Organoids as a Model for Precision Medicine in Malignant Pleural Mesothelioma: Where Are We Today?

MPM is an aggressive tumor originating from pleural mesothelial cells. A characteristic feature of the disease is the dominant prevalence of therapeutically intractable inactivating alterations in TSGs, making MPM one of the most difficult cancers to treat and the epitome of a cancer characterized by a significant lack of therapy options and an extremely poor prognosis (5-year survival rate of only 5% to 10%). Extensive interpatient heterogeneity poses another major challenge for targeted therapy of MPM, warranting stratified therapy for specific subgroups of MPM patients. Accurate preclinical models are critical for the discovery of new therapies and the development of personalized medicine. Organoids, an in vitro 'organ-like' 3D structure derived from patient tumor tissue that faithfully mimics the biology and complex architecture of cancer and largely overcomes the limitations of other existing models, are the next-generation tumor model. Although organoids have been successfully produced and used in many cancers, the development of MPM organoids is still in its infancy. Here, we provide an overview of recent advances in cancer organoids, focusing on the progress and challenges in MPM organoid development. We also elaborate the potential of MPM organoids for understanding MPM pathobiology, discovering new therapeutic targets, and developing personalized treatments for MPM patients

Zebrafish Microenvironment Elevates EMT and CSC-Like Phenotype of Engrafted Prostate Cancer Cells.

To visually and genetically trace single-cell dynamics of human prostate cancer (PCa) cells at the early stage of metastasis, a zebrafish (ZF) xenograft model was employed. The phenotypes of intravenously transplanted fluorescent cells were monitored by high-resolution, single-cell intravital confocal and light-sheet imaging. Engrafted osteotropic, androgen independent PCa cells were extravasated from caudle vein, invaded the neighboring tissue, proliferated and formed experimental metastases around caudal hematopoietic tissue (CHT) in four days. Gene expression comparison between cells in culture and in CHT revealed that engrafted PCa cells responded to the ZF microenvironment by elevating expression of epithelial-mesenchymal transition (EMT) and stemness markers. Next, metastatic potentials of ALDHhi cancer stem-like cells (CSCs) and ALDHlow non-CSCs were analyzed in ZF. Engraftment of CSCs induced faster metastatic onset, however after six days both cell subpopulations equally responded to the ZF microenvironment, resulting in the same increase of stemness genes expression including Nanog, Oct-4 and Cripto. Knockdown of Cripto significantly reduced the vimentin/E-cadherin ratio in engrafted cells, indicating that Cripto is required for transduction of the microenvironment signals from the ZF niche to increase mesenchymal potential of cells. Targeting of either Cripto or EMT transcriptional factors Snail 1 and Zeb1 significantly suppressed metastatic growth. These data indicated that zebrafish microenvironment governed the CSC/EMT plasticity of human PCa cells promoting metastasis initiation

Stromal-epithelial interaction induces GALNT14 in prostate carcinoma cells

IntroductionCell-cell communication is an important process in healthy tissue but also gains enhanced attention regarding pathological tissue. To date, the tumor microenvironment is gradually brought into focus when studying tumorigenesis. In the prostate gland, stromal and epithelial cells greatly interact to maintain homeostasis or tissue integrity. This study focuses on an indirect communication via soluble factors.MethodsTo investigate the cell-cell interaction via soluble factors, the prostate carcinoma cell line LNCaP and the stromal primary cells p21 were co-cultured without direct contact and RNA was isolated at defined time points. Differences in gene expression were finally analyzed by RNA sequencing.ResultsRNA sequencing revealed a time-depending differential expression profile. Selected factors were subsequently characterized at molecular level and analyzed in human prostate tissue of different developmental stages as well as pathology. GALNT14 was one of the highest induced co-culture-specific genes in LNCaP cells. Detection in healthy tissue and BPH revealed an age-dependent decrease in GALNT14 expression. Moreover, in prostate carcinoma, GALNT14 expression heavily varied independent of the Gleason score.ConclusionOverall, this work provides a basis for further studies related to paracrine stromal-epithelial interaction in prostate carcinoma and highlights the importance of GALNT14

Matching single cells across modalities with contrastive learning and optimal transport.

