Fold Change Increase Research Articles

Abstract Tu080: Unique Physiology of Fibroblast and Immune Cells in the Right Ventricle

Introduction: Failure of the right ventricle (RV) is a main component of the morbidity and mortality of pulmonary hypertension (PH). There exists no specific treatment for RV dysfunction and failure and the only cure for many patients (especially in the case of pulmonary arterial hypertension) remains transplantation. This is partially due to the critical lack of understanding of RV biology; therapeutic strategies that are beneficial for LV failure have worse outcomes when applied to those with RV failure. RV dysfunction is characterized by exaggerated fibrosis and divergent contractile and hypertrophic responses to several drugs when compared to LV dysfunction. Thus, the RV may contain distinct cellular physiology compared to the LV that has high significance for the treatment of PH. Methods: Tissue digests from the RV and LV of Sprague Dawley rats were collected. We utilized digests to collect RNA for RNA-seq, and prepared cells for flow cytometry and fibroblast isolations. Fibroblasts were cultured for proliferation assays and cell signaling studies. Results: We found that the RV and LV have significant differences in expression of immune-related and fibrotic genes. The RV of control rats had an average of four-fold higher density of immune cells, namely macrophages/monocytes, compared to the LV (P<0.05 via paired t-test). Fibroblasts from the RV of control rats, which had a higher proliferation rate and an alteration of metabolic genes than those from the LV (P<0.05 via two-way ANOVA), also had a higher fold-change increase in periostin and CTGF expression upon treatment with TGFβ. Conclusions: The results demonstrate that in comparison to the LV, the RV is made up of a unique population of cells and has a unique capacity for fibrotic response that is reflected in its molecular makeup, which may contribute to its distinct response to PH. Future experiments will focus on determining the molecular mechanisms driving these differences. This knowledge is expected to yield new therapeutic approaches for RV dysfunction and failure occurring with PH, which currently have no specific treatment.

Association between miRNAs in serum at 10-14 gestational weeks and spontaneous preterm delivery.

Preterm delivery (PTD) is the leading cause of death in children under 5 years of age. Cervical shortening detected by ultrasound can be used to predict PTD, but prediction is not perfect, and complementary diagnostic markers are needed. Recently, specific plasma microribonucleic acid (miRNAs) detected in early second trimester were shown to be associated with spontaneous PTD in high-risk women with a singleton pregnancy. The aim of this study was to explore to what extent these miRNAs are associated with spontaneous PTD and cervical length in a general population. This study is a nested case-control study within the CERVIX study. The CERVIX study evaluated the ability of cervical length screening with transvaginal ultrasound to identify women at risk of PTD. In the present study, women who delivered spontaneously <34 weeks (n = 61) were compared with a control group of women who delivered at full term (39 + 0 to 40 + 6 gestational weeks, n = 205). Archived serum samples were analyzed with RT-qPCR for miRNA expression levels of let-7a-5p, miR-150-5p, miR-15b-5p, miR-185-5p, miR-191-5p, miR-19b-3p, miR-23a-3p, miR-374a-5p, and miR-93-5p. The mean relative expression was compared between the groups. Sub-analyses were performed for women delivering <32, <30, and <28 weeks vs the full-term group. The analyzed miRNAs were not significantly differentially expressed in women delivering <34 weeks compared to those delivering at full term. MiR-191-5p and miR-93-5p were significantly overexpressed in women who delivered <32 weeks, and further increase in fold change was observed with decreasing gestational age at delivery. The level of miR-15b-5p was significantly higher in women delivering at <30 weeks compared to those delivering at full term. Our study shows that overexpression of miR-93-5p, miR-15b-5p, and miR-191-5p in serum at early gestation is associated with spontaneous PTD in a general population. Further research is needed to evaluate the potential of these miRNAs as future biomarkers for spontaneous PTD, as well as their pathophysiological role in spontaneous PTD.

Microbiota based personalized nutrition improves hyperglycaemia and hypertension parameters and reduces inflammation: a prospective, open label, controlled, randomized, comparative, proof of concept study.

