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Despite the advances in bone fracture treatment, a significant fraction of fracture patients will develop non-union. Most non-unions are treated with surgery since identifying the molecular causes of these defects is exceptionally challenging. In this study, compared with marrow bone, we generated a transcriptional atlas of human osteoprogenitor cells derived from healing callus and non-union fractures. Detailed comparison among the three conditions revealed a substantial similarity of callus and nonunion at the gene expression level. Nevertheless, when assayed functionally, they showed different osteogenic potential. Utilizing longitudinal transcriptional profiling of the osteoprogenitor cells, we identified FOS as a putative master regulator of non-union fractures. We validated FOS activity by profiling a validation cohort of 31 tissue samples. Our work identified new molecular targets for non-union classification and treatment while providing a valuable resource to better understand human bone healing biology.

Bladder Cancer (BLCa) inter-patient heterogeneity is considered the primary cause of tumor reoccurrence and treatment failure, suggesting that BLCa patients could benefit from a more personalized treatment approach. Patient-derived organoids (PDOs) have been successfully used as a functional model for predicting drug response in different cancer types. In our study, we established BLCa PDO cultures from different BLCa stages. BLCa PDOs preserve the histological and molecular heterogeneity of the parental tumors, including their multiclonal genetic landscapes. BLCa PDOs consistently share key genetic alterations detected in parental tumors, mirroring tumor evolution in longitudinal sampling. Our drug screening pipeline was implemented using BLCa PDOs, testing both standard-of-care and additional FDA-approved compounds for other solid tumors. Integrative analysis of drug response profiles with matched PDO genomic analysis was used to determine enrichment thresholds for candidate markers of therapy resistance and sensitivity. By assessing the clinical history of longitudinally sampled cases, the clonal evolution of the disease could be determined and matched with drug response profiles. In conclusion, we have developed a clinically relevant pipeline for drug response profile assessment and discovery of candidate markers of therapy resistance.

Introduction: Interpatient heterogeneity is one of the causes of treatment failure in bladder cancer (BLCa) patients, who would therefore profit from tailor-made therapies. Toward the direction of precision medicine, patient-derived organoids (PDOs), which retain parental tumor (PT) features, have been successfully tested in different cancer types and shown to predict patients’ responses. Therefore, our study aimed to explore the ability of BLCa PDOs to retain PT features and to determine patient drug sensitivity. We correlated PDO drug response profiles to their genomic profiles and the patient's clinical history. Methods: PDOs were derived from the suspension culture of single cells. Whole exome sequencing was performed on genomic DNA and scRNA sequencing on 3 PT/PDO pairs. Histological evaluation of PDO and PT marker expression and morphology was performed. PDOs were treated for 48h and drug response was evaluated by viability assay. Results: We derived PDOs from non-muscle-invasive and muscle-invasive BLCa. Genomic analysis of 15 representative PT/PDOs pairs showed high similarity in terms of copy number variations and single nucleotide variants (SNVs) with preservation of peculiar BLCa alterations. PT genomic heterogeneity was preserved in PDOs as demonstrated by the correlation between SNVs clonality profiles of PT/PDOs pairs. Moreover, marker analyses at the scRNA and protein levels supported that PDOs retained the main tumor phenotype. Interestingly, PDO grew accordingly to three morphologies linked to marker expression and PT stage.PDOs were used in drug screen assays for testing standard-of-care (SOC) and FDA-approved drugs. PDO responses were highly heterogeneous highlighting interpatient heterogeneity. In 2 case, PDO sensitivity mimicked the patient’s response to SOC and in a few cases correlated with the sample-specific genomic background. Due to the high relapse rate of BLCa, we performed 2 longitudinal studies. In the first one, PDOs from a relapse collected after epirubicin treatment showed less sensitivity to the drug compared to the baseline sample. Moreover, the SNVs' mean clonality of relapse PDOs was significantly higher than the baseline PDOs which were characterized by mutations in DNA damage repair genes (e.g., ATM, FANC1, ERCC2, and BRCA2). We hypothesize a drug-induced selection of a pre-existing epirubicin-resistant population. In the second study, PDOs from the cystectomy and the following relapse showed similar genomic and drug sensitivity profiles suggesting that the first sample could be informative for adjuvant therapy. Conclusion: We established PDOs from different BLCa stages and grades which preserve PT features and can be implemented in drug screening assays for personalized approaches. The power of PDOs in precision medicine was highlighted by the longitudinal studies that showed that PDOs mirrored patient response and tumor evolution in vitro. Citation Format: Martina Minoli, Thomas Cantore, Daniel Hanhart, Mirjam Kiener, Tarcisio Fedrizzi, Federico La Manna, Sofia Karkampouna, Panagiotis Chouvardas, Vera Genitsch, José Antonio Rodriguez-Calero, Eva Compérat, Irena Klima, Paola Gasperini, Bernhard Kiss, Roland Seiler-Blarer, Francesca Demichelis, George N. Thalmann, Marianna Kruithof-de Julio. Bladder cancer organoids as a functional system to model different disease stages and therapy response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 182.

