Tumor Model Research Articles

MCM4 Promotes the Progression of Malignant Melanoma by Activating the PI3K/AKT Pathway.

This study aims to elucidate the role of minichromosome maintenance protein 4 (MCM4) in malignant melanoma (MM) and explore the underlying mechanism. Initially, data from The Cancer Genome Atlas (TCGA) database and the Molecular Signature Database (MSigDB) were used to investigate the biological impact of MCM4 on MM. Further, a prognostic model using Cox regression analysis was developed to predict the overall survival (OS) rate in the MM patients. The effects of MCM4 on the proliferation, migration, and invasion abilities of MM (B16F0 and A375) cells were demonstrated using the CCK-8, colony formation, EDU, wound scratch, and Transwell assays. In subcutaneous tumor models in C57BL/6 mice invivo, the expression levels of MCM4 in MM cells and tumors were detected using Western blot and immunofluorescence approaches. The bioinformatics analysis indicated that MCM4 was expressed higher in MM tissues than in the normal tissues (p < 0.05). The established OS prediction model could significantly contribute to devising follow-up strategies and treating MM patients. MCM4 knockdown resulted in reduced proliferation, migration, and invasion abilities of MM cells, which were reversed by MCM4 overexpression (p < 0.05). Moreover, MCM4 could activate the phosphatidylinositol 3'-kinase (PI3K)/AKT pathway in MM cells. The PI3K inhibitor (LY294002) could reverse the effects of MCM4 on MM cells. MCM4 could substantially prompt the tumor growth of MM in mice through the PI3K/AKT pathway invivo. In summary, MCM4 prompted the development and metastasis of MM by activating the PI3K/AKT pathway.

NEK2 Promotes ESCC Malignant Progression by Inhibiting Cellular Senescence via the FOXM1/c-Myc/p27 Signaling Pathway.

Never in mitosis gene A (NIMA)-related kinase 2 (NEK2) is a crucial serine-threonine kinase involved in the process of cell mitosis. However, the precise relationship between NEK2 and esophageal squamous cell carcinoma (ESCC) remains inadequately understood. NEK2 expression in ESCC tissues was assessed through bioinformatics analysis, reverse transcription-quantitative PCR (RT-qPCR) and immunohistochemistry, revealing a correlation with ESCC patient prognosis. Cultured ESCC cells and human normal esophageal epithelial cells (HEEC) were used to investigate the effects of NEK2 knockdown on the development and progression of ESCC by integrated confluence algorithm, colony formation, wound-healing, transwell, and ESCC xenograft tumor model, in vitro and in vivo. In ESCC tissues, NEK2 was found to be significantly upregulated, and its expression correlated with poor prognosis in ESCC patients. NEK2 may facilitate ESCC development by regulating cell proliferation, migration, and invasion. Additionally, results from in vivo experiments suggested that NEK2 knockdown can inhibit tumor growth. Moreover, forkhead box M1 (FOXM1) was identified as a potential downstream target of NEK2 in the regulation of ESCC, with its overexpression reversing the effects of NEK2 knockdown on ESCC. Mechanistic studies also indicated that NEK2 may promote the malignant progression of ESCC by inhibiting cellular senescence through the activation of the FOXM1/c-Myc/p27 signaling pathways, which may provide a novel perspective for the management of ESCC.

Specifically Editing Cancer Sialoglycans for Enhanced In Vivo Immunotherapy through Aptamer‐Enzyme Chimeras

AbstractImmune checkpoint blockade (ICB) therapies have demonstrated remarkable clinical success in treating cancer. However, their objective response rate remains suboptimal because current therapies rely on limited immune checkpoints that fail to cover the multiple immune evasion pathways of cancer. To explore potential ICB strategies, we propose a glycoimmune checkpoint elimination (glycoICE) therapy based on targeted editing of sialoglycans on the tumor cell surface using an aptamer‐enzyme chimera (ApEC). The ApEC can be readily generated via a one‐step bioorthogonal procedure, allowing for large‐scale and uniform production. It specifically targets and desialylates cancer cells, disrupting the sialoglycan‐Siglec axis to activate immune cells and enhance immunotherapy efficacy, while its high tumor selectivity minimizes side effects from indiscriminate desialylation of normal tissues. Furthermore, the ApEC has the potential to be a versatile platform for specific editing of sialoglycans in different tumor models by adjusting the aptamer sequences to target specific protein markers. This research not only introduces a novel molecular tool for the effective editing of sialoglycans in complex environments, but also provides valuable insights for advancing DNA‐based drugs towards in vivo and clinical applications.