Recent studies suggest that gut microbiota composition, abundance and diversity can influence many chronic diseases such as type 2 diabetes. Modulating gut microbiota through targeted nutrition can provide beneficial effects leading to the concept of personalized nutrition for health improvement. In this prospective clinical trial, we evaluated the impact of a microbiome-based targeted personalized diet on hyperglycaemic and hyperlipidaemic individuals. Specifically, BugSpeaks®-a microbiome profile test that profiles microbiota using next generation sequencing and provides personalized nutritional recommendation based on the individual microbiota profile was evaluated. A total of 30 participants with type 2 diabetes and hyperlipidaemia were recruited for this study. The microbiome profile of the 15 participants (test arm) was evaluated using whole genome shotgun metagenomics and personalized nutritional recommendations based on their microbiota profile were provided. The remaining 15 participants (control arm) were provided with diabetic nutritional guidance for 3 months. Clinical and anthropometric parameters such as HbA1c, systolic/diastolic pressure, c-reactive protein levels and microbiota composition were measured and compared during the study. The test arm (microbiome-based nutrition) showed a statistically significant decrease in HbA1c level from 8.30 (95% confidence interval (CI), [7.74-8.85]) to 6.67 (95% CI [6.2-7.05]), p < 0.001 after 90 days. The test arm also showed a 5% decline in the systolic pressure whereas the control arm showed a 7% increase. Incidentally, a sub-cohort of the test arm of patients with >130 mm Hg systolic pressure showed a statistically significant decrease of systolic pressure by 14%. Interestingly, CRP level was also found to drop by 19.5%. Alpha diversity measures showed a significant increase in Shannon diversity measure (p < 0.05), after the microbiome-based personalized dietary intervention. The intervention led to a minimum two-fold (Log2 fold change increase in species like Phascolarctobacterium succinatutens, Bifidobacterium angulatum, and Levilactobacillus brevis which might have a beneficial role in the current context and a similar decrease in species like Alistipes finegoldii, and Sutterella faecalis which have been earlier shown to have some negative effects in the host. Overall, the study indicated a net positive impact of the microbiota based personalized dietary regime on the gut microbiome and correlated clinical parameters.

NF-κB in the Radiation Response of A549 Non-Small Cell Lung Cancer Cells to X-rays and Carbon Ions under Hypoxia.

Cellular hypoxia, detectable in up to 80% of non-small cell lung carcinoma (NSCLC) tumors, is a known cause of radioresistance. High linear energy transfer (LET) particle radiation might be effective in the treatment of hypoxic solid tumors, including NSCLC. Cellular hypoxia can activate nuclear factor κB (NF-κB), which can modulate radioresistance by influencing cancer cell survival. The effect of high-LET radiation on NF-κB activation in hypoxic NSCLC cells is unclear. Therefore, we compared the effect of low (X-rays)- and high (12C)-LET radiation on NF-κB responsive genes' upregulation, as well as its target cytokines' synthesis in normoxic and hypoxic A549 NSCLC cells. The cells were incubated under normoxia (20% O2) or hypoxia (1% O2) for 48 h, followed by irradiation with 8 Gy X-rays or 12C ions, maintaining the oxygen conditions until fixation or lysis. Regulation of NF-κB responsive genes was evaluated by mRNA sequencing. Secretion of NF-κB target cytokines, IL-6 and IL-8, was quantified by ELISA. A greater fold change increase in expression of NF-κB target genes in A549 cells following exposure to 12C ions compared to X-rays was observed, regardless of oxygenation status. These genes regulate cell migration, cell cycle, and cell survival. A greater number of NF-κB target genes was activated under hypoxia, regardless of irradiation status. These genes regulate cell migration, survival, proliferation, and inflammation. X-ray exposure under hypoxia additionally upregulated NF-κB target genes modulating immunosurveillance and epithelial-mesenchymal transition (EMT). Increased IL-6 and IL-8 secretion under hypoxia confirmed NF-κB-mediated expression of pro-inflammatory genes. Therefore, radiotherapy, particularly with X-rays, may increase tumor invasiveness in surviving hypoxic A549 cells.

Abstract CT022: Mutant KRAS peptide-based vaccine in patients at high risk of developing pancreatic cancer: Preliminary analysis from a phase I study

Abstract Background: The development of pancreatic ductal adenocarcinoma (PDAC) from the first driver event takes &amp;gt;10 years, providing a unique window of opportunity for early interception. As a frequent alteration identified in both PDAC and pancreatic premalignancy, mutant KRAS (mKRAS) has emerged as a potential target for immune-based interception strategies in individuals at high risk of developing pancreatic cancer. Methods: This is a single-arm, open-label, second-in-human phase I study of a pooled synthetic long peptide (SLP) vaccine with poly-ICLC adjuvant targeting up to six mKRAS subtypes (G12D, G12R, G12V, G12A, G12C, G13D) in patients with hereditary predisposition to developing PDAC (NCT05013216). Key inclusion: radiographic evidence of a pancreatic abnormality and eligibility under at least one of three high-risk groups based on family history of PDAC and/or germline mutation status. Vaccination schema: mKRAS vaccine given on weeks 1, 3, 5, and 13. Peripheral blood collected at baseline and weeks 5, 13, and 17 for analysis of immunogenicity by IFN-gamma ELISpot. Co-primary endpoints: safety and mKRAS-specific T cell response. Results: At the time of data cutoff (November 15, 2023), 15 out of 20 planned patients were vaccinated on study and evaluable for immunogenicity. All 15 patients achieved a mKRAS-specific T cell response, defined by the maximal fold change in the pooled average IFN-gamma-producing mKRAS-specific T cell density within 17 weeks post-vaccination (median: 12.3, range: 3.1-167). Differential immunogenicity elicited by individual mKRAS epitopes was noted in this cohort. Specifically, the mKRAS G12V peptide induced the highest fold change increase in antigen-specific T cell expansion (median: 32.6, range: 2.9-243) while the mKRAS G12D peptide was the least immunogenic (median: 7.6, range: 0.4-104). Regarding safety, all adverse events were grade 1 or grade 2 in severity per NCI CTCAE v5.0. The most frequent vaccine-related adverse events were injection site reactions (93.8%), fatigue (62.5%), chills (43.8%), and headache (37.5%). No safety signals were observed. Conclusions: For the first time, we demonstrate mKRAS-specific T cell responses induced by vaccination in a cohort of high-risk individuals with inherited predisposition to PDAC. These findings directly serve as proof-of-concept to advance vaccine strategies focused on intercepting the development of pancreatic cancer in high-risk populations. Support: Stand Up To Cancer, Lustgarten Foundation, and NIH/NCI. Citation Format: Saurav D. Haldar, Amanda Huff, Elizabeth Abou Diwan, Anna Ferguson, Carol Judkins, Jiayun Lu, Hao Wang, Hassan Sinan, Christopher Thoburn, Katherine M. Bever, Mark Yarchoan, Amy M. Thomas, Julie M. Nauroth, Daniel A. Laheru, Michael G. Goggins, Elizabeth M. Jaffee, Nilofer S. Azad, Neeha Zaidi. Mutant KRAS peptide-based vaccine in patients at high risk of developing pancreatic cancer: Preliminary analysis from a phase I study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT022.

