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OBJECTIVE: Chemoresistant-epithelial ovarian cancer (EOC) has a poor prognosis, prompting the search for new therapeutic drugs. The diphenylbutylpiperidine (DPBP) class of antipsychotic drugs used in schizophrenia has shown anticancer effects. This study aimed to investigate the preclinical efficacy of penfluridol, fluspirilene, and pimozide (DPBP) using in vitro and in vivo models of EOC. METHODS: Human EOC cell lines A2780, HeyA8, SKOV3ip1, A2780-CP20, HeyA8-MDR, and SKOV3-TR were treated with penfluridol, fluspirilene, and pimozide, and cell proliferation, apoptosis, and migration were assessed. The preclinical efficacy of DPBP was also investigated using in vivo mouse models, including cell lines and patient-derived xenografts (PDX) of EOC. RESULTS: DPBP drugs significantly decreased cell proliferation in chemosensitive (A2780, HeyA8, and SKOV3ip1) and chemoresistant (A2780-CP20, HeyA8-MDR, and SKOV3-TR) cell lines. Among these drugs, penfluridol exerted a relatively stronger cytotoxic effect on all cell lines. Penfluridol significantly increased apoptosis and inhibited migration of EOC cells. In the cell line xenograft mouse model with HeyA8, the penfluridol group showed significantly decreased tumor weight compared with the control group. In the paclitaxel-resistant model with HeyA8-MDR, the penfluridol group had significantly decreased tumor weight compared with the paclitaxel or control groups. Penfluridol exerted anticancer effects on the PDX model. CONCLUSION: Penfluridol exerted significant anticancer effects on EOC cells and xenograft models, including PDX. Thus, penfluridol therapy, as a drug repurposing strategy, might be a potential therapeutic for EOCs.

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α9-nAChR, a subtype of nicotinic acetylcholine receptor, is significantly overexpressed in female breast cancer tumor tissues compared to normal tissues. Previous studies have proposed that specific single nucleotide polymorphisms (SNPs) in the CHRNA9 (α9-nAChR) gene are associated with an increased risk of breast cancer in interaction with smoking. The study conducted a breast cancer risk assessment of the α9-nAChR SNP rs10009228 (NM_017581.4:c.1325A > G) in the Taiwanese female population, including 308 breast cancer patients and 198 healthy controls revealed that individuals with the heterozygous A/G or A/A wild genotype have an increased susceptibility to developing breast cancer in the presence of smoking compared to carriers of the G/G variant genotype. Our investigation confirmed the presence of this missense variation, resulting in an alteration of the amino acid sequence from asparagine (N442) to serine (S442) to facilitate phosphorylation within the α9-nAchR protein. Additionally, overexpression of N442 (A/A) in breast cancer cells significantly enhanced cell survival, migration, and cancer stemness compared to S442 (G/G). Four-line triple-negative breast cancer patient-derived xenograft (TNBC-PDX) models with distinct α9-nAChR rs10009228 SNP genotypes (A/A, A/G, G/G) further demonstrated that chronic nicotine exposure accelerated tumor growth through sustained activation of the α9-nAChR downstream oncogenic AKT/ERK/STAT3 pathway, particularly in individuals with the A/G or A/A genotype. Collectively, our study established the links between genetic variations in α9-nAChR and smoking exposure in promoting breast tumor development. This emphasizes the need to consider gene-environment interactions carefully while developing effective breast cancer prevention and treatment strategies.

