Trachyspermumammiis used traditionally to manage respiratory problems. The essential oil of the fruits was analyzed using GC-MS and the bronchodilator potential was explored using isolated guinea-pig trachea in an ex-vivo model. The GC-MS analysis revealed that thymol is major compound with 65% yield. T. ammi oil exhibited comparable potency in inhibiting contractions caused by carbachol (CCh, 1 mu M) and high K` (80 mM), with corresponding EC50 = 0.28 mg/mL (0.26-0.31, n=4) and 0.32 mg/mL (0.29-0.35, n=4) in a fashion similar to papaverine known to inhibit both Ca`2 channels and the phosphodiesterase enzyme (PDE). In contrast, Verapamil, a selective Ca`2 channel blocker, suppressed contractions induced by elevated K` levelswith significantly higher potency than those induced by CCh, as indicated by the EC50 = 0.82 mu M (0.68-1.02, n=5) and 17.84 mu M (15.64-1.86, n=5), respectively. Tissues preincubated with 0.03 and 0.1 mg/mL of the oil were able to attenuate the Ca`2 produced contractions and suppressing the maximum response in a manner comparable to verapamil and papaverine further supporting the Ca`2 channel inhibitory effect. PDE blockage was settled when tracheal tissues preincubated with T. ammi essential oil at 0.1 and 0.3 mg/mL expressed potentiation of isoprenaline relaxant effect against CCh, similar to papaverine. Preincubation with verapamil did not show any potentiation. Therefore, our results demonstrated that T. ammiessential oil have bronchodilator activities mediated by dual inhibition of Ca`2 channels and PDE, although other mechanisms may also be involved.

Introduction: Androgenetic alopecia (AGA) is a multifactorial and age-related dermatological disease that affects both males and females, usually at older ages. Traditional hair repair drugs exemplified by minoxidil have limitations such as skin irritation and hypertrichosis. Thus, attention has been shifted to the use of repurposing drugs. Metformin is an anti-diabetic drug, that can promote hair follicle regeneration via upregulation of the hair-inductive capability. Hence, the current study aims to fabricate a safe and effective nanoemulsion to improve metformin efficacy in targeting AGA. Methods: Rosemary oil was selected as the oily phase due to its ability to increase blood flow and hair growth. Rosemary-based nanoemulsions were statistically optimized by Box-Behnken experimental design, loaded with metformin, and incorporated into a hydrogel to form a nanoemulgel. Metformin-loaded nanoemulsions were assessed for their diametric size, uniformity, zeta potential, and metformin characteristics within the formulated nanosystem. The nanoemulgel was then evaluated in terms of its pH, percentage drug content, and in-vitro release performance. In-vivo study assessed the nanoemulgel's ability to augment hair growth in rats. Results: The experimental design displayed that using 50%w/w, 20%w/w, and 10%w/w of Cremophor (R), Labrafil (R), and deionized water, respectively, resulted in nanoemulsion formulation with the smallest globule size (125.01 +/- 0.534 nm), unimodal size distribution (PDI=0.103), negative surface charge (-19.9 +/- 2.01 mV) with a spherical morphological structure. Rosemary-based nanoemulgel displayed acceptable physicochemical characterizations namely; a neutral pH value of 6.7 +/- 0.15, high drug content (92.9 +/- 2.3%), and controlled metformin in-vitro release. Besides, the formulated nanoemulgel significantly increased the number of hair follicles in the animal model compared with other controls and tested groups. Conclusion: The designed nanoemulgel is a promising approach for treating androgenic alopecia.

