Stimulated Cell Proliferation Research Articles

Delivery of AKT1 phospho-forms to human cells reveals differential substrate selectivity.

Protein kinase B (AKT1) is a serine/threonine kinase that regulates fundamental cellular processes, including cell survival, proliferation, and metabolism. AKT1 activity is controlled by two regulatory phosphorylation sites (Thr308, Ser473) that stimulate a downstream signaling cascade through phosphorylation of many target proteins. At either or both regulatory sites, hyperphosphorylation is associated with poor survival outcomes in many human cancers. Our previous biochemical and chemoproteomic studies showed that the phosphorylated forms of AKT1 have differential selectivity toward peptide substrates. Here, we investigated AKT1-dependent activity in human cells, using a cell-penetrating peptide (transactivator of transcription, TAT) to deliver inactive AKT1 or active phospho-variants to cells. We used enzyme engineering and genetic code expansion relying on a phosphoseryl-transfer RNA (tRNA) synthetase (SepRS) and tRNASep pair to produce TAT-tagged AKT1 with programmed phosphorylation at one or both key regulatory sites. We found that all TAT-tagged AKT1 variants were efficiently delivered into human embryonic kidney (HEK 293T) cells and that only the phosphorylated AKT1 (pAKT1) variants stimulated downstream signaling. All TAT-pAKT1 variants induced glycogen synthase kinase (GSK)-3α phosphorylation, as well as phosphorylation of ribosomal protein S6 at Ser240/244, demonstrating stimulation of downstream AKT1 signaling. Fascinatingly, only the AKT1 variants phosphorylated at S473 (TAT-pAKT1S473 or TAT-pAKT1T308,S473) were able to increase phospho-GSK-3β levels. Although each TAT-pAKT1 variant significantly stimulated cell proliferation, cells transduced with TAT-pAKT1T308 grew significantly faster than with the other pAKT1 variants. The data demonstrate differential activity of the AKT1 phospho-forms in modulating downstream signaling and proliferation in human cells.

Features of Intracellular Signal Transduction in Neural Stem Cells under the Influence of Alkaloid Songorine, an Agonist of Fibroblast Growth Factor Receptors.

We performed a comparative in vitro study of the involvement of NF-κB, PI3K, cAMP, ERK1/2, p38, JAKs, STAT3, JNK, and p53-dependent intracellular signaling in the functioning of neural stem cells (NSC) under the influence of basic fibroblast growth factor (FGF) and FGF receptor agonist, diterpene alkaloid songorine. The significant differences in FGFR-mediated intracellular signaling in NSC were revealed for these ligands. In both cases, stimulation of progenitor cell proliferation occurs with the participation of NF-κB, PI3K, ERK1/2, JAKs, and STAT3, while JNK and p53, on the contrary, inhibit cell cycle progression. However, under the influence of songorin, cAMP- and p38-mediated cascades are additionally involved in the transmission of the NSC division-activating signal. In addition, unlike FGF, the alkaloid stimulates progenitor cell differentiation by activating ERK1/2, p38, JNK, p53, and STAT3.

High histone crotonylation modification in bovine fibroblasts promotes cell proliferation and the developmental efficiency of preimplantation nuclear transfer embryos

Lysine crotonylation (Kcr) is a recently discovered histone acylation modification that is closely associated with gene expression, cell proliferation, and the maintenance of stem cell pluripotency and indicates the transcriptional activity of genes and the regulation of various biological processes. During cell culture, the introduction of exogenous croconic acid disodium salt (Nacr) has been shown to modulate intracellular Kcr levels. Although research on Kcr has increased, its role in cell growth and proliferation and its potential regulatory mechanisms remain unclear compared to those of histone methylation and acetylation. Our investigation demonstrated that the addition of 5 mM Nacr to cultured bovine fibroblasts increased the expression of genes associated with Kcr modification, ultimately promoting cell growth and stimulating cell proliferation. Somatic cell nuclear transfer of donor cells cultured in 5 mM Nacr resulted in 38.1% blastocyst development, which was significantly greater than that in the control group (25.2%). This research is important for elucidating the crotonylation modification mechanism in fibroblast proliferation to promote the efficacy of somatic cell nuclear transfer.

Effects of Capsaicin on Migration and Alkaline Phosphatase Activity of Dental Pulp Cells.