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BACKGROUND: Patient-derived organoids (PDOs) are multi-cellular cultures with specific three-dimensional (3D) structures. Tumor organoids (TOs) offer a personalized perspective for assessing treatment response. However, the presence of normal organoid (NO) residuals poses a potential threat to their utility for personalized medicine. There is a crucial need for an effective platform capable of distinguishing between TO and NO in cancer organoid cultures. RESULTS: We introduced a whole-mount (WM) preparation protocol for in-situ visualization of the lipidomic distribution of organoids. To assess the efficacy of this method, nine breast cancer organoids (BCOs) and six normal breast organoids (NBOs) were analyzed. Poly-l-lysine (PLL) coated slides, equipped with 12 well chambers, were utilized as a carrier for the high-throughput analysis of PDOs. Optimizing the fixation time to 30 min, preserved the integrity of organoids and the fidelity of lipid compounds. The PDOs derived from the same organoid lines exhibited similar lipidomic profiles. BCOs and NBOs were obviously distinguished based on their lipidomic signatures detected by WM autofocusing (AF) scanning microprobe matrix-assisted laser desorption/ionization (SMALDI) mass spectrometry imaging (MSI). SIGNIFICANCE: A whole-mount (WM) preparation protocol was developed to visualize lipidomic distributions of the organoids' surface. Using poly-l-lysine coated slides for high-throughput analysis, the method preserved organoid integrity and distinguished breast cancer organoids (BCOs) from normal breast organoids (NBOs) based on their unique lipidomic profiles using autofocusing scanning microprobe matrix-assisted laser desorption/ionization (SMALDI) mass spectrometry imaging.

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Breast cancer represents the most prevalent tumor type worldwide, with hormone-responsive breast cancer the most common subtype. Despite the effectiveness of endocrine therapy, advanced disease forms represent an unmet clinical need. While drug combination therapies remain promising, differences in pharmacokinetic profiles result in suboptimal ratios of free drugs reaching tumors. We identified a synergistic combination of bisdemethoxycurcumin and exemestane through drug screening and rationally designed star-shaped poly-L-glutamic acid-based combination conjugates carrying these drugs conjugated through pH-responsive linkers for hormone-responsive breast cancer treatment. We synthesized/characterized single and combination conjugates with synergistic drug ratios/loadings. Physicochemical characterization/drug release kinetics studies suggested that lower drug loading prompted a less compact conjugate conformation that supported optimal release. Screening in monolayer and spheroid breast cancer cell cultures revealed that combination conjugates possessed enhanced cytotoxicity/synergism compared to physical mixtures of single-drug conjugates/free drugs; moreover, a combination conjugate with the lowest drug loading outperformed remaining conjugates. This candidate inhibited proliferation-associated signaling, reduced inflammatory chemokine/exosome levels, and promoted autophagy in spheroids; furthermore, it outperformed a physical mixture of single-drug conjugates/free drugs regarding cytotoxicity in patient-derived breast cancer organoids. Our findings highlight the importance of rational design and advanced in vitro models for the selection of polypeptide-based combination conjugates.

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BACKGROUND: mTORC1 activity is dependent on the presence of micronutrients, including Asparagine (Asn), to promote anabolic cell signaling in many cancers. We hypothesized that targeting Asn metabolism would inhibit tumor growth by reducing mTORC1 activity in well-differentiated (WD)/dedifferentiated (DD) liposarcoma (LPS). METHODS: Human tumor metabolomic analysis was utilized to compare abundance of Asn in WD vs. DD LPS. Gene set enrichment analysis (GSEA) compared relative expression among metabolic pathways upregulated in DD vs. WD LPS. Proliferation assays were performed for LPS cell lines and organoid models by using the combination treatment of electron transport chain (ETC) inhibitors with Asn-free media. 13C-Glucose-labeling metabolomics evaluated the effects of combination treatment on nucleotide synthesis. Murine xenograft models were used to assess the effects of ETC inhibition combined with PEGylated L-Asparaginase (PEG-Asnase) on tumor growth and mTORC1 signaling. RESULTS: Asn was enriched in DD LPS compared to WD LPS. GSEA indicated that mTORC1 signaling was upregulated in DD LPS. Within available LPS cell lines and organoid models, the combination of ETC inhibition with Asn-free media resulted in reduced cell proliferation. Combination treatment inhibited nucleotide synthesis and promoted cell cycle arrest. In vivo, the combination of ETC inhibition with PEG-Asnase restricted tumor growth. CONCLUSIONS: Asn enrichment and mTORC1 upregulation are important factors contributing to WD/DD LPS tumor progression. Effective targeting strategies require limiting access to extracellular Asn and inhibition of de novo synthesis mechanisms. The combination of PEG-Asnase with ETC inhibition is an effective therapy to restrict tumor growth in WD/DD LPS.