During disease progression from primary towards metastatic prostate cancer (PCa), and in particular bone metastases, the tumor microenvironment (TME) evolves in parallel with the cancer clones, altering extracellular matrix composition (ECM), vasculature architecture, and recruiting specialized tumor-supporting cells that favor tumor spread and colonization at distant sites. We introduce the clinical profile of advanced metastatic PCa in terms of common genetic alterations. Findings from recently developed models of PCa metastatic spread are discussed, focusing mainly on the role of the TME (mainly matrix and fibroblast cell types), at distinct stages: premetastatic niche orchestrated by the primary tumor towards the metastatic site and bone metastasis. We report evidence of premetastatic niche formation, such as the mechanisms of distant site conditioning by extracellular vesicles, chemokines and other tumor-derived mechanisms, including altered cancer cell-ECM interactions. Furthermore, evidence supporting the similarities of stroma alterations among the primary PCa and bone metastasis, and contribution of TME to androgen deprivation therapy resistance are also discussed. We summarize the available bone metastasis transgenic mouse models of PCa from a perspective of pro-metastatic TME alterations during disease progression and give an update on the current diagnostic and therapeutic radiological strategies for bone metastasis clinical management.

Precise characterization and targeting of host cell transcriptional machinery hijacked by SARS-CoV-2 remains challenging. To identify therapeutically targetable mechanisms that are critical for SARS-CoV-2 infection, here we elucidated the Master Regulator (MR) proteins representing mechanistic determinants of the gene expression signature induced by SARS-CoV-2. The analysis revealed coordinated inactivation of MR-proteins linked to regulatory programs potentiating efficiency of viral replication (detrimental host MR-signature) and activation of MR-proteins governing innate immune response programs (beneficial MR-signature). To identify MR-inverting compounds capable of rescuing activity of inactivated host MR-proteins, with-out adversely affecting the beneficial MR-signature, we developed the ViroTreat algorithm. Overall, >80% of drugs predicted to be effective by this methodology induced significant reduction of SARS-CoV-2 infection, without affecting cell viability. ViroTreat is fully generalizable and can be extended to identify drugs targeting the host cell-based MR signatures induced by virtually any pathogen.

To improve the success rate of current preclinical drug trials, there is a growing need for more complex and relevant models that can help predict clinical resistance to anticancer agents. Here, we present a three‐dimensional (3D) technology, based on biomimetic collagen scaffolds, that enables the modeling of the tumor hypoxic state and the prediction of in vivo chemotherapy responses in terms of efficacy, molecular alterations, and emergence of resistance mechanisms. The human breast cancer cell lines MDA‐MB‐231 (triple negative) and MCF‐7 (luminal A) were treated with scaling doses of doxorubicin in monolayer cultures, 3D collagen scaffolds, or orthotopically transplanted murine models. Lineage‐specific resistance mechanisms were revealed by the 3D tumor model. Reduced drug uptake, increased drug efflux, and drug lysosomal confinement were observed in triple‐negative MDA‐MB‐231 cells. In luminal A MCF‐7 cells, the selection of a drug‐resistant subline from parental cells with deregulation of p53 pathways occurred. These cells were demonstrated to be insensitive to DNA damage. Transcriptome analysis was carried out to identify differentially expressed genes (DEGs) in treated cells. DEG evaluation in breast cancer patients demonstrated their potential role as predictive biomarkers. High expression of the transporter associated with antigen processing 1 (TAP1) and the tumor protein p53‐inducible protein 3 (TP53I3) was associated with shorter relapse in patients affected by ER+ breast tumor. Likewise, the same clinical outcome was associated with high expression of the lysosomal‐associated membrane protein 1 LAMP1 in triple‐negative breast cancer. Hypoxia inhibition by resveratrol treatment was found to partially re‐sensitize cells to doxorubicin treatment. Our model might improve preclinical in vitro analysis for the translation of anticancer compounds as it provides: (a) more accurate data on drug efficacy and (b) enhanced understanding of resistance mechanisms and molecular drivers.