Development and validation of a controlled heating apparatus for long‐term MRI of 3D microfluidic tumor models

AbstractConventional testing of novel contrast agents for magnetic resonance imaging (MRI) involves cell and animal studies. However, 2D cultures lack dynamic flow and in vivo MRI is limited by regulatory approval of long‐term anesthesia use. Microfluidic tumor models (MTMs) offer a cost‐effective, reproducible, and high throughput platform for bridging cell and animal models. Yet, MRI of microfluidic devices is challenging, due to small fluid volumes generating low sensitivity. For the first time, an MRI of MTMs was performed at low field strength (1 T) using conventional imaging equipment without microcoils. To enable longitudinal MRI, we developed (1) CHAMP‐3 (controlled heating apparatus for microfluidics and portability) which heats MTMs during MRI scans and (2) an MRI‐compatible temperature monitoring system. CHAMP‐3 maintained chip surface temperature at ~37°C and the media inside at ~35.5°C. Enhanced T1‐weighted MRI contrast was achieved in 3D MTMs with free manganese (Mn2+) solutions and Mn2+ labeled tumor cells.

Patient-Derived Organoids on a Microarray for Drug Resistance Study in Breast Cancer.

Drug resistance is always a challenge in cancer treatment, whether for chemotherapy, targeting, or immunotherapy. Although tumor cell lines are derived from cancer patients, they gradually lost the original characteristics, including heterogeneity and tumor microenvironment (TME), during the long period of in vitro culturing. Therefore, it is urgent to use patient-derived tumor models instead of cancer cell lines to study tumor drug resistance. Herein, we developed a microarray device that serves as a platform for high-throughput and three-dimensional culture of breast cancer patient-derived organoids (BCOs) and investigated their resistance to adriamycin (ADM). Coupled with fluorescence microscopy, this system enabled on-chip drug response monitoring and cell viability assessment without the consumption of a large number of tumor cells. The organoids were divided into a resistant BCO group (RBCO) and a sensitive BCO group (SBCO) according to their half-inhibitory concentration (IC50). Different from cancer cell lines, BCOs demonstrated obvious heterogeneity in drug treatment. Ivermectin (IVM), a broad-spectrum antiparasitic agent approved by the Food and Drug Administration (FDA), was observed to synergistically augment ADM-induced cytotoxicity in organoids. The BCO chip provides a promising platform for investigation of drug resistance and preclinical drug screening based on clinical samples.

Dendritic cell maturation is induced by p53-armed oncolytic adenovirus via tumor-derived exosomes enhancing systemic antitumor immunity.

Dendritic cells (DCs) are crucial in cancer immunity, because they activate cytotoxic T cells by presenting tumor antigens. Recently, oncolytic virus therapy has been recognized as a systemic immune stimulator. We previously developed a telomerase-specific oncolytic adenovirus (OBP-301) and a p53-armed OBP-301 (OBP-702), demonstrating that these viruses strongly activate systemic antitumor immunity. However, their effects on DCs remained unclear. In the present study, the aim was to elucidate the mechanisms of DC activation by OBP-702, focusing particularly on tumor-derived exosomes. Exosomes (Exo53, Exo301, or Exo702) were isolated from conditioned media of human or murine pancreatic cancer cell lines (Panc-1, MiaPaCa-2, and PAN02) after treatment with Ad-p53, OBP-301, or OBP-702. Exo702 derived from Panc-1 and MiaPaCa-2 cells significantly upregulated CD86, CD80, CD83 (markers of DC maturation), and IFN-γ in DCs in vitro. Similarly, Exo702 derived from PAN02 cells upregulated CD86 and IFN-γ in bone marrow-derived DCs in a bilateral PAN02 subcutaneous tumor model. This DC maturation was inhibited by GW4869, an inhibitor of exosome release, and anti-CD63, an antibody targeting the exosome marker. Intratumoral injection of OBP-702 into PAN02 subcutaneous tumors significantly increased the presence of mature DCs and CD8-positive T cells in draining lymph nodes, leading to long-lasting antitumor effects through the durable activation of systemic antitumor immunity. In conclusion, tumor-derived exosomes play a significant role in DC maturation following OBP-702 treatment and are critical for the systemic activation of antitumor immunity, leading to the abscopal effect.