Understanding the interactions between the biomolecules that govern cellular behaviors remains an emergent question in biology. Recent advances in single-cell technologies have enabled the simultaneous quantification of multiple biomolecules in the same cell, opening new avenues for understanding cellular complexity and heterogeneity. Still, the resulting multimodal single-cell datasets present unique challenges arising from the high dimensionality and multiple sources of acquisition noise. Computational methods able to match cells across different modalities offer an appealing alternative towards this goal. In this work, we propose MatchCLOT, a novel method for modality matching inspired by recent promising developments in contrastive learning and optimal transport. MatchCLOT uses contrastive learning to learn a common representation between two modalities and applies entropic optimal transport as an approximate maximum weight bipartite matching algorithm. Our model obtains state-of-the-art performance on two curated benchmarking datasets and an independent test dataset, improving the top scoring method by 26.1% while preserving the underlying biological structure of the multimodal data. Importantly, MatchCLOT offers high gains in computational time and memory that, in contrast to existing methods, allows it to scale well with the number of cells. As single-cell datasets become increasingly large, MatchCLOT offers an accurate and efficient solution to the problem of modality matching

Concept development of an on-chip PET system.

BACKGROUND Organs-on-Chips (OOCs), microdevices mimicking in vivo organs, find growing applications in disease modeling and drug discovery. With the increasing number of uses comes a strong demand for imaging capabilities of OOCs as monitoring physiologic processes within OOCs is vital for the continuous improvement of this technology. Positron Emission Tomography (PET) would be ideal for OOC imaging, however, current PET systems are insufficient for this task due to their inadequate spatial resolution. In this work, we propose the concept of an On-Chip PET system capable of imaging OOCs and optimize its design using a Monte Carlo Simulation (MCS). MATERIAL AND METHODS The proposed system consists of four detectors arranged around the OOC device. Each detector is made of two monolithic LYSO crystals and covered with Silicon photomultipliers (SiPMs) on multiple surfaces. We use a Convolutional Neural Network (CNN) trained with data from a MCS to predict the first gamma-ray interaction position inside the detector from the light patterns that are recorded by the SiPMs on the detector's surfaces. RESULTS The CNN achieves a mean average prediction error of 0.80 mm in the best configuration. The proposed system achieves a sensitivity of 34.81% for 13 mm thick crystals and does not show a prediction degradation near the boundaries of the detector. We use the trained network to reconstruct an image of a grid of 21 point sources spread across the field-of-view and obtain a mean spatial resolution of 0.55 mm. We show that 25,000 Line of Responses (LORs) are needed to reconstruct a realistic OOC phantom with adequate image quality. CONCLUSIONS We demonstrate that it is possible to achieve a spatial resolution of almost 0.5 mm in a PET system made of multiple monolithic LYSO crystals by directly predicting the scintillation position from light patterns created with SiPMs. We observe that a thinner crystal performs better than a thicker one, that increasing the SiPM size from 3 mm to 6 mm only slightly decreases the prediction performance, and that certain surfaces encode significantly more information for the scintillation-point prediction than others

Connective tissue degeneration: Mechanisms of palmar fascia degeneration (Dupuytren’s disease)

Dupuytren’s disease is a connective tissue disorder of the hand causing excessive palmar fascial fibrosis with associated finger contracture and disability. The aetiology of the disease is heterogeneous, with both genetic and environmental components. The connective tissue is abnormally infiltrated by myofibroblasts that deposit collagen and other extracellular matrix proteins. We describe the clinical profile of Dupuytren’s disease along with current therapeutic schemes. Recent findings on molecular and cellular parameters that are dysregulated in Dupuytren’s disease, which may contribute to the onset of the disease, and the role of resident inflammation promoting fibrosis, are highlighted. We review recent literature focusing on non-myofibroblast cell types (stem cell-like cells), their pro-inflammatory and pro-fibrotic role that may account for abnormal wound healing response

Epiregulin expression and secretion is increased in castration-resistant prostate cancer.