Dental pulp, a specialized mesenchymal tissue within teeth, is pivotal in dental health and tissue repair. Capsaicin, the primary pungent component of chili peppers, is known for its diverse pharmacological properties. While capsaicin's effects on various cell types have been studied, its impact on dental pulp cells remains relatively unexplored. This study investigated the influence of pure capsaicin extract on dental pulp cell behavior, focusing on cell viability, proliferation, migration, and alkaline phosphatase (ALP) activity. Capsaicin solution was prepared and diluted to various concentrations (1 nM, 0.01 µM, 0.1 µM, 1 µM, 10 µM, and 100 µM), then was tested on rat dental pulp cells (RPC-C2A). Cell viability and proliferation were assessed using the MTT assay. Boyden chamber tests and wound healing were used for evaluating cell migration. The activity of ALP was determined to show cell function during dental pulp repair. The data were analyzed using a one-way analysis of variance or an independent-sample Kruskal-Wallis, followed by multiple comparison tests. Capsaicin of 100 µM exhibited cytotoxicity, whereas those with lower concentrations stimulated cell proliferation. Wound healing assays revealed increased cell migration, particularly when cultured with 1 nM capsaicin (p = 0.002). Boyden chamber assays demonstrated enhanced cell invasion without statistical significance. ALP activity of dental pulp cells increased significantly at 1 nM (p < 0.001) and 1 µM (p = 0.021) capsaicin concentrations, indicating potential dentinogenesis and pulp repair. Capsaicin of lower concentrations, less than 10 µM, is likely to promote proliferation, migration, and ALP activity of dental pulp cells. Our findings offer potential applications for capsaicin as a medication for dental pulp repair.

CD133 Stimulates Cell Proliferation via the Upregulation of Amphiregulin in Melanoma.

CD133, a cancer stem cell (CSC) marker in tumors, including melanoma, is associated with tumor recurrence, chemoresistance, and metastasis. Patient-derived melanoma cell lines were transduced with a Tet-on vector expressing CD133, generating doxycycline (Dox)-inducible cell lines. Cells were exposed to Dox for 24 h to induce CD133 expression, followed by RNA-seq and bioinformatic analyses, revealing genes and pathways that are significantly up- or downregulated by CD133. The most significantly upregulated gene after CD133 was amphiregulin (AREG), validated by qRT-PCR and immunoblot analyses. Induced CD133 expression significantly increased cell growth, percentage of cells in S-phase, BrdU incorporation into nascent DNA, and PCNA levels, indicating that CD133 stimulates cell proliferation. CD133 induction also activated EGFR and the MAPK pathway. Potential mechanisms highlighting the role(s) of CD133 and AREG in melanoma CSC were further delineated using AREG/EGFR inhibitors or siRNA knockdown of AREG mRNA. Treatment with the EGFR inhibitor gefitinib blocked CD133-induced cell growth increase and MAPK pathway activation. Importantly, siRNA knockdown of AREG reversed the stimulatory effects of CD133 on cell growth, indicating that AREG mediates the effects of CD133 on cell proliferation, thus serving as an attractive target for novel combinatorial therapeutics in melanoma and cancers with overexpression of both CD133 and AREG.

FANCA facilitates G1/S cell cycle advancement, proliferation, migration and invasion in gastric cancer.

The present study explores the function of FANCA gene, a pivotal member of the Fanconi anaemia (FA) pathway crucial for preserving genomic stability and preventing cancer, particularly in the context of gastric cancer (GC). Using immunohistochemistry, quantitative real-time PCR, and western blot analysis, we evaluate FANCA mRNA and protein expressions in GC cell lines. The relationship between FANCA expression and clinicopathological characteristics is also explored. Various assays, including CCK8, colony formation, wound healing, and Transwell assays, are used to assess functional changes in cells associated with FANCA. Flow cytometry is utilized to evaluate alterations in the cell cycle resulted from FANCA knockdown and overexpression. Our findings show elevated FANCA expression in GC cell lines, with levels correlated with pathologic stage and lymphatic metastasis. FANCA knockdown impedes cell proliferation, migration, and invasion and induces G1/S phase cell cycle arrest. Conversely, FANCA overexpression stimulates cell proliferation, migration, and invasion. In vivo xenograft experiments confirm the promotional role of FANCA in GC tumor progression. Moreover, FANCA overexpression is associated with the activation of cell cycle. Collectively, our results suggest that FANCA drives malignant cell behaviors in GC through the cell cycle pathway, highlighting its potential as a therapeutic target for the treatment of GC.

Food for thought: The effects of feeding on neurogenesis in the ball python, Python regius.