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STUDY QUESTION: Can a functional in vitro model, containing the main cellular components of the uterine wall, be generated from cells derived from patient tissues? SUMMARY ANSWER: We present a three-dimensional (3D) physiologically relevant, organ-on-a-chip model of the uterine wall containing primary endometrial and myometrial cellular participants, generated from human uterine tissue. WHAT IS KNOWN ALREADY: As a highly dynamic reproductive organ, the human uterus plays fundamental physiological roles in menstruation and childbirth. The endometrial-myometrial junction (EMJ) defines the interface between the inner mucosal layer (endometrium) and outer smooth muscle zone (myometrium) that comprises the uterine wall. The EMJ is implicit in several uterine pathologies of unknown aetiology, including adenomyosis and abnormally invasive placenta; however, despite this, no patient-derived in vitro models of the uterine wall containing all EMJ participants currently exist. STUDY DESIGN, SIZE, DURATION: We employed microfluidic technology to characterize multiple miniaturized models of the uterine wall. Protocols were tested that included variations in the seeding order of endometrial and myometrial fractions, and the addition of a low viscosity extracellular matrix to influence cell behaviour. Ultimately, functional hormone responses of patient-derived uterine wall models were assessed. PARTICIPANTS/MATERIALS, SETTING, METHODS: Endometrial (n = 9) and myometrial biopsies (n = 4) were enzymatically dissociated to create epithelial, stromal and myometrial cellular fractions. Cell suspensions were seeded into non-adhesive poly(dimethylsiloxane) microfluidic devices containing 5 × 5 microwell arrays. The fate of individual cell types was monitored in real-time using fluorescent tracers, and cell phenotype was characterized by immunocytochemistry. Model functionality was assessed by measuring Ca2+ responses to agonist stimulation, and both insulin-like growth factor binding protein 1 (IGFBP-1) and osteopontin secretion in response to hormone stimulation. MAIN RESULTS AND THE ROLE OF CHANCE: When subjected to microfluidic culture in isolation, endometrial stromal cells and smooth muscle myocytes formed compact spheroids, whilst epithelial cells produced diffuse aggregates. Tri-cultures were established by sequential seeding of individual or combined cell fractions at various ratios. Regardless of the protocol, epithelial cells localized to the outer periphery of tri-culture spheroids, which varied in morphology across the protocols. Incorporation of 5% [v/v] Matrigel® improved the reproducibility of 3D aggregates which exhibited robust self-assembly of a stromal/smooth muscle core encased in epithelium. Exposure of tri-cultures to oestradiol, medroxyprogesterone acetate and cyclic adenosine monophosphate (cAMP) increased secretion of IGFBP-1, which indicates stromal decidualization, and enhanced epithelial cell osteopontin secretion. Stimulation with endothelin-1 induced Ca2+ signalling in myocytes. LIMITATIONS, REASONS FOR CAUTION: Endometrial and myometrial tissue was collected from relatively few donors. Myometrial tissue was collected from pregnant donors, which may have influenced the myocyte phenotype. Furthermore, endometrial tissue sampling was from women not having a hysterectomy, thus may not include the deeper basalis region, which may limit the physiological mimicry of the final models. WIDER IMPLICATIONS OF THE FINDINGS: Our novel approach to modelling the uterine wall in 3D captures all of the main cell types in a medium-throughput system, enabling the screening of hundreds of cultures in parallel from a single biopsy. This system shows great promise for examining the cellular interplay between physiological cues and EMJ pathologies, such as the impact of uterine peristalsis and cyclical hormones on the pathogenesis of adenomyosis. STUDY FUNDING/COMPETING INTEREST(S): C.B. was supported by an Organ-on-a-Chip Technologies Network Pump Priming Project grant. C.J.H. was supported by a Wellbeing of Women project grant (RG2137), SRI/Bayer and Wellcome Trust IFFS3. D.K.H. was supported by a Wellbeing of Women project grant (RG2137) and MRC clinical research training fellowship (MR/V007238/1). M.Z. is Director and Co-Founder of ScreenIn3D Limited. The other authors declare no conflict of interest. TRIAL REGISTRATION NUMBER: N/A.