INTRODUCTION In prostate cancer, long-term treatment directed against androgens often leads to the development of metastatic castration-resistant prostate cancer, which is more aggressive and not curatively treatable. Androgen deprivation results in elevated epiregulin expression in LNCaP cells which is a ligand of EGFR. This study aims to reveal the expression and regulation of epiregulin in different prostate cancer stages enabling a more specific molecular characterization of different prostate carcinoma types. METHODS Five different prostate carcinoma cell lines were used to characterize the epiregulin expression on the RNA and protein levels. Epiregulin expression and its correlation with different patient conditions were further analyzed using clinical prostate cancer tissue samples. Additionally, the regulation of epiregulin biosynthesis was examined at transcriptional, post-transcriptional and release level. RESULTS An increased epiregulin secretion is detected in castration-resistant prostate cancer cell lines and prostate cancer tissue samples indicating a correlation of epiregulin expression with tumor recurrence, metastasis and increased grading. Analysis regarding the activity of different transcription factors suggests the involvement of SMAD2/3 in the regulation of epiregulin expression. In addition, miR-19a, -19b, and -20b are involved in post-transcriptional epiregulin regulation. The release of mature epiregulin occurs via proteolytic cleavage by ADAM17, MMP2, and MMP9 which are increased in castration-resistant prostate cancer cells. DISCUSSION The results demonstrate epiregulin regulation by different mechanism and suggest a potential role as a diagnostic tool to detect molecular alterations in prostate cancer progression. Additionally, although EGFR inhibitors false in prostate cancer, epiregulin could be a therapeutic target for patients with castration-resistant prostate cancer

Small Molecular Inhibitors Block TRPM4 Currents in Prostate Cancer Cells, with Limited Impact on Cancer Hallmark Functions.

Transient receptor potential melastatin 4 (TRPM4) is a broadly expressed Ca2+ activated monovalent cation channel that contributes to the pathophysiology of several diseases. For this study, we generated stable CRISPR/Cas9 TRPM4 knockout (K.O.) cells from the human prostate cancer cell line DU145 and analyzed the cells for changes in cancer hallmark functions. Both TRPM4-K.O. clones demonstrated lower proliferation and viability compared to the parental cells. Migration was also impaired in the TRPM4-K.O. cells. Additionally, analysis of 210 prostate cancer patient tissues demonstrates a positive association between TRPM4 protein expression and local/metastatic progression. Moreover, a decreased adhesion rate was detected in the two K.O. clones compared to DU145 cells. Next, we tested three novel TRPM4 inhibitors with whole-cell patch clamp technique for their potential to block TRPM4 currents. CBA, NBA and LBA partially inhibited TRPM4 currents in DU145 cells. However, none of these inhibitors demonstrated any TRPM4-specific effect in the cellular assays. To evaluate if the observed effect of TRPM4 K.O. on migration, viability, and cell cycle is linked to TRPM4 ion conductivity, we transfected TRPM4-K.O. cells with either TRPM4 wild-type or a dominant-negative mutant, non-permeable to Na+. Our data showed a partial rescue of the viability of cells expressing functional TRPM4, while the pore mutant was not able to rescue this phenotype. For cell cycle distribution, TRPM4 ion conductivity was not essential since TRPM4 wild-type and the pore mutant rescued the phenotype. In conclusion, TRPM4 contributes to viability, migration, cell cycle shift, and adhesion; however, blocking TRPM4 ion conductivity is insufficient to prevent its role in cancer hallmark functions in prostate cancer cells

Guidance of adjuvant instillation in intermediate-risk non-muscle invasive bladder cancer by drug screens in patient derived organoids: a single center, open-label, phase II trial.

BACKGROUND In intermediate-risk non-muscle invasive bladder cancer (NMIBC) clinical guidelines suggest an adjuvant instillation with a chemotherapeutic agent. However, the agent and regimen are not clearly defined. Worldwide, less than 15% of patients receive this adjuvant chemotherapeutic instillation. We recently developed a pipeline for the generation of patient derived organoids (PDO) in NMIBC. In this phase II trial, we aim to use our in vitro pipeline to select the most effective drug for chemotherapeutic instillation in NMIBC patients. METHODS Patients with first diagnosis of intermediate-risk NMIBC that are directed to transurethral resection of bladder tumor (TURBT) are enrolled. During TURBT, tumor is sampled, and specimens are directed to generate PDO. Once the PDO are formed, drug screens on them for Epirubicin, Mitomycin C, Gemcitabine and Docetaxel are performed. The drug with the highest antitumor activity in vitro will then be selected for 6 adjuvant intravesical instillations once weekly. Thereafter, patients are followed according to clinical guidelines by cystoscopy. DISCUSSION The aim of this trial is to use drug screens in PDO to precise treatment selection for adjuvant instillation therapies in patients with intermediate-risk NMIBC. The ultimate goal of this trial is to reduce the risk of cancer recurrence. In the future, we aim to conduct clinical multicenter trials with an increased sample size, a broader panel of compounds and a focus on the reduction of cancer recurrence by precision delivery of care. Trial registration NCT05024734