Introduction Pythons are a well-studied model of postprandial physiological plasticity. Consuming a meal evokes a suite of physiological changes in pythons including one of the largest documented increases in post-feeding metabolic rates relative to resting values. However, little is known about how this plasticity manifests in the brain. Previous work has shown that cell proliferation in the python brain increases six days following meal consumption. This study aimed to confirm these findings and build on them in the long term by tracking the survival and maturation of these newly created cells across a two-month period. Methods We investigated differences in neural cell proliferation in ball pythons six days after a meal with immunofluorescence using the cell-birth marker 5-bromo-12'-deoxyuridine (BrdU). We investigated differences in neural cell maturation in ball pythons two months after a meal using double-immunofluorescence for BrdU and a reptilian ortholog of the neuronal marker Fox3. Results We did not find significantly greater rates of cell proliferation in snakes six days after feeding, but we did observe more new cells in neurogenic regions in fed snakes two months after the meal. Feeding was not associated with higher rates of neurogenesis, but snakes that received a meal had higher numbers of newly created non-neuronal cells than fasted controls. We documented particularly high cell survival rates in the olfactory bulbs and lateral cortex. Conclusions Consuming a meal stimulates cell proliferation in the brains of ball pythons after digestion is complete, although this effect emerged at a later time point in this study than expected. Higher rates of proliferation partially account for greater numbers of newly created non-neuronal cells in the brains of fed snakes two months after the meal, but our results also suggest that feeding may have a mild neuroprotective effect. We captured a slight trend toward higher cell survival rates in fed snakes, and survival rates were particularly high in brain regions associated with olfactory perception and processing. These findings shed light on the relationship between energy balance and the creation of new neural cells in the brains of ball pythons.

Understanding the mechanism of ulcerative colitis-induced colorectal cancer and revealing the potential of Dillenia Indica in its management

Ulcerative colitis (UC) is a chronic inflammatory bowel disease (IBD) that significantly increases the risk of developing colorectal cancer (CRC). The overall prevalence of UC-induced CRC is on the rise due to the rapidly increasing frequency of UC. It is widely acknowledged that chronic inflammation that persists for a long period is a significant contributing factor to the development of CRC in individuals with UC. The activation of various proinflammatory pathways, such as nuclear factor kappa B (NF-κB), interleukin-6/signal transducer and activator of transcription 3 (IL-6/STAT3), cyclooxygenase-2/prostaglandin E2 (COX-2/PGE2), and interleukin-23/T-helper 17 (IL-23/Th17), and gut microbiota contributes to the initiation and progression of tumorigenesis by a different mechanism, such as triggering the production of inflammatory mediators, increasing the expression of antiapoptotic genes, and stimulating cell proliferation as well as angiogenesis. Managing this risk and finding potential nutraceutical interventions, such as Dillenia indica (DI), is of significant interest. This review delves into the literature concerning the mechanism correlated to the development of CRC in UC individuals and investigates the potential of DI, as a nutraceutical in managing this rapid risk of UC-CRC. By examining the different mechanisms involved in UC-induced CRC, we aim to provide insight into the therapeutic promise of DI in mitigating these mechanisms.

Transcription factor DREF regulates expression of the microRNA gene bantam in Drosophila melanogaster.

The bantam gene encodes a vital microRNA and has a complex expression pattern in various tissues at different stages of Drosophila development. This microRNA is involved in the control of normal development of the ocular and wing imaginal discs, the central nervous system, and also in maintaining the undifferentiated state of stem cells in the ovaries of adult females. At the cellular level, bantam stimulates cell proliferation and prevents apoptosis. The bantam gene is a target of several conserved signaling cascades, in particular, Hippo. At the moment, at least ten proteins are known to directly regulate the expression of this gene in different tissues of Drosophila. In this study, we found that the bantam regulatory region contains motifs characteristic of binding sites for DREF, a transcription factor that regulates the expression of Hippo cascade genes. Using transgenic lines containing a full-length bantam lethality-rescuing deletion fragment and a fragment with a disrupted DREF binding site, we show that these motifs are functionally significant because their disruption at the bantam locus reduces expression levels in the larvae and ovaries of homozygous flies, which correlates with reduced vitality and fertility. The effect of DREF binding to the promoter region of the bantam gene on its expression level suggests an additional level of complexity in the regulation of expression of this microRNA. A decrease in the number of eggs laid and a shortening of the reproductive period in females when the DREF binding site in the regulatory region of the bantam gene is disrupted suggests that, through bantam, DREF is also involved in the regulation of Drosophila oogenesis.