Acetyl-L-carnitine (ALC) is known for its potent antioxidant and anti-inflammatory properties. This research aimed to investigate the properties of ALC in combating pelvic inflammatory disease (PID) in a rat model. PID, a consequence of sexually transmitted infections in females, can impact the fallopian tubes and uterus, leading to complications. This study is a randomized control preclinical experiment. A total of 24 reproductively mature Sprague Dawley female rats were randomly and equally assigned (n = 4 per group) to six cohorts: control, PID, low-dose prophylactic, high-dose prophylactic, and low-dose therapeutic and high-dose therapeutic groups. PID was induced by injecting the rat cervix with a multi-pathogen solution. Acetyl-L-carnitine, 100 mg/kg (once a day), was orally administered to rats in the low-dose prophylactic group, while 200 mg/kg (once a day) to the high-dose prophylactic group, starting 1 day prior to induction of PID. The therapeutic groups were given similar doses of ALC 1 day after the PID model was confirmed. Samples from the right upper genital tract were collected for ELISA and antioxidant assays, while the left upper genital tract samples underwent histopathological analysis. According to the results, the ALC-treated groups showed a decreased level of cytokines (IL-1β, TNF-α) and oxidative stress markers (catalase, lipid peroxidation) when compared to the PID group. Histopathological examinations revealed that ALC treatment reduced the infiltration of neutrophils and lymphocytes in the uterus compared to the PID group. It was concluded that ALC showed potential antioxidant and anti-inflammatory effects in PID and, therefore, could be used as a possible option for the treatment of PID.

This study was undertaken to assess the antioxidant and neuropharmacological potentials of the methanol leaf extract of Acanthus ebracteatus (MAEL) through experimental and in silico methods. The phytochemical screening (PS) and GC-MS (gas chromatography-mass spectrometry) identified 28 phytochemicals with different classes in nature in MAEL. The MAEL revealed better antioxidant activity through various in vitro antioxidant assays. Additionally, in the tail suspension test (TST) and forced swimming test (FST), a dose-dependent reduction in immobility time was observed indicating antidepressant activity. In the elevated plus maze test (EPM), MAEL led to increased time spent and more entries in the open arms. At the same time, the hole board test (HBT) demonstrated an increase in head dipping compared to the control, both indicating anxiolytic activity. Moreover, a dose-dependent reduction in locomotor activities was observed in both the open field test (OFT) and hole cross test (HCT). Molecular docking showed better binding affinities of two compounds, CID-518982 and CID-236641. ADME/T analysis revealed good drug likeliness with no toxicity. Finally, the simulation demonstrated better structural stability with no significant fluctuations of the compounds with the selected receptors. In this study, compounds CID-518982 and CID-236641 might serve as drug candidates for treating anxiety and depression.

With a global towering prevalence of index acute myocardial infarction (nonrecurrent MI, NR-MI), a high incidence of recurrent MI (R-MI) has emerged in recent decades. Despite the extensive occurrence, the promising predictors of R-MI have been elusive within the cohort of survivors. This study investigates and validates the involvement of distinct gene expressions in R-MI and NR-MI. Bioinformatics tools were used to identify DEGs from the GEO dataset, functional annotation, pathway enrichment analysis, and the PPI network analysis to find hub genes. The validation of proposed genes was conceded by qRT-PCR and Western Blot analysis in experimentally induced NR-MI and R-MI models on a temporal basis. The temporal findings based on RT-PCR consequences reveal a significant and constant upregulation of the UBE2N in the NR-MI model out of the proposed three DEGs (UBE2N, UBB, and TMEM189), while no expression was reported in the R-MI model. Additionally, the proteomics study proposed five DEGs (IL2RB, NKG7, GZMH, CXCR6, and GZMK) for the R-MI model since IL2RB was spotted for significant and persistent downregulation with different time points. Further, Western Blot analysis validated these target genes' expressions temporally. I/R-induced NR-MI and R-MI models were confirmed by the biochemical parameters (CKMB, LDH, cTnI, serum nitrite/nitrate concentration, and inflammatory cytokines) and histological assessments of myocardial tissue. These results underscore the importance of understanding genetic mechanisms underlying MI and highlight the potential of UBE2N and IL2RB as biomarkers for non-recurrent and recurrent MI, respectively.

The vacuolar H+-ATPase (V-ATPase) is an ATP-dependent proton pump that functions to control the pH of intracellular compartments as well as to transport protons across the plasma membrane of various cell types, including cancer cells. We have previously shown that selective inhibition of plasma membrane V-ATPases in breast tumor cells inhibits the invasion of these cells in vitro. We have now developed a nanobody directed against an extracellular epitope of the mouse V-ATPase c subunit. We show that treatment of 4T1-12B mouse breast cancer cells with this nanobody inhibits V-ATPase-dependent acidification of the media and invasion of these cells in vitro. We further find that injection of this nanobody into mice implanted with 4T1-12B cells orthotopically in the mammary fat pad inhibits metastasis of tumor cells to lung. These results suggest that plasma membrane V-ATPases represent a novel therapeutic target to limit breast cancer metastasis.