Cripto is a small glycosylphosphatidylinisitol (GPI)-anchored and secreted oncofetal protein that plays important roles in regulating normal physiological processes, including stem cell differentiation, embryonal development, and tissue growth and remodeling, as well as pathological processes such as tumor initiation and progression. Cripto functions as a co-receptor for TGF-β ligands such as Nodal, GDF1, and GDF3. Soluble and secreted forms of Cripto also exhibit growth factor-like activity and activate SRC/MAPK/PI3K/AKT pathways. Glucose-Regulated Protein 78 kDa (GRP78) binds Cripto at the cell surface and has been shown to be required for Cripto signaling via both TGF-β and SRC/MAPK/PI3K/AKT pathways. To provide a comprehensive overview of the scientific literature related to Cripto, we performed, for the first time, a bibliometric analysis of the biological roles of Cripto as reported in the scientific literature covering the last 10 years. We present different fields of knowledge in comprehensive areas of research on Cripto, ranging from basic to translational research, using a keyword-driven approach. Our ultimate aim is to aid the scientific community in conducting targeted research by identifying areas where research has been conducted so far and, perhaps more importantly, where critical knowledge is still missing.

Therapy resistance and metastatic processes in prostate cancer (PCa) remain undefined, due to lack of experimental models that mimic different disease stages. We describe a novel androgen-dependent PCa patient-derived xenograft (PDX) model from treatment-naïve, soft tissue metastasis (PNPCa). RNA and whole-exome sequencing of the PDX tissue and organoids confirmed transcriptomic and genomic similarity to primary tumor. PNPCa harboursBRCA2 and CHD1somatic mutations, shows anSPOP/FOXA1-like transcriptomic signature and microsatellite instability, which occurs in 3% of advanced PCa and has never been modelledin vivo. Comparison of the treatment-naïve PNPCa with additional metastatic PDXs (BM18, LAPC9), in a medium-throughput organoid screen of FDA-approved compounds, revealed differential drug sensitivities. Multikinase inhibitors (ponatinib, sunitinib, sorafenib) were broadly effective on all PDX- and patient-derived organoids from advanced cases with acquired resistance to standard-of-care compounds. This proof-of-principle study may provide a preclinical tool to screen drug responses to standard-of-care and newly identified, repurposed compounds.

Therapy resistance and metastatic processes in prostate cancer (PCa) remain undefined, due to lack of experimental models that mimic different disease stages. We describe an androgen-dependent PCa patient-derived xenograft (PDX) model from treatment-naïve, soft tissue metastasis (PNPCa). RNA and whole-exome sequencing of the PDX tissue and organoids confirmed transcriptomic and genomic similarity to primary tumor. PNPCa harbors BRCA2 and CHD1 somatic mutations, shows an SPOP/FOXA1-like transcriptomic signature and microsatellite instability, which occurs in 3% of advanced PCa and has never been modeled in vivo. Comparison of the treatment-naïve PNPCa with additional metastatic PDXs (BM18, LAPC9), in a medium-throughput organoid screen of FDA-approved compounds, revealed differential drug sensitivities. Multikinase inhibitors (ponatinib, sunitinib, sorafenib) were broadly effective on all PDX- and patient-derived organoids from advanced cases with acquired resistance to standard-of-care compounds. This proof-of-principle study may provide a preclinical tool to screen drug responses to standard-of-care and newly identified, repurposed compounds., To date, patients still succumb to cancer, due to tumors not responding to therapy or ultimately acquiring resistance. Here the authors show that by exploiting patient derived organoids and a treatment-naïve patient derived xenograft, patient therapy can be personalized.