Gastric Cancer Mesenchymal Stem Cells Trigger Endothelial Cell Functional Changes to Promote Cancer Progression.

Our previous studies have highlighted the pivotal role of gastric cancer mesenchymal stem cells (GCMSCs) in tumor initiation, progression, and metastasis. In parallel, it is well-documented that endothelial cells (ECs) undergo functional alterations in response to challenging tumor microenvironment. This study aims to elucidate whether functional changes in ECs might be induced by GCMSCs and thus influence cancer progression. Cell proliferation was assessed through CCK-8 and colony formation assays, while cell migration and invasion capabilities were evaluated by wound-healing and Transwell assays. Immunohistochemistry was employed to examine protein distribution and expression levels. Additionally, quantitative analysis of protein and mRNA expression was carried out through Western blotting and qRT-PCR respectively, with gene knockdown achieved using siRNA. Our findings revealed that GCMSCs effectively stimulate cell proliferation, migration, and angiogenesis of human umbilical vein endothelial cells (HUVECs), both in vitro and in vivo. GCMSCs promote the migration and invasion of gastric cancer cells by inducing the expression of Slit2 in HUVECs. Notably, the inhibition of phosphorylated AKT partially mitigates the aforementioned effects. In conclusion, GCMSCs may exert regulatory control over Slit2 expression in ECs via the AKT signaling pathway, thereby inducing functional changes in ECs that promote tumor progression.

Conductive Hyaluronic Acid/Deep Eutectic Solvent Composite Hydrogel as a Wound Dressing for Promoting Skin Burn Healing Under Electrical Stimulation.

Burns can cause severe damage to the skin due to bacterial infection and severe inflammation. Although conductive hydrogels as electroactive burn-wound dressings achieve remarkable effects on accelerating wound healing, issues such as imbalance between their high conductivity and mechanical properties, easy dehydration, and low transparency must be addressed. Herein, a double-network conductive eutectogel is fabricated by integrating polymerizable deep eutectic solvents (PDESs)including acrylamide/choline chloride/glycerol (acrylamide-polymerization crosslink) and thiolated hyaluronic acid (disulfide-bonding crosslink). The introduction of PDESs provides the eutectogel with a conductivity (up to 0.25 S·m-1) and mechanical strength (tensile strain of 59-77%) simulating those of natural human skin, as well as satisfactory tissue adhesiveness, self-healing ability, and antibacterial properties. When combined with exogenous electrical stimulation, the conductive eutectogel exhibits the ability to reduce inflammation, stimulate cell proliferation and migration, promote collagen deposition and angiogenesis, and facilitate skin tissue remodeling. This conductive eutectogel shows great potential as a dressing for healing major burn wounds.

Use of vegetable oils in wound treatment

This study addresses the use of vegetable oils in wound treatment. These oils contain bioactive compounds that confer analgesic, antiseptic, soothing, healing, and emollient properties. Notably, candeia oil is recognized for its anti-inflammatory and antimicrobial properties, while grape seed oil is rich in proanthocyanidins with potent antioxidant activity. Lavender oil, in addition to stimulating cell proliferation and neovascularization, possesses antioxidant and antibacterial properties. Pumpkin seed oil, on the other hand, is rich in fatty acids and lipids, which help combat oxidative stress in wounds. Coconut oil, with its antibacterial and antifungal properties, and copaiba oil, with anti-inflammatory and healing actions, are also highlighted. Cucumber seed oil, besides its antioxidant properties, exhibits antimicrobial activity due to the presence of oleic acid. Lastly, tea tree oil, known for its antimicrobial action, also promotes wound healing. These oils represent a promising and natural alternative in wound treatment, although further studies are needed to determine their dosages and ensure their clinical safety.

Biocompatible Bletilla striata polysaccharide-based hemostatic sponges enhance wound healing and reduce the risk for infections

Bletilla striata polysaccharide (BSP) is an extract derived from the rare and valuable Chinese herb Bletilla striata [Thunb.] Reichb. f. Although BSP research has progressed considerably, producing portable BSP dressings with hemostatic efficacy remains challenging. This study describes a novel dressing with a reticulate structure. To create this structure, BSP sponges were cross-linked with chitosan and lyophilized. The resulting BSP sponges had excellent mechanical properties, a high density of pores, and an outstanding capacity for fluid absorption. Both in vitro and in vivo experiments demonstrated that the BSP sponges had excellent hemostatic, antibacterial, and anti-inflammatory properties; they also exhibited good stimulation of cell growth and wound healing properties. Furthermore, they were less likely to provoke complications than traditional materials. Cytocompatibility assays revealed that they also stimulated cell proliferation. Therefore, our study suggests that BSP sponges have promising clinical applications as wound dressings.