Lung cancer (LC) is the most common cancer in males. As per GLOBOCAN 2020, 8.1 % of deaths and 5.9 % of cases of LC were reported in India. Our laboratory has previously reported the significant anticancer potential of 5H-benzo[h]thiazolo[2,3-b]quinazoline analogues. In this study, we have explored the anticancer potential of 7A {4-(6,7-dihydro-5H-benzo[h]thiazolo[2,3-b]quinazolin-7-yl)phenol} and 9A {7-(4-chlorophenyl)-9-methyl-6,7dihydro-5H-benzo[h]thiazolo[2,3-b]quinazoline}by using in-vitro and in-vivo models of LC. In this study, we investigated the antiproliferative potential of quinazoline analogues using A549 cell line to identify the best compound of the series. The in-vitro and molecular docking studies revealed 7A and 9A compounds as potential analogues. We also performed acute toxicity study to determine the dose. After that, in-vivo studies using urethane-induced LC in male albino Wistar rats carried out further physiological, biochemical, and morphological evaluation (SEM and H&E) of the lung tissue. We have also evaluated the antioxidant level, inflammatory, and apoptotic marker expressions. 7A and 9A did not demonstrate any signs of acute toxicity. Animals treated with urethane showed a significant upregulation of oxidative stress. However, treatment with 7A and 9A restored antioxidant markers near-normal levels. SEM and H&E staining of the lung tissue demonstrated recovered architecture after treatment with 7A and 9A. Both analogues significantly restore inflammatory markers to normal level and upregulate the intrinsic apoptosis protein expression in the lung tissue. These experimental findings demonstrated the antiproliferative potential of the synthetic analogues 7A and 9A, potentially due to their antiinflammatory and apoptotic properties.

Abstract Background Cancer cells have accelerated glycolysis rate, resulting in excessive lactate generation, which is critical in rapidly growing cancerous cells. Lactate is primarily transported by MCT-1/MCT-4, the two H+/lactate transporters that promote cellular proliferation and growth. Through in-silico, in-vitro, and in-vivo investigation, we aimed to find new dual MCT-1 and MCT-4 inhibitor for therapeutic intervention in breast cancer. Material and Methods A library of 4098 natural product-like compounds (HY-L057L) was retrieved and screened based on structural similarity with Syrosingopine (above70%). Among them, we found LC-mHTT-AN2 as a potential molecule that inhibits MCT-1 and MCT-4 symporters through docking study, pharmacokinetic(ADMET) profiling.Further, compound was tested for the in-vitro cytotoxicity(via MTTassay) and antiapototic activity ( via DAPI, AO/EtBr,JC-1) against MCF-7 cells. we also examined the in-vivo anticancer activity against MNU (Methyl Nitrosourea) induced mammary gland carcinoma in Wistar rat through carmine staining, SEM,biochemical and western blotting analysis Results Our in-silico result revealed that LC-mHTT-AN2 has good docking score with both proteins( MCT-1 and MCT-4 ) and favourable ADMET profiling. Further ,in-vitro result demonstrated that LC-mHTT-AN2 has significant IC50 value (4.7µM) and antiapoptotic potential. Once scrutinized against MNU-induced mammary gland carcinoma, LC-mHTT-AN2 significantly restored the altered morphology and ameliorated histopathological, biochemical and lactate production. Furthermore, the western blotting analysis revealed that LC-mHTT-AN2 significantly regulate mitochondrial apoptotic pathway and has demarcating effect upon inhibition of lactate transport and hypoxic microenvironment, demonstrating the preclinical efficacy for treating breast cancer. Conclusions The overall findings from in -silico, in -vitro, and in -vivo support the pre-clinical efficacy of LC-mHTT-AN2 in the treatment of breast carcinoma by combined inhibition of MCT-1 and MCT-4. Further research is needed to verify its usefulness before clinical application.