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Benign prostatic hyperplasia (BPH) is a common condition marked by the enlargement of the prostate gland, which often leads to significant urinary symptoms and a decreased quality of life. The development of clinically relevant animal models is crucial for understanding the pathophysiology of BPH and improving treatment options. This study aims to establish a patient-derived xenograft (PDX) model using benign prostatic tissues to explore the molecular and cellular mechanisms of BPH. PDXs were generated by implanting fresh BPH (transition zone) and paired normal (peripheral zone) prostate tissue from 8 patients under the renal capsule of immunodeficient male mice. Tissue weight, architecture, cellular proliferation, apoptosis, prostate-specific marker expression, and molecular profiles of PDXs were assessed after 1 week and 1, 2, or 3 months of implantation by immunohistochemistry, enzyme-linked immunosorbent assay, transcriptomics, and proteomics. Responses to finasteride, a standard-of-care therapy, were evaluated. PDXs maintained histologic and molecular characteristics of the parental human tissues. BPH, but not normal PDXs, demonstrated significant increases in weight and cellular proliferation, particularly at 1 month. Molecular profiling revealed specific gene and protein expression patterns correlating with BPH pathophysiology. Specifically, an increased immune and stress response was observed at 1 week, followed by increased expression of proliferation markers and BPH-specific stromal signaling molecules, such as BMP5 and CXCL13, at 1 month. Graft stabilization to preimplant characteristics was apparent between 2 and 3 months. Treatment with finasteride reduced proliferation, increased apoptosis, and induced morphologic changes consistent with therapeutic responses observed in human BPH. Our PDX model recapitulates the morphologic, histologic, and molecular features of human BPH, offering a significant advancement in modeling the complex interactions of cell types in BPH microenvironments. These PDXs respond to therapeutic intervention as expected, providing a valuable tool for preclinical testing of new therapeutics that will improve the well-being of BPH patients.

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BACKGROUND: High-grade serous ovarian carcinoma (HGSOC) is the most common and aggressive subtype of epithelial ovarian carcinoma. It is primarily diagnosed at stage III or IV when the 5-year survival rate ranges between 20% and 40%. Here, we aimed to validate the hypothesis, based on HGSOC cell lines, that proposed the existence of two distinct groups of HGSOC cells with high and low oxidative phosphorylation (OXPHOS) metabolism, respectively, which are associated with their responses to glucose and glutamine withdrawal. METHODS: We isolated and cultivated primary cancer cell cultures from HGSOC and nontransformed ovarian fibroblasts from the surrounding ovarium of 45 HGSOC patients. We tested the metabolic flexibility of the primary cells, particularly in response to glucose and glutamine depletion, analyzed and modulated endoplasmic reticulum stress, and searched for indices of the existence of previously reported groups of HGSOC cells with high and low OXPHOS metabolism. RESULTS: The primary HGSOC cells did not form two groups with high and low OXPHOS that responded differently to glucose and glutamine availabilities in the cell culture medium. Instead, they exhibited a continuum of OXPHOS phenotypes. In most tumor cell isolates, the responses to glucose or glutamine withdrawal were mild and surprisingly correlated with those of nontransformed ovarian fibroblasts from the same patients. The growth of tumor-derived cells in the absence of glucose was positively correlated with the lipid trafficking regulator FABP4 and was negatively correlated with the expression levels of HK2 and HK1. The correlations between the expression of electron transport chain (ETC) proteins and the oxygen consumption rates or extracellular acidification rates were weak. ER stress markers were strongly expressed in all the analyzed tumors. ER stress was further potentiated by tunicamycin but not by the recently proposed ER stress inducers based on copper(II)-phenanthroline complexes. ER stress modulation increased autophagy in tumor cell isolates but not in nontransformed ovarian fibroblasts. CONCLUSIONS: Analysis of the metabolism of primary HGSOC cells rejects the previously proposed hypothesis that there are distinct groups of HGSOC cells with high and low OXPHOS metabolism that respond differently to glutamine or glucose withdrawal and are characterized by ETC protein levels.