Isolation, Characterization and Structure Elucidation of Antidiabetic Compound “2-({4-oxo-1H,2H,3H,4H,4aH-cyclopenta[b]pyridine-2-yl} methyl)-1H, 2H, 3H, 4H, 4aH-Cyclopenta[b]Pyridine-4-one” from the barks of Parkia timoriana (DC.) Merr.

Parkia timoriana (D.C.) Merr is a medicinal plant with long-known usage as a traditional medicine. This plant’s bark and pods have been used in medicine and food supplements. P. timoriana bark was extracted with ethyl acetate and purified using silica gel column chromatography. A pure compound, 2-({4-oxo-1H, 2H, 3H, 4H, 4aH-cyclopenta[b]pyridine-2-yl}methyl)-1H, 2H, 3H, 4H, 4Ah cyclopenta[b]pyridine-4-one (BCPP) was isolated and spectroscopic analysis was performed to determine its chemical structure. Antidiabetic studies were conducted using Bis-Cyclopenta-Pyridin (BCPP) for its enzymatic inhibition. In vitro cell culture assays, gene expression studies and in silico docking studies were done to ascertain the antidiabetic activity. P. timoriana bark was extracted using the solvents based on its polarity: hexane, ethyl acetate and methanol. At a 125 μg/ml concentration, BCPP demonstrated 75% inhibition of alpha-amylase, 74% inhibition of alpha-glucosidase and a 59% increase in glucose uptake by yeast cells (3.3 ± 0.0058 at 125 μg/ml). BCPP increased the expression of the PPARγ gene in 3T3-L1 cells by 0.23-fold. There is an increase in fold change of PPAR- γ at 25 μg/ml – 0.11-fold; at 50 μg /ml – 0.25-fold; at 75 μg /ml – 0.37-fold; 100 μg /ml – 0.73-fold; and at 125 μg/ml, there is an upregulation of 1.10-fold in 3T3-L1 cells. In silico docking studies confirmed the alpha-amylase and alpha-glucosidase inhibitory activities of BCPP. This study suggests the application of BCPP as a potential antidiabetic agent that can be used in drug design and pharmaceutics.

Abstract 3786: Genomic and transcriptomic characterization of peritoneal, lung and liver metastases of colorectal carcinoma reveals site-specific differences