Bone metastases are a frequent event in patients with solid tumors. Although great advances have been made in the treatment of these patients, the identification of novel, accurate indicators of bone response would greatly facilitate the clinical management of the disease. The receptor activator of nuclear factor-κB (RANK)/RANK ligand (RANKL)/osteoprotegerin (OPG) signaling pathway is significantly involved in bone metastasis formation. The main aim of the present study was to evaluate the role of circulating RANK, RANKL and OPG levels in predicting bone response. Marker accuracy was also compared with that of the conventional tumor marker N-terminal telopeptide of type I collagen (NTX). A prospective study was performed on 49 patients with bone metastases from breast, lung and prostate cancer, who were undergoing treatment with zoledronic acid. Patients were monitored for 1 year with blood tests, clinical evaluation and instrumental exams according to the response evaluation criteria of the University of Texas M. D. Anderson Cancer Center (Houston, TX, USA) and the Positron Emission Tomography Response Criteria in Solid Tumors. Circulating RANK/RANKL/OPG transcripts and NTX levels were evaluated by reverse transcription-quantitative polymerase chain reaction and immune enzymatic assay, respectively. The baseline RANKL levels differed significantly between responders and non-responders, whereas no differences in NTX levels were observed between the two groups. Receiver operating characteristic curve evaluation for all markers revealed that RANKL was the most accurate marker, with an area under the curve of 0.74 (95% confidence interval, 0.54-0.93). In addition, RANKL, which is the target of the novel monoclonal antibody denosumab, was the most accurate predictor of bone response in the present series of patients with bone metastases. Thus, the use of RANKL as a marker could potentially improve clinical practice, as current bone response evaluation is still somewhat problematic.

Therapy resistance and metastatic processes in prostate cancer (PCa) remain undefined, mainly due to the lack of experimental models representing different disease stages. We aim to provide functional experimental and preclinical drug tools applicable for direct use with patient-derived material. Biopsies from metastatic PCa were used for the establishment of patient-derived xenografts (PDXs) by subcutaneous implantation. In vivo tumor growth kinetic in response to androgens was assessed by surgical castration and testosterone supplementation. RNA and whole-exome sequencing (WES) and organoid drug screens were used to characterize the genomic, transcriptomic, and functional properties. Organoid culture methodology was adapted in order to set up an automated medium-throughput organoid drug screen (Nexus Theragnostics automated platform) for screening standard-of-care compounds and candidates for drug repurposing. A novel case of PCa xenograft model derived from soft tissue metastasis (PNPCa) was established. RNAseq and WES confirmed transcriptomic and genomic similarity to the primary tumor. PNPCa harbors BRAC2 and CHD1 mutations and shows SPOP-like and FOXA1-like transcriptomic signature with microsatellite instability. PNPCa tumor growth is inhibited after androgen deprivation by castration, while androgen replacement leads to tumor reformation. No spontaneous tumor growth occurred after prolonged period of castration; however, scarce micrometastases were found in the bone marrow and lymph nodes. The treatment-naive and androgen-dependent properties of the PNPCa made it a candidate model for identification of potent drug compounds. PNPCa-derived organoid screens on standard-of-care and 74 FDA-approved compounds showed that mTOR pathway and multi-tyrosine kinase inhibitors, used for clinical treatment of other cancer types, have high impact on PCa organoid viability. Application of the organoid drug screen panel on two additional metastatic PDXs of advanced disease has allowed shortlisting of compounds for repurposing use in patient-derived organoid screens that can be routinely performed in a timeframe of few weeks, and thus, provide information on treatment decision. Treatment response of a treatment-naive, early metastatic PCa case (PNPCa PDX) highlights the potential of organoid screens to assess therapy response with extended applications for personalized screens. Citation Format: Sofia Karkampouna, Federico la Manna, Maria R. De Filippo, Mirjam Kiener, Marta De Menna, Eugenio Zoni, Joel Grosjean, Irena Klima, Andrea Garofoli, Marco Bolis, Jean-Philippe Theurillat, Vera Genitsch, David Keller, Booij Tijmen, Christian U. Stirnimann, Andrea Sboner, Charlotte K.Y. Ng, Salvatore Piscuoglio, Martin Spahn, Mark A. Rubin, George N. Thalmann, Marianna Kruithof-de Julio. Patient-derived xenograft and organoids models of prostate cancer [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr B18.