Abstract 7072: MYC acetylated lysine residues drive oncogenic cell transformation and regulate select genetic programs for cell adhesion-independent growth and survival

Abstract The MYC oncogenic transcription factor is acetylated by the p300 and GCN5 histone acetyltransferases. The significance of MYC acetylation and functions of specific acetylated lysine (K) residues have remained unclear. Here, we show that the major p300-acetylated K148(149) and K157(158) sites in human (or mouse) MYC and the main GCN5-acetylated K323 residue are reversibly acetylated in various malignant and non-malignant cells. Oncogenic overexpression of MYC enhances its acetylation and alters regulation of site-specific acetylation by proteasome and deacetylase inhibitors. Acetylation of MYC at different K residues differentially affects its stability in a cell type- dependent manner. Lysine-to-arginine substitutions indicate that although none of the acetylated K (AcK) residues is required for MYC stimulation of adherent cell proliferation, individual AcK sites have gene-specific functions controlling select MYC-regulated processes in cell adhesion, contact inhibition, apoptosis, and/or metabolism, and are required for the malignant cell transformation activity of MYC. Each AcK site is required for anchorage-independent growth of MYC-overexpressing cells in vitro, and both the AcK148(149) and AcK157(158) residues are also important for the tumorigenic activity of MYC-transformed cells in vivo. These AcK residues may also preferentially activate transcription via facilitating MYC binding to several cancer-associated target gene promoters and via enhancing RNAP II recruitment. With the knowledge that MYC lysine residues can be acetylated by HATs GCN5 and P300, some HATs and HAT associated cofactors (i.e. YEATS2 and PIN1) are important for MYC recruitment to DNA binding. By identifying MYC acetylation-dependent coregulators and their oncogenic function, we proposed to understand whether and how MYC acetylation residues cooperate with its cofactors and influence malignant transformation in breast cancer cells.Overall, this study will uncover the functions and mechanisms of MYC acetylation in regulation of malignant transformation and the impact on breast cancer initiation and progression. By analyzing deregulated target genes and MYC-relevant coactivators, the MYC AcK site-specific signaling pathways identified may offer new avenues for selective therapeutic targeting of MYC oncogenic activities. Citation Format: Jeffrey J. Pino, Matthew Hurd, Yifan Zhao, Kay Jang, Michael Allevato, Marina Vorontchikhina, Wataru Ichikawa, Ryan Gates, Emily Villalpando, Michael Hamilton, Francesco Failo, Songqin Pan, Yue Qi, Yu-wen Hung, Thomas Girke, David Ann, Victoria Seewaldt, Ernest Martinez. MYC acetylated lysine residues drive oncogenic cell transformation and regulate select genetic programs for cell adhesion-independent growth and survival [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 7072.

Abstract 5913: TEAD1/4 inhibitors exhibit deeper biological impact and broader activity compared to TEAD1-only inhibitors in both monotherapy and combination without additional kidney toxicity