Abstract Background: Metastasis in colorectal cancer (CRC) is a critical factor in patient outcomes, with 20% of patients affected at the time of diagnosis and up to 50% developing metachronous metastatic disease later on. Despite the growing importance of metastatic sites as independent predictors of overall survival (OS) and the ongoing development of new treatments, there has been limited effort to personalize systemic treatment based on specific metastatic locations. Aim: This study aims to compare the genomic landscape of different metastatic sites in CRC to detect novel diagnostic and therapeutic targets. Methods: DNA whole-genome-sequencing and RNA gene expression data from metastatic CRC samples were retrieved from the Hartwig Medical Foundation (HMF). Single nucleotide variants (SNVs), somatic copy number aberrations (SCNAs) and structural variants (SVs) were reported by HMF with their pipelines. In addition, focal amplifications and deletions were identified using GISTIC2 (v6.15.30). We included 464 microsatellite stable samples derived from liver metastases (n=369), peritoneal metastases (n=58) and lung metastases (n=37). We compared DNA mutation frequencies between tumors from different metastatic sites (two-sided Mann-Whitney tests) and performed differential gene expression and pathway enrichment analysis. Results: Liver metastases had the highest number of focal SCNAs per sample (median 33 vs 17 in peritoneum (p &amp;lt; 0.001); vs 18 in lung (p &amp;lt; 0.001)). Furthermore, we observed 39 site-specific SCNAs in liver, 9 in peritoneal and 15 in lung metastases. Of these, the most significant focal SCNAs were an amplification within 20q13.12 (79%) and deletion within 1p35.3 (64%) in peritoneal metastases and a deletion within 18q21.1 (84%) in lung metastases. Furthermore, peritoneal metastases displayed less genomic instability, featuring lower ploidy (median 2.65 vs 2.94 in liver (p=0.025); vs 3.15 in lung (p=0.025)) and lower aneuploidy scores (median 19 vs 26 in liver (p=0.016); vs 30 in lung (p=0.016)) while having no significant differences in tumor mutational burden or individual SNVs. Analysis of SVs revealed that peritoneal metastases exhibited fewer deletions (≥10kb; 1.3x fold change increase in liver and lung) and fewer duplications (≥1kb; 1.4x fold change increase in liver). Differential gene expression analysis uncovered higher expression of gene sets involved in the apical junction and the TGF-beta-signaling pathway in peritoneal metastases. Conclusion: Specific focal SCNA and differences in genomic instability exhibit an association with preferential organotropism in CRC metastasis and could offer potential targets for the development of new diagnostic and therapeutic approaches. Citation Format: Nerma Crnovrsanin, Soufyan Lakbir, Renske d. Wit, Steven L. Ketelaars, Niels F. Kok, Petur Snaebjornsson, Gerrit A. Meijer, Sanne Abeln, Remond J. Fijneman. Genomic and transcriptomic characterization of peritoneal, lung and liver metastases of colorectal carcinoma reveals site-specific differences [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3786.

Liver and ovarian toxicities boosted by bisphenol and gamma radiation in female albino rats.

Bisphenol A (BPA), a carbon-based synthetic polymer compound, was newly classified as an environmental toxicant and an endocrine-disrupting chemical leading to abnormalities in cell proliferation, apoptosis, or migration that contributes to cancer development and progression. This study aims to evaluate the effect of the elevation of γ- radiation dose and BPA on the liver and ovaries of female rats. In this study, eighty female albino rats (130-150g) were used in this work. Rats in this experiment received BPA in ethanol (50mg/kg b. wt.) for 30days, day after day, and in the irradiated groups, animals were administered BPA and then exposed to γ- radiation in doses (2, 4, and 6Gy) one shot dose. Several members of the cytochrome family were examined. Exposure to γ-radiation and BPA showed an increase in cytochrome P450 and b5 fold change. Further, BPA and γ-radiation activate α and β estrogen receptors and also downregulate aromatase (CYT19) fold change. The current results also revealed that BPA and/or γ-radiation regulate the protein expression of the PI3K/Akt signaling pathway. The steroidogenic acute regulatory protein (StAR) appeared to be targeted by BPA and γ-radiation and its relative expression was elevated significantly by raising the γ-radiation dose. In conclusion, exposure to BPA, an endocrine-disrupting chemical, leads to marked toxicity. Additionally, toxicity is heightened by increasing the γ-radiation dose, either alone or in combination with BPA.

Airbrushed nanofibers with bioactive core and antibacterial shell for wound healing application

Acute and chronic wounds are vulnerable to infection and delayed healing and require critical care and advanced wound protection. To overcome the challenges, dual therapy of antibacterial and growth factors will be a novel wound care strategy. The present study explores airbrushed core–shell nanofiber for dual delivery of epidermal growth factor (EGF) and amoxicillin (AMOX) in a sustained manner. A blend of polycaprolactone (PCL)-polyethylene oxide (PEO) was used to prepare the shell compartment for amoxicillin loading and poly-DL-lactide (PDLLA) core for EGF loading by using a customized airbrush setup. Characterization result shows a uniform distribution of nanofibers ranging between 200 and 500 nm in diameter. Amoxicillin loading in the shell compartment offers an initial burst release followed by a sustained release for up to 14 days. Whereas EGF in the core part shows a continuous sustained release throughout the release study.In-vitrostudy indicates the biocompatibility of EGF-AMOX loaded core–shell nanofibers with human dermal fibroblast cell (HDF) cells and a higher cellular proliferation compared to control samples. Gene expression data show an increase in fold change of collagen I and tropoelastin expression, indicating the regenerative properties of EGF-AMOX encapsulated nanofiber. The combination of bioactive core (EGF) and antibiotic shell (amoxicillin) in an airbrushed nanofibrous scaffold is a novel approach, which is the first time explored to deliver sustainable therapy to treat skin wounds. Our results demonstrate that PCL-PEO-Amoxicillin/PDLLA-EGF-loaded core–shell nanofibers are promising dual therapy scaffolds to deliver effective skin wound care, with the possibility of direct deposition on the wound.

A Composite Decision Rule of CD8+ T-cell Density in Tumor Biopsies Predicts Efficacy in Early-stage, Immunotherapy Trials.