Stem cell therapy has emerged as a promising approach for regenerative medicine, offering potential treatments for a wide range of diseases and injuries. Although stem cell therapy has great promise, several obstacles have prevented its broad clinical adoption. The effectiveness of therapy has been inhibited by problems such as ineffective stem cell differentiation, low post-transplantation survival rates, and restricted control over stem cell behavior. Furthermore, the implementation of stem cell therapies is further complicated by the possibility of immunological rejection and cancer. Innovative strategies that provide precise control over stem cell characteristics and maximize their therapeutic potential are desperately needed to overcome these obstacles. Recent studies have shown that the effectiveness of stem cell treatments can be greatly increased by nanoscale advances. By establishing an ideal microenvironment and precisely offering growth factors, nanomaterials such as nanoparticles, nanocomposites, and quantum dots have been demonstrated to improve stem cell differentiation and proliferation. This article provides an overview of the recent trends and applications of nanoscale innovations in the context of stem cell therapy. The recent development of precision medicine has been facilitated by the incorporation of nanotechnology into stem cell therapy. The ability to manipulate stem cells at the nanoscale offers unprecedented control over their behavior and function, opening up exciting possibilities for personalized and highly effective therapeutic interventions. This review paper highlights the recent trends and applications of nanotechnology in advancing stem cell therapy, showcasing its potential to revolutionize regenerative medicine.

Lung cancer (LC) ranks second most prevalent cancer in females after breast cancer and second in males after prostate cancer. Based on the GLOBOCAN 2020 report, India represented 5.9% of LC cases and 8.1% of deaths caused by the disease. Several clinical studies have shown that LC occurs because of biological and morphological abnormalities and the involvement of altered level of antioxidants, cytokines, and apoptotic markers. In the present study, we explored the antiproliferative activity of indeno[1,2-d]thiazolo[3,2-a]pyrimidine analogues against LC using in-vitro, in-silico, and in-vivo models. In-vitro screening against A549 cells revealed compounds 9B (8-methoxy-5-(3,4,5-trimethoxyphenyl)-5,6-dihydroindeno[1,2-d]thiazolo[3,2-a]pyrimidine) and 12B (5-(4-chlorophenyl)-5,6-dihydroindeno[1,2-d]thiazolo[3,2-a]pyrimidine) as potential pyrimidine analogues against LC. Compounds 9B and 12B were docked with different molecular targets IL-6, Cyt-C, Caspase9, and Caspase3 using AutoDock Vina 4.1 to evaluate the binding affinity. Subsequently, in-vivo studies were conducted in albino Wistar rats through ethyl-carbamate (EC)- induced LC. 9B and 12B imparted significant effects on physiological (weight variation), and biochemical (anti-oxidant [TBAR's, SOD, ProC, and GSH), lipid (TC, TG, LDL, VLDL, and HDL)], and cytokine (IL-2, IL-6, IL-10, and IL-1β) markers in EC-induced LC in albino Wistar rats. Morphological examination (SEM and H&E) and western blotting (IL-6, STAT3, Cyt-C, BAX, Bcl-2, Caspase3, and caspase9) showed that compounds 9B and 12B had antiproliferative effects. Accordingly, from the in-vitro, in-silico, and in-vivo experimental findings, we concluded that 9B and 12B have significant antiproliferative potential and are potential candidates for further evaluation to meet the requirements of investigation of new drug application.

Isorhamnetin (C16H12O7), a 3′-O-methylated derivative of quercetin from the class of flavonoids, is predominantly present in the leaves and fruits of several plants, many of which have traditionally been employed as remedies due to its diverse therapeutic activities. The objective of this in-depth analysis is to concentrate on Isorhamnetin by addressing its molecular insights as an effective anticancer compound and its synergistic activity with other anticancer drugs. The main contributors to Isorhamnetin’s anti-malignant activities at the molecular level have been identified as alterations of a variety of signal transduction processes and transcriptional agents. These include ROS-mediated cell cycle arrest and apoptosis, inhibition of mTOR and P13K pathway, suppression of MEK1, PI3K, NF-κB, and Akt/ERK pathways, and inhibition of Hypoxia Inducible Factor (HIF)-1α expression. A significant number of in vitro and in vivo research studies have confirmed that it destroys cancerous cells by arresting cell cycle at the G2/M phase and S-phase, down-regulating COX-2 protein expression, PI3K, Akt, mTOR, MEK1, ERKs, and PI3K signaling pathways, and up-regulating apoptosis-induced genes (Casp3, Casp9, and Apaf1), Bax, Caspase-3, P53 gene expression and mitochondrial-dependent apoptosis pathway. Its ability to suppress malignant cells, evidence of synergistic effects, and design of drugs based on nanomedicine are also well supported to treat cancer patients effectively. Together, our findings establish a crucial foundation for understanding Isorhamnetin's underlying anti-cancer mechanism in cancer cells and reinforce the case for the requirement to assess more exact molecular signaling pathways relating to specific cancer and in vivo anti-cancer activities.