A bispecific CD40 agonistic antibody allowing for antibody-peptide conjugate formation to enable cancer-specific peptide delivery, resulting in improved T proliferation and anti-tumor immunity in mice

Current antibody-based immunotherapy depends on tumor antigen shedding for proper T cell priming. Here we select a novel human CD40 agonistic drug candidate and generate a bispecific antibody, herein named BiA9*2_HF, that allows for rapid antibody-peptide conjugate formation. The format is designed to facilitate peptide antigen delivery to CD40 expressing cells combined with simultaneous CD40 agonistic activity. In vivo, the selected bispecific antibody BiA9*2_HF loaded with peptide cargos induces improved antigen-specific proliferation of CD8+ (10-15 fold) and CD4+ T cells (2-7 fold) over control in draining lymph nodes. In both virus-induced and neoantigen-based mouse tumor models, BiA9*2_HF demonstrates therapeutic efficacy and elevated safety profile, with complete tumor clearance, as well as measured abscopal impact on tumor growth. The BiA9*2_HF drug candidate can thus be utilized to tailor immunotherapeutics for cancer patients.

Evaluation of a Radioiodinated G-Quadruplex Binder in Cervical Cancer Models.

We herein describe the radiosynthesis of a 125I-labeled acridine orange derivative ([125I]-C8), acting as a G-quadruplex binder, and its biological evaluation in cervical cancer models, aiming to enlighten its potential as a radioligand for Auger Electron Radiopharmaceutical Therapy (AE-RPT) of cancer. [125I]-C8 was synthesized with a moderate radiochemical yield (ca. 60 %) by a [125I]iodo-destannylation reaction. Its evaluation in cervical cancer HeLa cells demonstrated that the radiocompound has a significant cellular internalization with a notorious accumulation in the cell nucleus. In line with these results, [125I]-C8 strongly compromised the viability of HeLa cells in a dose-dependent manner, inducing non-repairable DNA lesions that are most probably due to the AEs emitted by 125I in close proximity to the DNA molecule. Biodistribution studies in a murine HeLa xenograft model showed that [125I]-C8 has fast blood clearance and high in vivo stability but poor tumor uptake, after systemic administration. The respective supramolecular conjugate with the AS1411 aptamer ([125I]-C8/AS1411) led to a slower blood clearance in the same animal tumor model, although without improving the tumor uptake. To take advantage of the radiotoxicity of [125I]-C8 against cervical cancer cells other strategies need to be studied, based namely on alternative nanodelivery carriers and/or intratumoral injection approaches.

Controlling the Redox Chemistry of Cobalt Radiopharmaceuticals.

The elementally matched 55Co2+/3+ (t1/2=17.53 h, Iβ+=77 %)/58mCo2+/3+ (t1/2=9.10 h, internal conversion=100 %) radioisotope pair is of interest for development of paired diagnostic/therapeutic radiopharmaceuticals. Due to the accessibility of the nat/55Co2+/3+ redox couple, the redox state can be readily modulated. Here, we show that macroscopic and radiochemical redox reactions can be closely monitored and controlled using spectroscopic and radiochemical methods. We employ model systems to inform how to selectively synthesize thermodynamically favored oxidation state coordination complexes. In addition to exogenous oxidants, our data indicates that 55Co-induced radiolysis of water efficiently and directly drives selective oxidation to the 55Co3+ species under no-carrier added (n.c.a.) conditions. Our synthetic strategies subsequently stabilize the respective 55Co2+ or 55Co3+ species for targeted positron emission tomography imaging in a mouse tumor model.