To examine whether CD8+ T-cell numbers in paired tumor biopsies in early-stage clinical trials can be used as an early indicator of clinical benefit for cancer immunotherapies. Paraffin sections of tumor biopsies were stained immunohistochemically for CD8+ T cells, which were digitally enumerated. The tumor biopsies were from cancer patients in early-phase trials testing novel immunotherapeutic agents. Paired biopsies taken before the start of treatment and on-treatment were compared. A total of 155 patients were used as the training set and an additional 221 patients were used as the validation set. Using the Cox proportional hazard model, a ≥0.9- increase in fold change (FC) on a ln scale in CD8+ T cells (corresponding to a 2.5-fold increase on the linear scale), from baseline, demonstrated a greater association with prolonged progression-free survival and allowed improved differentiation between groups above and below the threshold. Similarly, a ≥6.2 threshold in geometric mean of the on-treatment density (OTD) of T cells, which approximately corresponds to 500 cells/mm2, correlated with longer PFS. The combination of both criteria (FC and OTD) provided the best discrimination between clinically nonactive and active compounds. We propose that a composite score of CD8+ T-cell density in paired biopsies taken before and on-treatment may be a new biomarker to inform on clinical outcomes in early immunotherapy clinical trials.

Urban wastewater disinfection by iron chelates mediated solar photo-Fenton: Effects on seven pathogens and antibiotic resistance transfer potential

The effects of solar photo-Fenton (SPF) process mediated by the iron chelate Fe3+ imminodisuccinic acid (Fe:IDS) on both the inactivation of seven relevant pathogens and the potential for antibiotic resistance transfer (degradation of antibiotic resistance genes (ARGs) and after treatment regrowth), in real secondary treated urban wastewater, were investigated for the first time. A comparison with results obtained by sunlight/H2O2 process and Fe3+ ethylenediaminedisuccinic acid (Fe:EDDS) SPF was also carried out. ARGs were quantified by polymerase chain reaction (PCR) in samples before and after (3 h) the treatment. The persistence of the selected pathogens and ARGs was also evaluated in regrowth tests (72 h) under environmentally mimicking conditions. Fe:IDS SPF resulted to be more effective (from 1.4 log removal for Staphylococcus spp. to 4.3 log removal for Escherichia coli) than Fe:EDDS SPF (from 0.8 log removal for Pseudomonas aeruginosa to 2.0 log removal for Total coliphages) and sunlight/H2O2 (from 1.2 log removal for Clostridium perfringens to 3.3 log removal for E. coli) processes for the seven pathogens investigated. Potential pathogens regrowth was also severely affected, as no substantial regrowth was observed, both in presence and absence of catalase. A similar trend was observed for ARGs removal too (until 0.001 fold change expression for qnrS after 3 h). However, a poor effect and a slight increase in fold change was observed after treatment especially for gyrA, mefA and intl1. Overall, the effect of the investigated processes on ARGs was found to be ARG dependent. Noteworthy, coliphages can regrow after sunlight/H2O2 treatment unlike SPF processes, increasing the risk of antibiotic resistance transfer by transduction mechanism. In conclusion, Fe:IDS SPF is an attractive solution for tertiary treatment of urban wastewater in small wastewater treatment plants as it can provide effective disinfection and a higher protection against antibiotic resistance transfer than the other investigated processes.

PSV-19 Supplementation of a Multi-Strain Probiotic Increases the Expression of Biomarkers of Immunity in Savannah X Spanish Goat Blood

Abstract Probiotics are considered a safe, non-chemical alternative to antibiotics and have been shown to boost the host immune response of livestock. The present study investigated the effect of Gut Candy (GC), a multi-strain probiotic supplementation, on immune gene expression in Spanish X Savannah goats. Seven adult female Spanish X Savannah goats (n = 7) received a daily dose (28 g) of a GC (containing Lactobacillus acidophilus, Enterococcus faecium, Bacillus subtilis, Propioni bacterium freudenreichii, and Aspergillus oryza) for 60 days. The control group received no probiotics (n = 7). Blood (10 mL) was collected aseptically from the jugular vein on days 0 and 60. Total RNA was extracted from whole blood, reverse transcribed to cDNA, and quantitative real-time PCR (RT-PCR) was performed using specific primers for TLR2, TLR4, TNF-α, IL1B, IL10, and MPO. The fold changes (FC) in relative gene abundance were analyzed using Livak's method. β-actin and GAPDH were used as internal controls and for normalization. Fold changes (FC) were considered significant at -2 ≥ FC ≥ 2. After 60 days of GC supplementation, there was an increase in fold change in relative gene abundance in IL10 (FC = 3.66), TNF-α (FC = 11.7), IL1β (FC = 19.59), TLR4 (FC = 5.09), and MPO (FC = 2.05). GC also tended to increase the FC of TLR 2(FC = 1.86). The results from the study demonstrate that probiotic supplementation modulates the expression of immune genes in goats. Incorporating probiotics supplementation into the feed regimen of livestock could be beneficial to the animals' health and overall performance.