ABSTRACT Gastrointestinal stromal tumors are rare malignancies characterized by c-kit and PDGFR-α mutations targeted by imatinib. Pleural effusion is a very rare side effect of imatinib treatment. A 65-year-old female with metastatic gastrointestinal stromal tumor developed electrolyte imbalance, severe peripheral edema and progressively worsening dyspnea 2 months after starting imatinib. Having excluded cardiovascular and pulmonary disorders, imatinib was discontinued and prednisone 25 mg orally daily was begun. The patient's condition improved substantially over the next 48 h with a progressive decrease in dyspnea and a reduction in pleural effusion and peripheral edema. All side effects had resolved within 1 month. In view of the partial response obtained, the patient re-started imatinib after a 1-week interruption. Prednisone was maintained and there was no further toxicity.

Although bone metastases represent a major challenge in the natural history of breast cancer (BC), the complex interactions involved have hindered the development of robust in vitro models. The aim of this work is the development of a preclinical model of cancer and bone stromal cells to mimic the bone microenvironment. We studied the effects on osteoclastogenesis of BC cells and Mesenchymal stem cells (MSC) cultured alone or in combination. We also analyzed: (a) whether the blockade of the Epithelial Growth Factor Receptor (EGFR) pathway modified their influence on monocytes towards differentiation, and (b) the efficacy of bone-targeted therapy on osteoclasts. We evaluated the osteoclastogenesis modulation of human peripheral blood monocytes (PBMC) indirectly induced by the conditioned medium (CM) of the human BC cell line SCP2, cultured singly or with MSC. Osteoclastogenesis was evaluated by TRAP analysis. The effect of the EGFR blockade was assessed by treating the cells with gefitinib, and analyzed with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Western Blot (WB). We observed that SCP2 co-cultured with MSC increased the differentiation of PBMC. This effect was underpinned upon pre-treatment of the co-culture with gefitinib. Co-culture of SCP2 with MSC increased the expression of both the bone-related marker Receptor Activator of Nuclear Factor κB (RANK) and EGFR in BC cells. These upregulations were not affected by the EGFR blockade. The effects of the CM obtained by the cells treated with gefitinib in combination with the treatment of the preosteoclasts with the bone-targeted agents and everolimus enhanced the inhibition of the osteoclastogenesis. Finally, we developed a fully human co-culture system of BC cells and bone progenitor cells. We observed that the interaction of MSC with cancer cells induced in the latter molecular changes and a higher power of inducing osteoclastogenesis. We found that blocking EGFR signaling could be an efficacious strategy for breaking the interactions between cancer and bone cells in order to inhibit bone metastasis.

Alberto Bongiovanni,1 Nada Riva,1 Sebastiano Calpona,1 Marianna Ricci,1 Erica Gunelli,1 Chiara Liverani,1 Federico La Manna,1 Alessandro De Vita,1 Manuela Monti,1 Stefano Severi,2 Federica Pieri,3 Elena Amadori,1 Riccardo Galassi,1 Davide Cavaliere,4 Alberto Zaccaroni,5 Andreas Tartaglia,6 Veronica Lunedei,7 Andrea Gardini,8 Laura Mercatali,1 Dino Amadori,1 Toni Ibrahim11Osteoncology and Rare Tumors Center, 2Nuclear Medicine Unit, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Meldola, 3Pathology Unit, 4Unit of Oncological Surgery and Advanced Therapies, 5Endocrine Surgery Unit, 6Endocrinology Unit, 7Gastroenterology and Digestive Endoscopy Unit, 8Department of General Surgery, Morgagni-Pierantoni Hospital, Forlì, ItalyBackground: We present a retrospective analysis of metronomic capecitabine in metastatic gastroenteropancreatic neuroendrocrine tumors (GEP-NETs). A review of the literature is also presented.Methods: From January 2007 to December 2013, ten patients with metastatic GEP-NETs (four pancreatic and six ileal) who progressed after treatment with somatostatin analogs and other cytotoxic agents received oral capecitabine 1,500 mg/day continuously. The median patient age was 68 (range 29–82) years. The median treatment duration was 8 months.Results: Five (50%) patients achieved a partial radiographic response, four (40%) showed stable disease, and one (10%) progressed. Median overall survival was 56 months. Three of the four pancreatic patients achieved a partial radiographic response that lasted for a median of 15.5 months; overall survival and progression-free survival in this subgroup was 58 and 6 months, respectively.Conclusion: Data in the literature show that capecitabine has only occasionally been used as a single agent, with increased toxicity. Only one study using single-agent capecitabine reported a progression-free survival of 9.9 months and overall survival of 36.5 months, without an objective response or major toxicity. In our experience, metronomic capecitabine was well tolerated, although minor side effects may have been underestimated due to the retrospective nature of our study. This regimen also seems to be feasible in elderly people. Although high response rates and prolonged response duration indicate the potential efficacy of this treatment, our results should be interpreted cautiously because of the small number of patients involved. Capecitabine was most effective in the pancreatic subgroup, which would seem to be more sensitive to chemotherapy.Keywords: capecitabine, gastroenteropancreatic neuroendocrine tumors, metronomic chemotherapy