IMPA1-derived inositol maintains stemness in castration-resistant prostate cancer via IMPDH2 activation.

Acquisition of prostate cancer stem cells (PCSCs) manifested during androgen ablation therapy (ABT) contributes to castration-resistant prostate cancer (CRPC). However, little is known about the specific metabolites critically orchestrating this process. Here, we show that IMPA1-derived inositol enriched in PCSCs is a key metabolite crucially maintaining PCSCs for CRPC progression and ABT resistance. Notably, conditional Impa1 knockout in the prostate abrogates the pool and properties of PCSCs to orchestrate CRPC progression and prolong the survival of TRAMP mice. IMPA1-derived inositol serves as a cofactor that directly binds to and activates IMPDH2, which synthesizes guanylate nucleotides for maintaining PCSCs with ARlow/- features leading to CRPC progression and ABT resistance. IMPA1/inositol/IMPDH2 axis is upregulated in human prostate cancer, and its overexpression predicts poor survival outcomes. Genetically and pharmacologically targeting the IMPA1/inositol/IMPDH2 axis abrogates CRPC and overcomes ABT resistance in various CRPC xenografts, patient-derived xenograft (PDX) tumor models, and TRAMP mouse models. Our study identifies IMPDH2 as an inositol sensor whose activation by inositol represents a key mechanism for maintaining PCSCs for CRPC and ABT resistance.

Treatment of Conjunctival Melanoma Cell Lines With a Light-Activated Virus-Like Drug Conjugate Induces Immunogenic Cell Death.

Conjunctival melanoma (CJM) is a rare malignant ocular surface tumor, which often leads to local recurrences and metastases. In murine models of subcutaneous tumors, treatment with a novel virus-like drug conjugate (VDC; Bel-sar) showed a dual mechanism of action with direct tumor cell killing as well as stimulation of an antitumoral immune response. Bel-sar is currently being evaluated for the treatment of primary uveal melanoma and indeterminate nevi in a phase III clinical trial. We determined whether Bel-sar also has direct antitumor efficiency and a potential immunostimulatory capacity in CJM cells. Three human tumor-derived CJM lines were used. Bel-sar's subcellular and intracellular locations were determined with tracers. Following light activation of Bel-sar, cytotoxicity and exposure of damage-associated molecular patterns (DAMPs) were assessed. Treated tumor cells were co-cultured with THP-1 derived macrophages to assess tumor-cell phagocytosis. Bel-sar was bound and internalized by CJM cells and subsequently found in the cell membrane, lysosome, Golgi apparatus, and mitochondria. Bel-sar activation induced near complete cell death with half-maximal inhibitory concentration (IC50) values between 30 pM and 60 pM. Finally, light-activated Bel-sar enhanced exposure of DAMPs, including calreticulin, heat shock protein 90, and stimulated phagocytosis by macrophages. Treatment with a novel VDC (Bel-sar) induced pro-immunogenic cell death in all three CJM cell lines. The in vitro cytotoxicity was accompanied by exposure of DAMPs, suggesting Bel-sar is a potential treatment for CJM by a dual mechanism of action. This dual mechanism may provide a targeted and direct killing of tumor cells and induce an immune response which might decrease local recurrences and metastasis.

Leveraging long-acting IL-15 agonists for intratumoral delivery and enhanced antimetastatic activity