Comparative RNA-seq analysis of resistant and susceptible banana genotypes reveals molecular mechanisms in response to banana bunchy top virus (BBTV) infection

Bananas hold significant economic importance as an agricultural commodity, serving as a primary livelihood source, a favorite fruit, and a staple crop in various regions across the world. However, Banana bunchy top disease (BBTD), which is caused by banana bunchy top virus (BBTV), poses a considerable threat to banana cultivation. To understand the resistance mechanism and the interplay of host suitability factors in the presence of BBTV, we conducted RNA-seq-based comparative transcriptomics analysis on mock-inoculated and BBTV-inoculated samples from resistant (wild Musa balbisiana) and susceptible (Musa acuminata ‘Lakatan’) genotypes. We observed common patterns of expression for 62 differentially expressed genes (DEGs) in both genotypes, which represent the typical defense response of bananas to BBTV. Furthermore, we identified 99 DEGs exclusive to the 'Lakatan' banana cultivar, offering insights into the host factors and susceptibility mechanisms that facilitate successful BBTV infection. In parallel, we identified 151 DEGs unique to the wild M. balbisiana, shedding light on the multifaceted mechanisms of BBTV resistance, involving processes such as secondary metabolite biosynthesis, cell wall modification, and pathogen perception. Notably, our validation efforts via RT-qPCR confirmed the up-regulation of the glucuronoxylan 4-O-methyltransferase gene (14.28 fold-change increase), implicated in xylan modification and degradation. Furthermore, our experiments highlighted the potential recruitment of host's substrate adaptor ADO (30.31 fold-change increase) by BBTV, which may play a role in enhancing banana susceptibility to the viral pathogen. The DEGs identified in this work can be used as basis in designing associated gene markers for the precise integration of resistance genes in marker-assisted breeding programs. Furthermore, the findings can be applied to develop genome-edited banana cultivars targeting the resistance and susceptibility genes, thus developing novel cultivars that are resilient to important diseases.

The Effects of Adjuvant Chemoradiotherapy (CRT) on Myeloid-Derived Suppressor Cell (MDSC) Subsets in Glioblastoma (GBM)

GBM continues to be a diagnosis with an exceedingly low survival rate despite standard therapy of resection followed by concurrent CRT. MDSC, immunosuppressive myeloid cells that aid immune system evasion by the tumor, expand during fractionated RT. To date, little is known about the effects of CRT on MDSC subsets. The goal of our pilot study is to compare peripheral blood MDSC subset frequency in patients (pts) undergoing CRT. We hypothesize that the pro-tumor, monocytic MDSC (M-MDSC) subset increases following CRT. Pts over the age of 18 yo with a new diagnosis of GBM from a single institution participated in the study between 7/2022 -1/2023. Exclusion criteria included prior history of brain RT. Baseline peripheral blood samples were collected within one week prior to CRT start and post-CRT samples were collected within the last week of CRT. Peripheral blood mononuclear cells (PBMC) were isolated from whole blood and freshly stained for intracellular flow cytometric analyses. Total MDSC (singlet, viable, non-lymphocyte, CD11b+CD33+ HLA-DR- cells) and subsets (M-MDSC, granulocytic (G-MDSC), and early MDSC (eMDSC)) were identified. Percent frequency (%fx) of total and MDSC subsets as well as MDSC activation status (n = 3) (interleukin (IL)-10, transforming growth factor-β (TGF-β), interferon gamma (IFNγ)) was compared before and after CRT. The average age of pts receiving CRT (n = 4) was 70 yo. Fractionated RT consisted of CF RT (n = 2), 6000 cGy in 30 fractions, and HF RT (n = 2), 4005 cGy in 15 fractions. All pts received concurrent chemotherapy with temozolomide. The %fx of MDSC of non-lymphocytes started at 29.2% prior to CRT and decreased to 9.9% at the end of CRT. The fold change of %fx of total MDSC in CF RT and HF RT was 0.39 and 0.29, respectively. The mean change in %fx of MDSC subsets before and after CRT are shown in Table 1. The mean %fx of TGF-β-expressing MDSC in all GBM pts increased by 3.5% after CRT. The mean IFNγ+ MDSC %fx for all GBM pts decreased after CRT from 24.2% to 16.2% with a corresponding decrease in geometric mean fluorescence intensity (GMFI). The mean %fx of IL-10+ MDSC decreased by 1.5% at the end of CRT with a decrease in GMFI from 5550 to 1806. In this limited dataset of pts with GBM receiving standard of care adjuvant therapy, we identified an expansion in the eMDSC and decrease in the M-MDSC subsets. These early results suggest HF RT may promote immunity that is more supportive of anti-tumor function, with a lesser increase in fold change of total MDSC after CRT. A more detailed understanding of the effect of RT on myeloid subpopulations is essential to addressing immune suppression in pts with GBM.