Abstract The TEAD transcription factors in association with the YAP/TAZ co-activators drive the expression of pro-proliferative and pro-oncogenic genes that underlies the transformed phenotype of many carcinomas. In addition, YAP-TEAD transcriptional activity is emerging as a major resistance mechanism for diverse precision oncology drugs, with the most extensive data for resistance to drugs targeting the MAPK pathway. There is a strong interest in generating inhibitors of YAP/TEAD transcriptional activity with TEAD palmitic acid site inhibitors having demonstrated encouraging pre-clinical activity and now clinical activity with confirmed objective responses with the TEAD1/2/3 inhibitor VT3989. However, the TEAD1-preferential inhibitor IK930 did not yield any objective responses but also showed a more favorable safety profile especially with respect to proteinuria. These contrasting clinical read-outs provide a good lead-in to a critical design challenge/question of TEAD palmitic acid site inhibitors: the optimization of inhibitory profile against the respective four TEAD paralogs (TEAD1-4). In depth bioinformatic analyses led Sporos investigators to conclude that while TEAD1 inhibition was a minimum requirement for anti-neoplastic activity, inhibition of other paralogs would be necessary for maximizing biological impact and a TEAD1/4 inhibitor would provide the best balance of anti-neoplastic activity and toxicity. Activity against TEAD2 was identified as counter-productive associated with a context-specific paradoxical stimulation of cell proliferation and tumor growth while activity against TEAD3 was flagged as a major driver of podocyte effacement and kidney toxicity. Here, we provide novel corroborating data supporting this selection of inhibitory profile. We provide an update on the pre-clinical efficacy and toxicology of SPR1, Sporos’s TEAD1/4 preferential inhibitor which favorably contrasts with other TEAD inhibitors such as the TEAD1/3/4 inhibitor VT3989 and the TEAD1 inhibitor VT103 and IK930 in both the monotherapy and combination setting. We show that 1) SPR1 displays broader and deeper cell-based activity and extends the utility of TEAD inhibitors outside of mesothelioma and NF2 mutants 2) SPR1 shows stronger activity than TEAD1-only inhibitors in combination with MAPK and EGFR inhibitors in vitro and in vivo 3) SPR1 does not cause proteinuria in mice; dogs or rats even above therapeutic doses 4) SPR1 does not show the context-specific stimulation of tumor growth in Lung PDX previously observed with VT3989 and other inhibitors that include TEAD2 in their profile. Taken together - the data suggests SPR1 is positioned to become a best-in-class TEAD palmitic acid site inhibitor with broad utility in both monotherapy and combination setting. Citation Format: Florian Muller, Selvi Kunnimalaiyaan, Parth Mangrolia, Jill Olson. TEAD1/4 inhibitors exhibit deeper biological impact and broader activity compared to TEAD1-only inhibitors in both monotherapy and combination without additional kidney toxicity [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 5913.

Abstract 3305: Golcadomide (CC-99282) is a novel CELMoD® agent with antiproliferative activity and combinatorial potential in disease models of chronic lymphocytic leukemia

Abstract Chronic Lymphocytic Leukemia (CLL) is an adult leukemia characterized by the accumulation of incompetent B lymphocytes expressing CD19, CD20, CD23, and CD5. Current therapies include chemotherapy and combination with targeted therapies such as ibrutinib (Bruton’s tyrosine kinase inhibitor), venetoclax (BCL2 inhibitor), and obinutuzumab (anti-CD20). The tumor microenvironment promotes CLL development and drug resistance. Golcadomide (GOLCA, CC-99282) is an oral cereblon E3 ligase modulator (CELMoD®) agent with immunomodulatory and tumor cell-autonomous activities under clinical investigation for R/R non-Hodgkin lymphomas and CLL/SLL. Here we investigated the effect of GOLCA on inhibiting proliferation and inducing apoptosis of CLL preclinical models by degrading Ikaros and Aiolos. We also studied its combination with other anti-CLL agents. A panel of 10 CLL cell lines and 15 primary CLL patient samples were used to assess efficacy of GOLCA alone or in combination with ibrutinib (Ibru), venetoclax (Ven), and obinutuzumab (Obi). Primary CLL cells were grown ex vivo in a co-culture system with CD40L expressing fibroblasts designed to mimic the CLL lymph node microenvironment. Substrate degradation, viability, cell cycle and immunophenotyping were assessed by flow cytometry. GOLCA showed potent in vitro antiproliferative activity on 6 out of 10 CLL cell lines, including those with high-risk features, with IC50 of 1-20 nM. It also inhibited CLL stimulated cell proliferation and induced apoptosis in all evaluated patient samples in the co-culture system, with low to sub-nanomolar IC50, independent of IGHV mutation status and other chromosomal characteristics. Cell-cycle analysis confirmed inhibition of proliferation in the 5 patient samples evaluated after 3 days of treatment with GOLCA, with a dose-dependent decrease in the fraction of cells in the S phase. GOLCA also demonstrated degradation of the proximal substrates Ikaros and Aiolos in tumor and T cells from CLL patient samples. In combination with Obi, Ven, and Ibru, GOLCA led to synergistic or additive tumor cell toxicity in most patient samples evaluated, suggesting potential clinical benefit of these targeted agent combinations. Additive tumor toxicity was observed in two out of four patient samples when GOLCA was combined with Ibru. We have previously demonstrated that GOLCA induces immune activation in T cells from healthy donor PBMCs. In co-cultures of CLL PBMCs and CD40L-expressing fibroblasts, GOLCA increased T cell numbers and activation characteristics. Ven inhibited this immune activation, while Obi or Ibru maintained the immune activation. Collectively, these findings suggest that GOLCA, alone or in combination with targeted agents, may produce clinical benefit in CLL patients. Citation Format: Antonia Lopez-Girona, Maria Dolores Jimenez-Nunez, Diego Sobradillo, Soraya Carrancio, Preethi Janardhanan, Gauri Deb, Lynda Groocock, Daniel Pierce, Neil Bence, Mark Rolfe. Golcadomide (CC-99282) is a novel CELMoD® agent with antiproliferative activity and combinatorial potential in disease models of chronic lymphocytic leukemia [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 3305.