IntroductionIL-15 agonists hold promise as immunotherapeutics due to their ability to induce the proliferation and expansion of cytotoxic immune cells including natural killer (NK) and CD8+ T cells. However, they generally have short half-lives that necessitate frequent administration to achieve efficacy. To address this limitation, we have developed a half-life extension technology using hydrogel microspheres (MS). Here, the therapeutic is tethered to MSs by a releasable linker with pre-programed cleavage rates. We previously showed the MS conjugate of single-chain IL-15, MS~IL-15, effectively increased the half-life of IL-15 to approximately 1 week and enhanced the pharmacodynamics. We sought to determine whether the same would be true with a MS conjugate of the IL-15 agonist, receptor-linker IL-15 (RLI).MethodsWe prepared a long acting MS conjugate of RLI, MS~RLI. The pharmacokinetics and pharmacodynamics of MS~RLI were measured in C57BL/6J mice and compared to MS~IL-15. The antitumor efficacy of MS~RLI was measured when delivered subcutaneously or intratumorally in the CT26 tumor model and intratumorally in the orthotopic EO771 tumor model.ResultsMS~RLI exhibited a half-life of 30 h, longer than most IL-15 agonists but shorter than MS~IL-15. The shorter than expected half-life of MS~RLI was shown to be due to target-mediated-disposition caused by an IL-15 induced cytokine sink. MS~RLI resulted in very potent stimulation of NK and CD44hiCD8+ T cells, but also caused significant injection-site toxicity that may preclude subcutaneous administration. We thus pivoted our efforts toward studying the MS~RLI for long-acting intra-tumoral therapy, where some degree of necrosis might be beneficial. When delivered intra- tumorally, both MS~IL-15 and MS~RLI had modest anti-tumor efficacy, but high anti- metastatic activity.ConclusionIntra-tumoral MS~RLI and MS~RLI combined with systemic treatment with other agents could provide beneficial antitumor and anti-metastatic effects without the toxic effects of systemic IL-15 agonists. Our findings demonstrate that intra-tumorally administered long-acting IL-15 agonists counter two criticisms of loco-regional therapy: the necessity for frequent injections and the challenge of managing metastases.

Minimalist Adjuvant-Free Nano-Vaccine Based on Antigen Self-Assembled Amyloid-Like Fibrils to Induce Potent Immune Response.

The development of cancer vaccines is at the forefront of cancer immunotherapy. Most existing strategies to induce an efficient anti-tumor immune response rely on molecular adjuvants and the incorporation of complex synthetic vectors into vaccine formulations. In contrast, this study introduces a one-step engineering technique to assemble the model antigen, Ovalbumin (OVA), into amyloid aggregates, leveraging biomimetic folding and aggregation to create non-fibrillar OVA globular aggregates and OVA amyloid-like fibrils as single-component, adjuvant-free vaccines. Notably, the OVA amyloid-like fibrils induced stronger immune responses compared to the native form, as evidenced by robust humoral immune reactions and the establishment of immune memory. These enhanced responses can be attributed to the self-adjuvant effect of the unique assembled structure, which preserves antigenic epitopes, improves antigen stability, facilitates antigen internalization, prolongs retention at the injection site, enhances antigen trafficking to the lymphoid organs, and promotes increased secretion of antibodies and cytokines. Furthermore, the efficacy of the vaccine was validated in a high OVA-expressing tumor model, demonstrating the potential of OVA amyloid-like fibrils as an effective vaccine for cancer immunoprevention. This minimalist self-adjuvant vaccine strategy holds promising implications for cancer immunotherapy and can inform the design of other protein antigen-based vaccines.

Injectable nanocomposite hydrogel with cascade drug release for treatment of uveal melanoma

Uveal melanoma (UM) is the most common malignant intraocular tumor with the trait of distant metastases. Currently, the standard clinical therapy results in suboptimal outcomes due to ineffective inhibition of tumor metastasis. Thus, developing novel therapeutic modalities for UM remains a critical priority. Herein, we have developed an injectable nanocomposite hydrogel (HA-DOX/LAP gel) through integrating hyaluronic acid-based drug-loaded nanoparticles into an alginate-dopamine gel, delivering the chemotherapeutic drugs, lapatinib and doxorubicin for combinational treatment of UM. HA-DOX/LAP gel is fabricated in situ by a simple injection of the mixed precursor solution into tumor sites and maintains in vivo for more than 21 days. The entrapped drug-loaded nanoparticles can gradually release from HA-DOX/LAP gel, enhancing tumor targeting and penetration, and synchronously releasing lapatinib and doxorubicin into the acidic intracellular environment to synergistically destroy UM cells. In vivo anti-tumor studies conducted in MuM-2B tumor models demonstrated that HA-DOX/LAP gel significantly impedes tumor growth, diminishes postoperative recurrence, and prolongs overall survivals of UM tumor-bearing mice through only single injection. Remarkably, the escaped drug-loaded nanoparticles effectively reduce the risk of tumor metastases. Our findings provide new insights for the development of multifunctional nanocomposite-incorporating combination therapy against UM by targeting tumor recurrence and metastases.