Pioglitazone (PGL), an antidiabetic drug within the thiazolidinedione class, is utilized in managing type-2-diabetes-mellitus. Additionally, it has been observed to exhibit therapeutic effects on various inflammatory markers. PGL falls under BCS Class II, which is defined by its low water solubility and slow dissolution rate. The dual aim of our study was to optimize an oral as well as topical PGL-loaded nanoemulsion (PGL-NE) in order to enhance their bioavailability orally as well as topically. Using the Box-Behnken Design Expert software, Pioglitazone-loaded nanoemulsions (NanoE1 to NanoE17) were optimized. The ultrasonication method was employed following excipients-screening and the preparation of PTPD. The optimized nanoformulation (NanoE13) was determined based on its particle size (137.7 +/- 7.92 nm), %Transmittance (91.07 +/- 1.03), and PDI (0.221 +/- 0.004). NanoE13 exhibited sustained drug release, adhering to the Korsemeyer-Peppas model. When converted into a gel (PGL-NE-Gel), it enhanced permeation of the skin and topical bioavailability. The results of dermatokinetic demonstrated a significant i.e., p.0.001 increase in C-Skinmax & AUC(0-8h) in skin treated with the optimized PGL-NE-Gel compared to PGL-NE and conventional PGL gel. After 2 weeks, reductions of 40.06 %, 27.34 %, and 37.45 % were observed for diabetic control, PGL-S, and PGL-NE, respectively. PGL-NE also significantly lowered the levels of blood-glucose i.e., p < 0.05 as compared to the diabetic-control-group throughout the experiment. We successfully developed a novel PGL-NE and PGL-NE-Gel, which enhanced the solubility, skin permeation, and bioavailability of Pioglitazone for both oral and topical applications.

PURPOSE:The present study investigated the nephron-testicular protective effects of sesamin against cisplatin (CP)-induced acute renal and testicular injuries. METHODS:Thirty-two male Wistar rats were allocated to receive carboxymethylcellulose (0.5%, as sesamin vehicle), CP (a single i.p. 5 mg/kg dose), CP plus sesamin at 10 or 20 mg/kg orally for 10 days. RESULTS:Data analysis showed significant increases in serum urea, creatinine, interleukin (IL)-1, IL-6, and tumor necrosis factor-α (TNF-α), as well as renal and testicular tissue malondialdehyde and nitric-oxide concentrations in CP-intoxicated rats in comparison to control animals. On the contrary, rats treated with CP only exhibited significantly lower (p < .05) serum testosterone, tissue glutathione, and activities of endogenous antioxidant enzymes compared to control rats. Histopathologically examining CP-intoxicated rats' tissues using H&E and PAS stains showed atrophied glomeruli, interstitial inflammatory cells, atypic tubular epithelium with focal apoptosis, and reduced mucopolysaccharide content. Further, immunohistochemical staining of the same group revealed an increase in p53 and cyclooxygenase-II (Cox-II) expression in renal and testicular tissues. Treatment with sesamin alleviated almost all the changes mentioned above in a dose-dependent manner, with the 20 mg/kg dose restoring several parameters' concentrations to normal ranges. CONCLUSIONS:In brief, sesamin could protect the kidneys and testes against CP toxicity through its antioxidant, anti-inflammatory, and anti-apoptotic effects.

Enhancing the brain bioavailability of paliperidone by developing a novel mucoadhesive-CS-coated-PLP-NE and examining the quantity of PLP by developing a novel method of ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) in schizophrenic rat brain treatment.