Antiproliferative and pro-apoptotic properties against colon carcinoma cell line of the marine sponge haliclona sp. From poblacion, kauswagan, lanao del norte, Philippines

Marine sponge Haliclona sp. was examined for its anticancer potential focusing on the antiproliferative and pro-apoptotic properties against HCT116 (colon) cancer cell line. The 16 fractions obtained from Haliclona sp. were subjected to antiproliferative and pro-apoptotic evaluation via MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and Caspase-Glo 3/7 assay, respectively, at 30 μg/mL concentration. Fractions 10, 13, 14, and 15 exhibited a significant decrease (p &lt; 0&gt; 50�ll viability) against HCT116. Only fraction 8 showed a significant increase (p &lt; 0.05) in relative light unit (RLU) and fold change (FC) (1809.78 and 2.42), more than two times larger, compared to the negative control (754.89 RLU and 1.00 FC). This result demands further study on fraction 8 to identify and characterize the structure of the compounds responsible for its bioactivity and hence could be a source of drug leads.

Investigation of Expression Profile of Placenta-specific 1 (PLAC1) in Acute Myeloid and Lymphoid Leukemias.

Placenta-specific 1 (PLAC1) is one of the cancer-testis-placenta antigens that has no expression in normal tissue except placenta trophoblast and testicular germ cells, but is overexpressed in a variety of solid tumors. There is a lack of studies on the expression of PLAC1 in leukemia. We investigated expression of PLAC1 in Acute Myeloid Leukemia (AML) and Acute Lymphoblastic Leukemia (ALL). In this study, we investigated expression pattern of PLAC1 gene in peripheral blood and bone marrow mononuclear cells of newly-diagnosed patients with AML (n=31) and ALL (n=31) using quantitative real-time PCR. Normal subjects (n=17) were considered as control. The PLAC1 protein expression in the samples were also detected using western blotting. Our data demonstrated that PLAC1 transcripts had 2.7 and 2.9 fold-change increase in AML and ALL, respectively, compared to normal samples. PLAC1 transcript expression was totally negative in all studied normal subjects. Level of PLAC1 mRNA expression in ALL statistically increased compared to normal samples (p=0.038). However, relative mRNA expression of PLAC1 in AML was not significant in comparison to normal subjects (p=0.848). Furthermore, relative mRNA expression of PLAC1 in AML subtypes was not statistically significant (p=0.756). PLAC1 gene expression showed no difference in demographical clinical and para-clinical parameters. Western blotting confirmed expression of PLAC1 in the ALL and AML samples. Considering PLAC1 expression profile in acute leukemia, PLAC1 could be a potential marker in leukemia which needs complementary studies in the future.

Bacteroides fragilis expresses three proteins similar to Porphyromonas gingivalis HmuY: Hemophore-like proteins differentially evolved to participate in heme acquisition in oral and gut microbiomes.

Oral and gut microbiomes are important for the maintenance of homeostasis in the human body. Altered or disturbed mutualism between their members results in dysbiosis with local injury and subsequent systemic diseases. The high bacterial density causes intense competition among microbiome residents to acquire nutrients, including iron and heme, the latter of high importance for heme auxotrophic members of the Bacteroidetes phylum. Our main hypothesis is that the heme acquisition mechanism, with the leading role played by a novel HmuY family of hemophore-like proteins, can be used to fulfill nutritional requirements and increase virulence. We characterized HmuY homologs expressed by Bacteroides fragilis and compared their properties with the first representative of this family, the HmuY protein of Porphyromonas gingivalis. In contrast to other Bacteroidetes members, B. fragilis produces three HmuY homologs (Bfr proteins). All bfr transcripts were produced at higher levels in bacteria starved of iron and heme (fold change increase ~60, ~90, and ~70 for bfrA, bfrB, and bfrC, respectively). X-ray protein crystallography showed that B. fragilis Bfr proteins are structurally similar to P. gingivalis HmuY and to other homologs, except for differences in the potential heme-binding pockets. BfrA binds heme, mesoheme, and deuteroheme, but preferentially under reducing conditions, using Met175 and Met146 to coordinate heme iron. BfrB binds iron-free protoporphyrin IX and coproporphyrin III, whereas BfrC does not bind porphyrins. HmuY is capable of heme sequestration from BfrA, which might increase the ability of P. gingivalis to cause dysbiosis also in the gut microbiome.

Abstract OT2-08-03: Immunogenicity of pembrolizumab and doxorubicin in a phase I trial for patients with metastatic triple negative breast cancer