ST2L promotes VEGFA-mediated angiogenesis in gastric cancer by activating TRAF6/PI3K/Akt/NF-κB pathway via IL-33

Suppression of Tumorigenicity 2 (ST2) is a member of the interleukin-1 receptor/ Toll-like receptor superfamily, and its specific ligand is Interleukin-33 (IL-33). IL-33/ ST2 signaling has been implicated in numerous inflammatory and allergic diseases, as well as in promoting malignant behavior of tumor cells and angiogenesis. However, the precise role of ST2 in gastric cancer angiogenesis remains incompletely elucidated. We observed a significant correlation between high expression of ST2 in gastric cancer tissues and poor prognosis, along with various clinicopathological features. In vitro experiments demonstrated that the IL-33/ ST2 axis activates the PI3K/AKT/NF-κB signaling pathway through TRAF6, thereby promoting VEGFA-mediated tumor angiogenesis; meanwhile sST2 acts as a decoy receptor to regulate the IL-33/ST2L axis. Consistent findings were also observed in subcutaneous xenograft tumor models in nude mice. Furthermore, we investigated the molecular mechanism by which IL-33 promotes ST2L expression in GC cells via upregulation of transcription factors YY1 and GATA2 through intracellular signaling pathways.

Ly6E on tumor cells impairs anti-tumor T-cell responses: a novel mechanism of tumor-induced immune exclusion

BackgroundLymphocyte antigen 6 complex, locus E (Ly6E) has been initially demonstrated to involve in T cell activity and impair viral infectivity. Recently, high expression levels of Ly6E have been reported in tumor microenvironment (TME) of various types of cancers. However, the immunoregulatory mechanism of Ly6E manipulating TME remains unknown.MethodsTCGA database and Kaplan–Meier plotter database were used to evaluate the correlation between Ly6E expression levels and cancer patient survival. After analyzing Ly6E expression levels in human breast cancer tissues and tumor cell lines, we generated Ly6E knockout (KO) and overexpression (OE) mouse cell lines. Cell proliferation ability in vitro and the ability of growth and metastasis in mouse tumor models were compared between KO/OE and wild-type tumor cells. On day 7 after tumor implantation, tumor tissues were separated for flow cytometric assay, bulk RNA sequencing and single-cell RNA sequencing (ScRNA-seq). The role of Ly6E-expressing tumor cell on macrophage was analyzed in vitro.ResultsOur result surprisingly found that high Ly6E expression levels were associated with CD8+ T cell exclusion in tumor tissues and resistance to immunotherapy. Our data showed that knockout of Ly6E in tumor cells prompts tumor regression and inhibits tumor metastases, and Ly6E-OE tumor cells vice versa. The enhanced anti-tumor effect of Ly6E knockout in tumor cells was dependent on T cell response and formed long-lasting memory. The increase in the CD8+ T-cell infiltration into the tumor islet of Ly6E-KO tumors confirmed the role of Ly6E on T cell exclusion. ScRNA-seq analysis showed that M2 macrophages are particularly abundant in the Ly6E-expressing tumor tissues, especially M2-4 macrophage cluster identified by high levels of Arg-1, indicates that Ly6E-expressing tumor cells might restrict T cell infiltration via M2 macrophages. Moreover, in vitro assay showed that cell culture media derived from Ly6E-positive tumor cells promoted macrophage migration and M2 polarization.ConclusionOur study illuminated that Ly6E-expressing tumor cells facilitated the accumulation of M2 macrophages in TME, which contributes to CD8+ T cell exclusion and provides new insights for improving efficacy of cancer immunotherapy.

Spatial context of immune checkpoints as predictors of overall survival in patients with resectable colorectal cancer independent of standard TNM stages.