Patient-derived specimens are an invaluable resource to investigate tumor biology. However

Patient-derived specimens are an invaluable resource to investigate tumor biology. However, in vivo studies on primary cultures are often limited by the small amount of material available, while conventional in vitro systems might alter the features and behavior that characterize cancer cells. We present our data obtained on primary dedifferentiated liposarcoma cells cultured in a 3D scaffold-based system and injected into a zebrafish model. Primary cells were characterized in vitro for their morphological features, sensitivity to drugs and biomarker expression, and in vivo for their engraftment and invasiveness abilities. The 3D culture showed a higher enrichment in cancer cells than the standard monolayer culture and a better preservation of liposarcoma-associated markers. We also successfully grafted primary cells into zebrafish, showing their local migratory and invasive abilities. Our work provides proof of concept of the ability of 3D cultures to maintain the original phenotype of ex vivo cells, and highlights the potential of the zebrafish model to provide a versatile in vivo system for studies with limited biological material. Such models could be used in translational research studies for biomolecular analyses, drug screenings and tumor aggressiveness assays.

Alberto Bongiovanni,1 Nada Riva,1 Marianna Ricci,1 Chiara Liverani,1 Federico La Manna,1 Alessandro De Vita,1 Flavia Foca,2 Laura Mercatali,1 Stefano Severi,3 Dino Amadori,4 Toni Ibrahim1 1Osteoncology and Rare Tumors Center, 2Unit of Biostatistics and Clinical Trials, 3Nuclear Medicine Unit, 4Department of Medical Oncology, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Meldola, FC, Italy Background and aim: To investigate the efficacy of platinum-based chemotherapy in patients with metastatic gastroenteropancreatic neuroendocrine carcinoma (mGEP-NEC) and define predictive and prognostic factors. Methods: Twenty mGEP-NEC patients were treated with cisplatin or carboplatin/etoposide between April 2010 and October 2014. Both large-cell and small-cell histologies were included. Cisplatin 25 mg/m2 was administered on days 1–3 followed by etoposide 100 mg/m2 on days 1–3 every 21 days. Carboplatin 300 mg/m2 was administered on day 1 followed by etoposide 100 mg/m2 on days 1–3. Results: Of the 19 evaluable patients, 13 obtained a partial response, four showed stable disease, and two progressed. Median overall survival (mOS) was 13.5 months and median progression-free survival (mPFS) was 10.9 months. Gallium-68 positron emission tomography/computerizsed tomography-positive patients had a higher, albeit not significantly, OS than those with negative results (75% vs 34.3% at 18 months; P=0.06). mPFS was 19.3 and 6.3 months (P55%, respectively. mOS was 8.1 months in the latter group but was not reached in the Ki67 ≤55% group (P-value =0.039). Patients with a lower body mass index (BMI) had a better prognosis in terms of both OS and PFS. Patients with BMI≥25 had a mOS of 11.7 months (P=0.0293) and a mPFS of 6.2 months (P=0.0057). Conclusion: Platinum-based chemotherapy showed good efficacy in mGEP-NEC patients. Those with Ki67 ≤55%, positive Gallium-68 positron emission tomography/computerized tomography and BMI

Prostate cancer (PCa) prognosis and clinical outcome is directly dependent on metastatic occurrence. The bone microenvironment is a favorable metastatic niche. Different biological processes have been suggested to contribute to the osteotropism of PCa such as hemodynamics, bone-specific signaling interactions, and the "seed and soil" hypothesis. However, prevalence of disseminating tumor cells in the bone is not proportional to the actual occurrence of metastases, as not all patients will develop bone metastases. The fate and tumor-reforming ability of a metastatic cell is greatly influenced by the microenvironment. In this review, the molecular mechanisms of bone and soft-tissue metastasis in PCa are discussed. Specific attention is dedicated to the residual disease, novel approaches, and animal models used in oncological translational research are illustrated.