Abstract 1280: Role of the tumor microbiome in the lung adenocarcinoma immune microenvironment through multi meta-omics analysis

Abstract Lung cancer is the leading cause of death by cancer worldwide. The tumor microbiome has shed light on its role, influence, and modulation in cancer immunosurveillance, where the tumor-bacterial component emerges as a potential mediator for the antitumoral-immune response. Nevertheless, the complexities and functional mechanisms underlying this interaction remain elusive. Here, we outline how the tumor microbiota can impact the host´s immune response to lung adenocarcinoma (LUAD). We explored the microbiome within LUAD tumors and non-tumoral adjacent tissues (NAT) from Chilean patients. We used 10 paired (LUAD and NAT) fresh-frozen samples, 20 exclusively tumor fresh-frozen samples, and 40 exclusively tumor formalin-fixed paraffin-embedded samples. Using 16S rRNA gene sequencing, we assessed revealed variations in alpha diversity, with an intra-tumoral microbiome enriched in Gammaproteobacteria and Actinobacteria, particularly genera Pseudomonas and Corynebacterium in undifferentiated LUAD samples. Notably, a consistent depletion of Verrucomicrobiota with genus Akkermansia was evident in samples where cell differentiation was lost. Metaproteomics analysis revealed bacteria-associated peptides from Pseudomonas and Streptomyces enriched in the tumor, linked to Major facilitator superfamily (MFS) transporters and Glutathione S-transferase N-terminal domain-containing protein, both associated with transport activity and negatively correlated with cancer. The bacterial metabolic analysis uncovered 10 enriched microbial-associated metabolites in the tumor, highlighting adenosine and inosine, two known inducers of immunosuppression on T-cells in the tumor microenvironment. Host proteomics gene-ontology analysis highlighted an enrichment of proteins in the Interferon pathways. Concurrently, a downregulation of proteins like T-cell surface-glycoprotein CD8 and HLA-class I histocompatibility antigen was observed, linked to MHC protein complex binding function, and associated with immune evasion through diminished antigen presentation. Furthermore, upregulated proteins such as macrophage migration inhibition factor (MIF) and eIF-2-alpha kinase GCN2, known to stimulate cell proliferation, were identified. In summary, our multi-omics and bioinformatic analyses unveil a distinct intra-tumoral microbiome and microbe-derived metabolites could induce immune suppression, altering the metabolic profile in the LUAD tumor microenvironment and collectively contribute to tumor progression. Funding information: This work was supported by FONDECYT1231629 and ANID-Subdirección de capital humano/Doctorado Nacional/2023/21231386. Citation Format: Ivania Valdés, Alberto Martin, Eduardo Martínez, Daniel Carvajal Hausdorf, Erick Riquelme. Role of the tumor microbiome in the lung adenocarcinoma immune microenvironment through multi meta-omics analysis [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 1280.

Exploring a Synergistic Approach: Dual GLP-1 Agonist Combined with Degludec Basal Insulin for Early Type 1 Diabetes Treatment and its Impact on Albumin-Insulin Producing Cells Expression."

This manuscript explores various aspects related to the use of dual GLP-1 agonist with degludec basal insulin as a potential treatment approach for early type 1 diabetes. The background section highlights the destruction of beta cells in type 1 diabetes and the emergence of GLP-1 agonists as a promising option for managing obesity and type 2 diabetes. The authors discuss a retrospective analysis of the efficacy of semaglutide, a GLP-1 agonist, in patients with newly diagnosed type 1 diabetes. The results show the elimination of prandial and basal insulin, increased C-peptide levels, and improved glycemic control. However, the study's retrospective nature and lack of a control group emphasize the need for larger prospective trials. The interpretation section highlights the potential of GLP-1 agonists in protecting residual beta cells, stimulating cell proliferation, and reprogramming liver cells into insulin-producing cells. Moreover, modifying GLP-1 agonists with albumin ligands shows promise in extending their half-life and enhancing their anti-diabetic effects. The perspective section provides a comprehensive overview of the synergistic approach, considering the pharmacokinetic properties of degludec, the plasticity of adult human hepatic tissue, and the benefits of modified GLP-1 derivatives. The conclusion emphasizes the need for further research to explore the full potential of this approach in type 1 diabetes treatment. The proposed approach offers a promising avenue for the treatment of type 1 diabetes, integrating the autoimmune hypothesis, the proliferative effects of GLP-1, and modifications using albumin ligands. By combining these elements, we can strive towards restoring beta cell mass and function, ultimately improving the lives of individuals living with type 1 diabetes. The manuscript is planned to undergo clinical trials in 2024, registered as 'Amr Ahmed, Maher M. Akl, Semaglutide GLP1 Agonists with Degludec Basal-bolus Insulin in Early Type 1 Diabetes to Basal-bolus' with ClinicalTrials.gov Identifier NCT06057077.