Abstract Background: Combination of immune checkpoint inhibitor pembrolizumab and chemotherapy are standard of care therapy for patients with programmed death-ligand 1 positive (PD-L1+) metastatic triple negative breast cancer (mTNBC). However, a greater understanding of how immune checkpoint inhibitors and chemotherapies synergize to yield anti-tumor T cell responses is needed. The current phase I study evaluated the immunogenicity of doxorubicin plus pembrolizumab in patients with mTNBC. Patients and Methods: Patients with mTNBC, no prior anthracycline use, and 0-2 lines of prior systemic chemotherapies received pembrolizumab 200 mg IV and doxorubicin 50-60 mg/m2 IV every 3 weeks for 6 cycles followed by pembrolizumab maintenance until disease progression or intolerance. Patients were not selected based on PD-L1 expression. The primary objectives were safety and objective response rate per RECIST 1.1. Peripheral blood samples were collected at baseline, Cycle 2 Day 1 (C2D1), and post Cycle 3 for analysis by high parameter flow cytometry. Results: Ten patients were enrolled between March 2016 and November 2019. Best responses included one patient with complete response (CR), five with partial responses (PR), two with stable disease (SD), and one with progression of disease (PD). Flow cytometry showed increased CD3+ T cells (p=0.03) from pre-treatment to C2D1 with substantial inter-patient variation in CD3+ T cells. The expansion of proliferative exhausted-like PD-1+ CD8+ T cell population was identified in 8/9 patients, and exhausted CD8+ T cells were significantly expanded from pre-treatment to C2D1 in the patient with CR (p=0.01). Notably, frequencies of PD-1 high proliferative CD8+ T cells contracted significantly from C2D1 to post Cycle 3 (p=0.005), with a return to near baseline frequencies in the majority of the patients. In the context of all identified T cell subsets, PD-1hi proliferative CD8+ T cells demonstrated the greatest increase in fold change from pre-treatment to C2D1. In contrast, PD-1lo proliferative CD8+ T cells demonstrated the greatest decrease in fold change from pre-treatment to C3D1. Conclusion: Anthracycline-naïve patients with mTNBC treated with the combination of pembrolizumab and doxorubicin yielded robust peripheral blood T cell responses. Further studies dissecting the dynamics and durability of anti-tumor T cell responses and concurrent tumor immune microenvironment changes are needed to optimize combined immune checkpoint inhibitor and chemotherapy treatment for patients with mTNBC. Citation Format: Colt A. Egelston, Weihua Guo, Susan E. Yost, Xuan Ge, Jin Sun Lee, Paul H. Frankel, Yujie Cui, Christopher Ruel, Daniel Schmolze, Mireya Murga, Aileen Tang, Norma Martinez, Misagh Karimi, George Somlo, Peter Lee, James Waisman, Yuan Yuan. Immunogenicity of pembrolizumab and doxorubicin in a phase I trial for patients with metastatic triple negative breast cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr OT2-08-03.

The Antimelanoma Biological Assessment of Triterpenic Acid Functionalized Gold Nanoparticles.

One of several promising strategies for increasing the bioavailability and therapeutic potential of high-lipophilic biologically active compounds is gold nanoparticle formulation. The current study describes the synthesis and biological antimelanoma evaluation of three triterpen-functionalized gold nanoparticles, obtained using our previously reported antimelanoma benzotriazole-triterpenic acid esters. Functionalized gold nanoparticle (GNP) formation was validated through UV-VIS and FTIR spectroscopy. The conjugate's cytotoxic effects were investigated using HaCaT healthy keratinocytes and A375 human melanoma cells. On A375 cells, all three conjugates demonstrated dose-dependent cytotoxic activity, but no significant cytotoxic effects were observed on normal HaCaT keratinocytes. GNP-conjugates were found to be more cytotoxic than their parent compounds. After treatment with all three GNP-conjugates, 4,6'-diamidino-2-phenylindole (DAPI) staining revealed morphological changes consistent with apoptosis in A375 melanoma cells. Quantitative real-time polymerase chain reaction (RT-qPCR) analysis revealed that the triterpene-GNP conjugate treated A375 melanoma cells had a fold change increase in Bcl-2-associated X protein (BAX) expression and a fold change decrease in B-cell lymphoma 2 (Bcl-2) expression. In A735 melanoma cells, high-resolution respirometry studies revealed that all three GNP-conjugates act as selective inhibitors of mitochondrial function. Furthermore, by examining the effect on each mitochondrial respiratory rate, the results indicate that all three conjugates are capable of increasing the production of reactive oxygen species (ROS), an apoptosis trigger in cancer cells.

Evaluation and Comparison of Plasma miRNA-31 in Oral Squamous Cell Carcinoma

Background/Purpose: Oral Squamous cell carcinoma is sixth most common cancer with considerable morbidity and mortality. The microRNAs (miRNAs) are set of short RNAs involved in regulating the expression of protein coding genes. They are up or down-regulated in carcinogenesis and in oral cancer. The miRNA-31 (miR-31) is increased in oral cancer. Objective: To evaluate and compare the expression of miRNA-31 in plasma of Oral squamous cell carcinoma and control subjects. Materials and Methods: Case control study was carried out in 25 cases of oral squamous cell carcinoma subjects and 25 normal control subjects. The level of miRNA-31 in blood plasma was evaluated by miRNA easy kit (quagen) and miRNA-based qRT-PCR. The fold change was observed and compared between OSCC and controls. Results: The plasma level of miRNA-31 was significantly increased in OSCC patients compared to controls (p&lt;0.001). The patients with moderately differentiated, grade 4 OSCC patients showed significant increase in fold change compared to control, well differentiated and grade 3 OSCC (p&lt;0.001). Conclusion: Our results indicate that plasma miR-31 may be used as an adjuvant biomarker the detection of OSCC patient.