Abstract ID 128681Poster Board 056Melanoma remains one of the difficult malignancies to treat, due to its ability to evade anti-tumor immune responses via employing tolerance mechanisms such as negative immune checkpoint molecules. While immune checkpoint inhibitors (ICIs) have been increasingly used to treat melanoma, a considerable number of patients do not respond to this treatment, indicating the development of resistance mechanisms such as intrinsic tumor characteristics and an immunosuppressive tumor microenvironment (TME). Multiple studies have documented that an inflamed TME increases the therapeutic efficacy of ICIs via upregulating the expression of the T-cell inflamed signature, a set of genes linked with dendritic cell and T-cell anti-tumor response. Notably, histone deacetylases (HDACs) are frequently deregulated in melanomas. In addition, it has been demonstrated that HDACs can modulate the transcription of PD-1/PD-L1, and other genes linked to immune evasion. The objective of our research was to examine the association between the T-cell inflamed signature and histone deacetylases (HDACs) in melanoma patients. The data were obtained by use of the cBioPortal for Cancer Genomics and were sourced from melanoma dataset: "Skin Cutaneous Melanoma (TCGA, Firehose Legacy) (n = 287)”. The analysis was focused on the co-expression of T-cell inflamed signature genes and the HDAC family. The study revealed a negative correlation between the mRNA expression of HDAC4 and several T-cell inflamed signature genes, including PD-L1, LAG3, IDO1, TIGIT, ICOS, STAT1, IFNG, CD8A, CD4, IRF1, PRF1, GZMA, GZMB, GZMK, CCL3, CCL4, CCL5, CXCL9, CXCL10, CXCR6, CMKLR1, NKG7, PSMB10, HLA-DMA, HLA-DMB, HLA-DOA, HLA-DOB, HLA-DQA1, HLA-DRB1, and HLA-E. Furthermore, we observed a significant methylation of T-cell inflamed signature genes in the group exhibiting elevated HDAC4 expression. Consequently, their transcriptions were rescued in the group who had a low HDAC4 expression. Based on our findings, HDAC4 may negatively modulate TME and ICI efficacy in melanoma. Furthermore, our research suggested a new method of targeting a specific epigenetic change, which has implications for developing a novel therapeutic strategy. Specifically, we propose combining an HDAC4 inhibitor with ICIs to enhance their effectiveness in melanoma. Further studies are required to validate our results.

Background: The combination of patients' unsustainable use of antibiotics and doctors' inappropriate prescribing practices has resulted in challenges for disease control in the community and inadequate delivery of healthcare services. Methods: The General Public's understanding and awareness of AMR and the One Health Approach policy were evaluated through the use of a cross-sectional study questionnaire. Results were analysed statistically using SPSS software Study participants' knowledge, attitude, and perspective scores were predicted using ordinal logistic regression analysis. Results: Participants who were self-employed have good knowledge (41.8%) about AMR and One Health Approach policy and also had higher KAP scores (CI = 4.885). Factors like age and gender were observed to have no impact on aggregate KAP scores. Conclusion: In light of the knowledge gaps identified in our survey, development and implementation of health education and campaign will help to improve awareness among General Public. DOI: https://doi.org/10.52783/jchr.v13.i4.1339

ID 53622 Poster Board 435 Small cell lung cancer (SCLC) is a more aggressive type of lung cancer with a worse prognosis. Immune checkpoint inhibitors (ICIs) showed effectiveness against SCLC, however, the response rate was low. Several studies have reported that therapeutic efficacy of ICIs was improved when the tumor microenvironment (TME) exhibited inflamed phenotype including the expression of a T-cell inflamed signature which is a set of genes associated with T-cells and dendritic cells anti-tumor response. The induction of this signature can predict ICI effectiveness. However, epigenetic modifications such as DNA methylation can switch the TME into non-inflamed phenotype and eventually lead to ICI resistance. The aim of the study was to investigate the alterations in DNA methylation levels of T-cell inflamed signature genes in SCLC. Publicly available 450K DNA methylation array data of SCLC was used for our in silico analysis. CpG sites at promoter region of the T-cell inflamed genes were profiled, evaluated, and differentially methylated sites with a methylation difference (delta beta) of ≥ 5% were selected. The investigation was conducted on a dataset containing 13 controls and 11 SCLC cases. The analysis revealed that 18 of the total 37 T-cell inflamed signature genes, including CCL2, CCL5, CD27, CD276, CD4, CMKLR1, FOXP3, GZMB, GZMK, IDO1, IFNG, IRF1, LAG3, NKG7, PRF1, PSMB10, TIGIT, and TNF, had at least one differentially methylated site at the promotor area and 15 of them had at least two sites. The differentially methylated genes were hypermethylated in SCLC with methylation differences ranging between 5 % and 36 %. Interestingly, LAG3 and PRF1 were the most densely methylated genes with eight differentially methylated sites at the promoter region (average methylation difference = 23% and 17%, respectively). Our discovery of changes in DNA methylation status of T-cell inflamed genes in SCLC can open a novel avenue for targeting this epigenetic modification to reprogram TME and promote ICI susceptibility. Moreover, our data provided support for a novel therapeutic approach of combining hypomethylating agents such as decitabine or azacytidine with ICIs to improve their outcomes in SCLC. However, additional in vitro and in vivo studies are required to validate our results.