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INTRODUCTION: Colorectal cancer (CRC) remains one of the leading causes of cancer-related morbidity and mortality worldwide. This disease is complex and heterogeneous, influenced by a variety of genetic, epigenetic, and environmental factors that drive CRC initiation and progression. Despite advances in therapeutic strategies, the five-year survival rate for metastatic CRC is alarmingly low. Traditional two-dimensional (2D) cell culture systems have been the foundation of cancer research, but their inability to replicate the complex tumor microenvironment (TME) limits their effectiveness. AREAS COVERED: This paper explores the evolution of CRC models, starting with the limitations of traditional 2D cell culture systems and the significant advancements offered by 3D models. Additionally, it highlights 3D bioprinting and on-chip CRC models, which have enhanced the ability to mimic in vivo conditions. EXPERT OPINION: The transition to advanced 3D models represents a pivotal shift in CRC research, offering considerable improvements over the established 2D models. These models hold promise for the development of patient-specific models that better mimic in vivo conditions. However, the inherent complexity of CRC continues to pose challenges in developing models that can fully capture the disease's multifaceted nature. This complexity and high costs associated with these technologies, along with the need for standardized protocols, pose significant challenges to their widespread adoption.

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Malignant transformation (MT) is commonly seen in IDH-mutant gliomas. There has been a growing research interest in revealing its underlying mechanisms and intervening prior to MT at the early stages of the transforming process. Here we established a unique pair of matched 3D cell models: 403L, derived from a low-grade glioma (LGG), and 403H, derived from a high-grade glioma (HGG), by utilizing IDH-mutant astrocytoma samples from the same patient when the tumor was diagnosed as WHO grade 2 (tumor mutational burden (TMB) of 3.96/Mb) and later as grade 4 (TMB of 70.07/Mb), respectively. Both cell models were authenticated to a patient's sample retaining endogenous expression of IDH1 R132H. DNA methylation profiles of the parental tumors referred to LGG and HGG IDH-mutant glioma clusters. The immunopositivity of SOX2, NESTIN, GFAP, OLIG2, and beta 3-Tubulin suggested the multilineage potential of both models. 403H was more prompt to cell invasion and developed infiltrative HGG in vivo. The differentially expressed genes (DEGs) from the RNA sequencing analysis revealed the tumor invasion and aggressiveness related genes exclusively upregulated in the 403H model. Pathway analysis showcased an enrichment of genes associated with epithelial-mesenchymal transition (EMT) and Notch signaling pathways in 403H and 403L, respectively. Mass spectrometry-based targeted metabolomics and hyperpolarized (HP) 1-13C pyruvate in-cell NMR analyses demonstrated significant alterations in the TCA cycle and fatty acid metabolism. Citrate, glutamine, and 2-HG levels were significantly higher in 403H. To our knowledge, this is the first report describing the development of a matched pair of 3D patient-derived cell models representative of MT and temozolomide (TMZ)-induced hypermutator phenotype (HMP) in IDH-mutant glioma, providing insights into genetic and metabolic changes during MT/HMP. This novel in vitro model allows further investigation of the mechanisms of MT at the cellular level.

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Paraneoplastic leukocytosis (PNL) in genitourinary cancer, though rare, can indicate aggressive behavior and poor outcomes. It has been potentially linked to cancer expressing G-CSF and GM-CSF, along with their respective receptors, exerting an autocrine/paracrine effect. In our study, we successfully established four patient-derived xenograft (PDX) lines and related cell lines from urothelial cancer (UC), conducting next-generation sequencing (NGS) for genetic studies. UC-PDX-LN1, originating from bladder cancer, exhibited two druggable targets - HRAS and ERCC2 - responding well to chemotherapy and targeted therapy, though not to tipifarnib, an HRAS inhibitor. Transcriptome analysis post-treatment illuminated potential mechanisms, with index protein analysis confirming their anticancer pathways. Mice implanted with UC-PDX-LN1 mirrored PNL observed in the patient's original tumor. Cytokine array and RT-PCR analyses revealed high levels of G-CSF and GM-CSF in our PDX and cell lines, along with their presence in culture media and tumor cysts.Leukocytosis within small vessels in and around the tumor, associated with NETosis and thrombus formation, suggested a mechanism wherein secreted growth factors were retained, further fueling tumor growth via autocrine/paracrine signaling. Disrupting this cancer cell-NETosis-thrombosis cycle, we demonstrated that anti-neutrophil or anticoagulant interventions enhanced chemotherapy's antitumor effects or prolonged survival in mice, even though these drugs lacked direct antitumor efficacy when used independently. Clinical observations in bladder cancer patients revealed PNL in 1.61% of cases (35/2162) with associated poor prognosis. These findings propose a novel approach, advocating for the combination of anticancer/NETosis/thrombosis strategies for managing UC patients presenting with PNL in clinical settings.