While immune checkpoint blockade (ICB) therapy has shown promising results in a small subset of colorectal cancer patients with high microsatellite instability (MSI-H), the majority of patients with colorectal cancer do not respond to ICB therapy. The main obstacle to the success of immunotherapy in cancer treatment is the exhaustion of tumor-infiltrating lymphocytes (TILs). Elucidating the spatial organization of immune checkpoints within the tumor microenvironment could pave the way for the development of novel prognostic tools and therapeutic strategies to enhance antitumor immune responses. To clarify the spatial and functional diversity of tumor-infiltrating lymphocytes (TILs) in the colorectal tumor microenvironment (TME), we performed multiplexed IHC to examine the exhaustion of TILs in the TME (the expression of PD-1 and TIM-3 (T-cell immunoglobulin and mucin-domain-containing protein 3), which are major biomarkers of T-cell exhaustion) and Lasso-Cox analyses of the correlation between CRC prognosis and TME features. For proof of concept, the antitumor efficacy of TIM-3 and PD-1 dual blockade in CRC was further evaluated in a CT26 subcutaneous tumor model of human CRC. We found that the spatial context of PD-1 and TIM-3 successfully predicted the overall survival of CRC patients independent of TNM stage. Dual targeting of PD-1 and TIM-3 in mouse tumor models inhibited tumor progression and reduced T-cell exhaustion, indicating a potential strategy for improving the clinical treatment of CRC.

Activity-Based Dicyanoisophorone Derivatives: Fluorogenic Toolbox Enables Direct Visualization and Monitoring of Esterase Activity in Tumor Models.

The visualization and spatiotemporal monitoring of endogenous esterase activity are crucial for clinical diagnostics and treatment of liver diseases. Our research adopts a novel substrate hydrolysis-enzymatic activity (SHEA) approach using dicyanoisophorone-based fluorogenic ester substrates DCIP-R (R = R1-R6) to evaluate esterase preferences on diverse substrate libraries. Esterase-mediated hydrolysis yielded fluorescent DCIP-OH with a nanomolar detection limit in vitro. These probes effectively monitor ester hydrolysis kinetics with a turnover number of 4.73 s-1 and catalytic efficiency (kcat/Km) of 106 M-1 s-1 (DCIP-R1). Comparative studies utilizing two-photon imaging have indicated that substrates containing alkyl groups (DCIP-R1) as recognition elements exhibit enhanced enzymatic cleavage compared to those containing phenyl substitution on alkyl chains (DCIP-R4). Time-dependent variations in endogenous esterase levels were tracked in healthy and liver tumor models, especially in diethylnitrosamine (DEN)-induced tumors and HepG2-transplanted liver tumors. Overall, fluorescence signal quantifications demonstrated the excellent proficiency of DCIP-R1 in detecting esterase activity both in vitro and in vivo, showing promising potential for biomedical applications.

Acoustically tunable intra-droplet assembly of organoids towards high-throughput tumor model construction

Three-dimensional cell organoids and spheroids are increasingly recognized as physiologically relevant models for ex vivo research and therapeutic screening. However, there remains a need for a cost-effective and convenient platform to fabricate these models with well-defined architectures for downstream applications. Here, this work proposes micro models constructing in the droplet platform, by acoustic forces, that can obtain 3D cellular organoids with tunable structures at high yield. By modulating signals applied to acoustic devices, a periodic potential array with adjustable structures is formed, enabling uniform trapping and patterning of droplets. Simultaneously, the localized potential is also formed inside droplets, resulting in the active assembly of cells. Then, hydrogel droplets containing the constructed models are crosslinked, providing ECM to mimic the 3D scaffolds. In experiment, MCF-7 cells are assembled into spherical, linear and cross structures. This method can generate thousands of tumor spheroids in one minute. The long-term culturing results indicate an enhanced efficiency in forming mature tumor models compared with control groups. This biomanufacturing technology has the advantages of being high-throughput, low-cost, and tunable, making it a robust tool for droplet-based applications such as tissue engineering, drug screening, and disease modeling etc.

CDK4/6 inhibitors potently inhibit cell viability in pre-clinical models of adult-type ovarian granulosa cell tumors

CDK4/6 inhibitors potently inhibit cell viability in pre-clinical models of adult-type ovarian granulosa cell tumors