Chromatin profiling reveals TFAP4 as a critical transcriptional regulator of bovine satellite cell differentiation

BackgroundSatellite cells are myogenic precursor cells in adult skeletal muscle and play a crucial role in skeletal muscle regeneration, maintenance, and growth. Like embryonic myoblasts, satellite cells have the ability to proliferate, differentiate, and fuse to form multinucleated myofibers. In this study, we aimed to identify additional transcription factors that control gene expression during bovine satellite cell proliferation and differentiation.ResultsUsing chromatin immunoprecipitation followed by sequencing, we identified 56,973 and 54,470 genomic regions marked with both the histone modifications H3K4me1 and H3K27ac, which were considered active enhancers, and 50,956 and 59,174 genomic regions marked with H3K27me3, which were considered repressed enhancers, in proliferating and differentiating bovine satellite cells, respectively. In addition, we identified 1,216 and 1,171 super-enhancers in proliferating and differentiating bovine satellite cells, respectively. Analyzing these enhancers showed that in proliferating bovine satellite cells, active enhancers were associated with genes stimulating cell proliferation or inhibiting myoblast differentiation whereas repressed enhancers were associated with genes essential for myoblast differentiation, and that in differentiating satellite cells, active enhancers were associated with genes essential for myoblast differentiation or muscle contraction whereas repressed enhancers were associated with genes stimulating cell proliferation or inhibiting myoblast differentiation. Active enhancers in proliferating bovine satellite cells were enriched with binding sites for many transcription factors such as MYF5 and the AP-1 family transcription factors; active enhancers in differentiating bovine satellite cells were enriched with binding sites for many transcription factors such as MYOG and TFAP4; and repressed enhancers in both proliferating and differentiating bovine satellite cells were enriched with binding sites for NF-kB, ZEB-1, and several other transcription factors. The role of TFAP4 in satellite cell or myoblast differentiation was previously unknown, and through gene knockdown and overexpression, we experimentally validated a critical role for TFAP4 in the differentiation and fusion of bovine satellite cells into myofibers.ConclusionsSatellite cell proliferation and differentiation are controlled by many transcription factors such as AP-1, TFAP4, NF-kB, and ZEB-1 whose roles in these processes were previously unknown in addition to those transcription factors such as MYF5 and MYOG whose roles in these processes are widely known.

Beyond Traditional Boundaries: A Review on Calotropis gigantea’s Efficacy in Wound Healing

Introduction: Wounds result from the disruption of cellular and anatomical continuity in living tissues due to external factors. Calotropis gigantea, also known as crown flower or giant milkweed (or "biduri" in Indonesia), is a small tree or shrub with white or purple flowers, widely used in traditional remedies. Its properties, including antimicrobial, cytotoxic, and wound-healing activities, have prompted this literature review to assess its efficacy in wound healing. Method: A thorough analysis of scientific findings and research conducted in the last 10 years, specifically after January 1, 2014, was undertaken. Data were collected from reputable databases, such as PubMed, Web of Science, and ScienceDirect. After applying publication limitations, three manuscripts were selected as the final outcomes of the review process. Result: The analysis of multiple studies reveals that C. gigantea possesses wound healing properties attributed to phytochemical constituents like flavonoids, tannins, alkaloids, and saponin. These constituents play vital roles in each stage of wound healing, including the inflammatory reaction, proliferation, and remodeling stages. Additionally, C. gigantea promotes wound healing, leaving minimal scar residues. Conclusion: Calotropis gigantea has demonstrated its ability to enhance wound healing through various mechanisms, including astringency, antibacterial activity, promotion of homeostasis, facilitation of collagen synthesis, enhancement of re-epithelialization, and stimulation of cell proliferation via its phytochemical constituents. However, due to the scarcity of recent studies, the plant's true efficacy in wound healing remains unclear.