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Endometrial cancer (EC) is one of the most common gynecologic malignancies, which lacking effective drugs for intractable conditions or patients unsuitable for surgeries. Recently, the patient-derived organoids (PDOs) are found feasible for cancer research and drug discoveries. Here, we established a series of PDOs from EC and performed drug repurposing screening and mechanism analysis for cancer treatment. We confirmed that the regulatory ß subunit of methionine adenosyltransferase (MAT2B) is highly correlated with malignant progression in endometrial cancer. Through drug screening on PDOs, we identify JX24120, chlorpromazine derivative, as a specific inhibitor for MAT2B, which directly binds to MAT2B (Kd = 4.724µM) and inhibits the viability of EC PDOs and canonical cell lines. Correspondingly, gene editing assessment demonstrates that JX24120 suppresses tumor growth depending on the presence of MAT2B in vivo and in vitro. Mechanistically, JX24120 induces inhibition of S-adenosylmethionine (SAMe) synthesis, leading to suppressed mTORC1 signaling, abnormal energy metabolism and protein synthesis, and eventually apoptosis. Taken together, our study offers a novel approach for drug discovery and efficacy assessment by using the PDOs model. These findings suggested that JX24120 may be a potent MAT2B inhibitor and will hopefully serve as a prospective compound for endometrial cancer therapy.

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Extracellular vesicles (EVs) have demonstrated significant potential in drug delivery and anti-tumor therapy. Despite this promising strategy, challenges such as specific targeting, EVs purification persist. In this study, a personalized nanodrug delivery platform using patient-derived tumor EVs (PT-EVs) based on a microchip is presented. The microchip integrates multiple functions, including capture, enrichment, drug loading, and elution of PT-EVs. The isolation and drug-carrying procedures are completed within a 12 h timeframe, achieving a recovery rate of 65%, significantly surpassing the conventional ultracentrifuge (UC) method. Furthermore, PT-EVs derived from patient tumor models are first utilized as natural drug carriers, capitalizing on their inherent homing ability to precisely target homologous tumors. Lenvatinib and doxorubicin (DOX), two commonly utilized drugs in the clinical treatment of hepatocellular carcinoma (HCC), are loaded into PT-EVs and delivered to a matched in vitro tumor model that recapitulates original tumors for drug susceptibility testing. As is proven, PT-EVs exhibit robust tumor cell targeting and efficient receptor-mediated cellular uptake, and the efficacy of chemotherapeutic drugs is improved significantly. These results suggest that this platform could be a valuable tool for efficient isolation of PT-EVs and personalized drug customization, particularly when working with limited clinical samples, thus supporting personalized and precision medicine.

