 gene targets. Further, we observed that apoptosis was mediated through P53 activation leading to higher BAX/BCL-2 ratio and cleaved caspase-3 levels. It was also seen that P276-00 treatment reduced expression of tumor micro-environment proteins such as IL-6, secreted EGFR and HSPA8. Finally, P276-00 treatment resulted in significant tumor growth inhibition in xenograft tumor models via lowered proliferative activity of E2F1 and aggravated P53 mediated apoptosis. CONCLUSION: In summary, we have observed that P276-00 inhibits cyclin-D/CDK4/P16/pRB/E2F axis and induces apoptosis by increased P53 phosphorylation in HNSCC cells. These results suggest a novel indication for P276-00 in head and neck cancer with a potential role for IL-6 and HSPA8 as candidate serum biomarkers.

Potentiation of in vitro and in vivo antitumor efficacy of doxorubicin by cyclin-dependent kinase inhibitor P276-00 in human non-small cell lung cancer cells.

BACKGROUND: In the present study, we show that the combination of doxorubicin with the cyclin-dependent kinase inhibitor P276-00 was synergistic at suboptimal doses in the non-small cell lung carcinoma (NSCLC) cell lines and induces extensive apoptosis than either drug alone in H-460 human NSCLC cells. METHODS: Synergistic effects of P276-00 and doxorubicin on growth inhibition was studied using the Propidium Iodide (PI) assay. The doses showing the best synergistic effect was determined and these doses were used for further mechanistic studies such as western blotting, cell cycle analysis and RT-PCR. The in vivo efficacy of the combination was evaluated using the H-460 xenograft model. RESULTS: The combination of 100 nM doxorubicin followed by 1200 nM P276-00 showed synergistic effect in the p53-positive and p53-mutated cell lines H-460 and H23 respectively as compared to the p53-null cell line H1299. Abrogation of doxorubicin-induced G2/M arrest and induction of apoptosis was observed in the combination treatment. This was associated with induction of tumor suppressor protein p53 and reduction of anti-apoptotic protein Bcl-2. Furthermore, doxorubicin alone greatly induced COX-2, a NF-κB target and Cdk-1, a target of P276-00, which was downregulated by P276-00 in the combination. Doxorubicin when combined with P276-00 in a sequence-specific manner significantly inhibited tumor growth, compared with either doxorubicin or P276-00 alone in H-460 xenograft model. CONCLUSION: These findings suggest that this combination may increase the therapeutic index over doxorubicin alone and reduce systemic toxicity of doxorubicin most likely via an inhibition of doxorubicin-induced chemoresistance involving NF-κB signaling and inhibition of Cdk-1 which is involved in cell cycle progression.