The fixed dose combination of valsartan (VAL) and hydrochlorothiazide (HCTZ) is the most commonly prescribed medicine for the effective treatment of hypertension. In this study, a simple sensitive and accurate liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the simultaneous quantitation of VAL and HCTZ in human plasma by using irbesartan (IRB) and hydroflumethiazide (HFMZ) as their specific internal standards (ISs). HLB cartridge-based solid-phase extraction was used for the extraction of analytes and ISs. The chromatographic separation was achieved on Lichrocart RP Select (125 × 4 mm), 5 nm with the mobile phase composition of acetonitrile: 10 mM ammonium acetate buffer: 95:05, v/v, at flow rate of 0.5 mL/min. The turbo ion electrospray ionization in negative mode was used as ion source for the sample ionization. The precursor to product ion transitions were 434.10 > 179.10 (VAL), 295.70 > 204.90 (HCTZ), 427.10 > 192.90 (IRB), and 329.90 > 302.40 (HFMZ) for detection and quantification of analytes and their ISs. The retention times of VAL and HCTZ were 1.90 min and 2.30 min, respectively. The range for the calibration curves of VAL and HCTZ were 50.2–6018.6 ng/mL and 1.25–507.63 ng/mL, respectively, with good linearity having correlation coefficient values of ≥0.995 for both VAL and HCTZ. All validation parameter results (selectivity, precision and accuracy, matrix effects and stabilities) were within the acceptable range as per USFDA guideline for bioanalytical method validation. The intra-day and inter-day accuracy data for VAL were within the range of 105.68–114.22% and 98.41–108.16%, respectively, whereas for HCTZ they were 87.01–101.18% and 95.16–99.37%, respectively. The ion suppression effects produced for VAL and ion enhancement effects produced for HCTZ were insignificant according to the proposed sample cleanup procedure. The developed LC-MS/MS method was successfully applied to bioequivalence study on healthy volunteers.

ID 53598 Poster Board 438 Solid tumors in children represent approximatively 30% of all pediatric cancers. The most prevalent types are Wilms tumor, retinoblastoma, neuroblastoma, and Ewing9s sarcoma. The literature suggested that there is a degree of similarity between these conditions (such as age of diagnosis, prognosis, etc.), indicating that they may share a similar pathogenesis. The objective of our study was to investigate the DNA methylation profile of common childhood solid tumors in order to find a common signature that can be used to understand the underlying common molecular mechanism and to identify new pharmacological targets. This was an in silico study which used publicly available data from a DNA methylation array. Illumina 450K array datasets of the four conditions were downloaded from the GEO data repository and processed on RStudio. Differentially methylated sites were selected based on scoring Student’s t-test P-value ≤ 0.001 and a methylation difference (delta beta) ≥ 20%. Differentially methylated genes were defined as genes having at least two differentially methylated sites according to the human GRCh37/hg19 annotation. The study included 82 controls and 357 cases of childhood solid tumors. The analysis revealed that there were 13,472 novel differentially methylated sites that shared a common methylation profile between all the tumors. Using the hierarchical clustering method, the differentially methylated sites properly clustered all cases of childhood solid tumors. In addition, the analysis identified 17 differentially methylated genes that were involved in cell-cell communication, signal transduction and immune system pathways. The novel discovery of the common differentially methylated sites and genes among common childhood solid tumors provided an insight towards understanding their shared molecular pathogenesis. In addition, the identified candidate genes can be used as biomarkers for the clinical diagnosis and as therapeutic pharmacoepigenomic targets of childhood solid tumors.