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BACKGROUND AND OBJECTIVE: Bladder cancer (BLCa) remains a prevalent malignancy with high recurrence rates and limited treatment options. In recent years, patient-derived organoids (PDOs) have emerged as a promising platform for studying cancer biology and therapeutic responses in a personalized manner. Using drug screening, PDOs facilitate the identification of novel therapeutic agents and translational treatment strategies. Moreover, their ability to model patient-specific responses to treatments holds promise for predicting clinical outcomes and guiding treatment decisions. This exploratory review aims to investigate the potential of PDOs in advancing BLCa research and treatment, with an emphasis on translational clinical approaches. Furthermore, we analyze the feasibility of deriving PDOs from minimally invasive blood and urine samples. METHODS: In addition to exploring hypothetical applications of PDOs for predicting patient outcomes and their ability to model different stages of BLCa, we conducted a comprehensive PubMed search on already published data as well as comprehensive screening of currently ongoing trials implementing PDOs in precision medicine in cancer patients irrespective of the tumor entity. KEY FINDINGS AND LIMITATIONS: While the research on BLCa PDOs is advancing rapidly, data on both BLCa PDO research and their clinical application are scarce. Owing to this fact, a narrative review format was chosen for this publication. CONCLUSIONS AND CLINICAL IMPLICATIONS: BLCa PDOs have the potential to influence the domain of precision medicine and enhance personalized cancer treatment strategies. However, standardized protocols for PDO generation, their ideal clinical application, as well as their impact on outcomes remain to be determined. PATIENT SUMMARY: In this review, we discuss the current state and future needs for the use of patient-derived organoids, small three-dimensional avatars of tumor cells, in bladder cancer. Patient-derived bladder cancer organoids offer a more personalized approach to studying and treating bladder cancer, providing a model that closely resembles the patient's own tumor. These organoids can help researchers identify new treatment options and predict how individual patients may respond to standard therapies. By using minimally invasive samples such as blood and urine, patients can participate in research studies more easily, potentially leading to improved outcomes in bladder cancer treatment.

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BACKGROUND AND OBJECTIVE: Predicting response to therapy for each patient's tumor is critical to improving long-term outcomes for muscle-invasive bladder cancer. This study aims to establish ex vivo bladder cancer patient-derived organoid (PDO) models that are representative of patients' tumors and determine the potential efficacy of standard of care and curated experimental therapies. METHODS: Tumor material was collected prospectively from consented bladder cancer patients to generate short-term PDO models, which were screened against a panel of clinically relevant drugs in ex vivo three-dimensional culture. Multiomic profiling was utilized to validate the PDO models, establish the molecular characteristics of each tumor, and identify potential biomarkers of drug response. Gene expression (GEX) patterns between paired primary tissue and PDO samples were assessed using Spearman's rank correlation coefficients. Molecular correlates of therapy response were identified using Pearson correlation coefficients and Kruskal-Wallis tests with Dunn's post hoc pairwise comparison testing. KEY FINDINGS AND LIMITATIONS: A total of 106 tumors were collected from 97 patients, with 65 samples yielding sufficient material for complete multiomic molecular characterization and PDO screening with six to 32 drugs/combinations. Short-term PDOs faithfully represent the tumor molecular characteristics, maintain diverse cell types, and avoid shifts in GEX-based subtyping that accompany long-term PDO cultures. Utilizing an integrative approach, novel correlations between ex vivo drug responses and genomic alterations, GEX, and protein expression were identified, including a multiomic signature of gemcitabine response. The positive predictive value of ex vivo drug responses and the novel multiomic gemcitabine response signature need to be validated in future studies. CONCLUSIONS AND CLINICAL IMPLICATIONS: Short-term PDO cultures retain the molecular characteristics of tumor tissue and avoid shifts in expression-based subtyping that have plagued long-term cultures. Integration of multiomic profiling and ex vivo drug screening data identifies potential predictive biomarkers, including a novel signature of gemcitabine response. PATIENT SUMMARY: Better models are needed to predict patient response to therapy in bladder cancer. We developed a platform that uses short-term culture to best mimic each patient's tumor and assess potential sensitivity to therapeutics.

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BACKGROUND: To explore the most effective adjuvant chemotherapy regimen for malignant peritoneal mesothelioma (MPM) through patient derived tumor-like cell clusters (PTC) drug sensitivity test. METHODS: PTC were cultured in vitro with intraoperative specimens, and drug sensitivity test was performed to calculate the most effective chemotherapy regimen for MPM. The patients were divided into conventional and individualized chemotherapy group according to whether they received PTC drug testing. Univariate and multivariate analyses were conducted to identify independent prognostic factors. RESULTS: Among 186 MPM patients included, 63 underwent PTC culture and drug sensitivity test. The results showed that the most effective chemotherapy regimen was oxaliplatin + gemcitabine. After propensity score matching, a total of 64 patients were enrolled in the following study, including 32 patients receiving individualized chemotherapy guided by PTC drug results as group 1 and 32 patients receiving conventional chemotherapy as group 2. Survival analysis showed that the median OS of group 1 was not reached, significantly longer than that of group 2 (23.5 months) (p < 0.05). CONCLUSIONS: Compared with conventional chemotherapy, individualized chemotherapy guided by PTC drug sensitivity tests can prolong patient survival, and oxaliplatin + gemcitabine + apatinib could be the optimal adjuvant treatment regimen for MPM.

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Liver diseases represent a growing global health challenge, and the increasing prevalence of obesity and metabolic disorders is set to exacerbate this crisis. To meet evolving regulatory demands, patient-specific in vitro liver models are essential for understanding disease mechanisms and developing new therapeutic approaches. Organoid models, which faithfully recapitulate liver biology, can be established from both non-malignant and malignant liver tissues, offering insight into various liver conditions, from acute injuries to chronic diseases and cancer. Improved understanding of liver microenvironments, innovative biomaterials, and advanced imaging techniques now facilitate comprehensive and unbiased data analysis, paving the way for personalised medicine. In this review, we discuss state-of-the-art patient-derived liver organoid models, recent technological advancements, and strategies to enhance their clinical impact.

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Background: The effectiveness of radiotherapy for pancreatic cancer is debated. Patient-derived organoids (PDOs) already mimicked clinical radiation response in other cancer types, which could be valuable in pancreatic cancer as well. This study aimed to investigate whether PDOs can be used to model RT response in pancreatic cancer and to explore the presence of a dose-response correlation. Methods: PDOs derived from two pancreatic cancer patients (HUB-08-B2-022A and HUB-08-B2-026B) were irradiated with doses ranging from 0 to 40 Gray. Viability assessments were conducted after seven and 10 days by measuring ATP-levels. Results were normalized, defining the viability at 0 Gray as 100 % and an absolute viability of 0 as 0 %. The relative area under the curve (rAUC) was calculated (0 = total sensitivity, 1 = total resistance). Results: With a readout time of seven days, both HUB-08-B2-022A and HUB-08-B2-026B exhibited viability above 50 % at the highest dose of 12 Gy (rAUC of 0.79 and 0.69, respectively). With a readout time of 10 days, both PDOs showed a dose-response relation although HUB-08-B2-022A was more sensitive than HUB-08-B2-026B (rAUC of 0.37 and 0.51, respectively). Increasing the radiation dose to 40 Gy did not further affect viability, but the dose-response relation remained present (rAUC of 0.13 and 0.26, respectively). In the final experiment with a readout time of 10 days and a maximum dose of 14 Gy, the dose-response correlation was paramount in both PDOs (rAUC 0.28 and 0.45, respectively), with HUB-08-B2-022A being most sensitive. Conclusions: In this setup, both pancreatic cancer PDOs showed an irradiation dose-response correlation. These preliminary findings suggest that pancreatic cancer PDOs are suitable for assessing radiation response in vitro. Further experiments are needed to eventually simulate treatment responses to personalized treatment strategies.

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Musculoskeletal sarcomas pose major challenges to researchers and clinicians due to their rarity and heterogeneity. Xenografting human cells or tumor fragments in rodents is a mainstay for the generation of cancer models and for the preclinical trial of novel drugs. Lately, though, technical, intrinsic and ethical concerns together with stricter regulations have significantly curbed the employment of murine patient-derived xenografts (mPDX). In alternatives to murine PDXs, researchers have focused on embryonal systems such as chorioallantoic membrane (CAM) and zebrafish embryos. These systems are time- and cost-effective hosts for tumor fragments and near-patient cells. The CAM of the chick embryo represents a unique vascularized environment to host xenografts with high engraftment rates, allowing for ease of visualization and molecular detection of metastatic cells. Thanks to the transparency of the larvae, zebrafish allow for the tracking of tumor development and metastatization, enabling high-throughput drug screening. This review will focus on xenograft models of musculoskeletal sarcomas to highlight the intrinsic and technically distinctive features of the different hosts, and how they can be exploited to elucidate biological mechanisms beneath the different phases of the tumor's natural history and in drug development. Ultimately, the review suggests the combination of different models as an advantageous approach to boost basic and translational research.