Introduction & objectives: Prostate cancer (PCa)-associated mortality results from metastasis to bone and resistance to androgen deprivation or cytotoxic therapy. Despite early detection of primary PCa, advanced castration resistant prostate cancer (CRPC) and bone metastases (BM) are detected in 10% of patients already at the time of initial diagnosis. The majority of recurrences might be due to cancer cells with stem cell-like properties (cancer stem cell-like, CSC-like). These could be therapy-resistant in a dormant state at the primary site or have metastasize prior to diagnosis of the primary tumor. CSC-like cells are the most tumorigenic and metastatic, however, current treatments target the differentiated tumor bulk cells. Understanding the mechanisms of PCa tumor initiation and metastasis by CSC-like cells is crucial for proper prognosis of high risk patient groups. Materials & methods: To model CSC-like cells we generated organoids from patient-derived tissues (here termed as “canceroids”) and established patient-derived xenografts (PDXs). Canceroids were derived from bulk tumor tissues; primary PCa, bone metastasis from PCa tissue (BM-PCa), established PDX models LAPC9 and BM18 and lymph node (LN) metastasis from PCa (LAPC4) PDX models. Cytotoxic compounds and androgen inhibitors are tested on the canceroids from different BM or LN tissues using viability assays (CellTiter Glo assay) and 3D imaging by confocal and light sheet microscopy. Results: We have generated several canceroid lines from human primary PCa and from established PDX models that maintain key features of the CSC-like cells, previously characterized on the tumor itself (AR, PSA, Cytokeratins). The luminal phenotype of BM18 is evidently maintained in the BM18 canceroids, based on positive cytokeratin (CK)18 and absent CK5 expression. LAPC4 organoids contain CK5 and CK18 cells, in line with the mixed basal and luminal phenotype. Imaging of PSA and cytokeratin distribution confirms the luminal phenotype of the canceroids. BM18 and LAPC9 canceroids are maintained both in presence and absence of dihydrotestosterone indicating that androgen independent growth properties of the tumor are conserved in the canceroids. Viability assays indicate that canceroids respond to chemotherapeutics (cabacitaxel, docetaxel) and partially to hormone inhibitors (abiraterone, enzalutamide). Conclusions: Identification of the oncogenic properties of metastatic CSC-like (sub)populations has both prognostic and therapeutic applications. Establishment of CSC-derived organoids is the first step towards routine derivation from metastasis or primary PCa tissues as a potential platform for personalized drug compound evaluation. Citation Format: Sofia Karkampouna, Federico la Manna, Eugenio Zoni, Lijkele Beimers, Peter Kloen, Antoinette Wetterwald, Joël Grosjean, Irena Klima, Marco Giovanni Cecchini, Martin Spahn, George Niklaus Thalmann, Marianna Kruithof- de Julio. Establishment, characterization and drug response of primary and metastatic prostate organoids [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3885. doi:10.1158/1538-7445.AM2017-3885

Metastatic bone disease has a major impact on the morbidity and mortality of breast cancer patients, and studies on bone metastasis biology have led to the development of the most widely used drugs for bone metastases treatment: zoledronate (Zol) and denosumab (Den). The aim of the present study was to assess the effect of soluble mediators produced by breast cancer cells on human osteoclast maturation in a co-culture model. We also tested the ability of zoledronate, denosumab and 5H4, an antibody directed against CSF-1, to interfere with the osteoclastogenic potential of breast cancer. The study was performed on the triple negative cell line MDA-MB-231 and on human osteoclasts obtained from the differentiation of peripheral blood monocytes of a healthy volunteer. Osteoclastogenesis was evaluated by TRAP assay after 14days of differentiation with 10% MDA-MB-231-conditioned media or with CSF-1 and RANKL. Den, Zol and 5H4 were administered after 7days of differentiation. MDA-MB-231-conditioned media doubled the differentiation of monocytes into osteoclasts. MDA-MB-231 secreted CSF-1, especially when cells were cultured to confluence. Induced osteoclasts were sensitive to bone-targeted drugs: Den and 5H4 blocked osteoclast differentiation and survival, while Zol induced osteoclast apoptosis. Osteoclasts differentiated by breast cancer cells were less sensitive to Zol than those induced by differentiation factors, whereas sensitivity to Den was similar. Conversely, breast cancer-induced osteoclast activation resulted in a higher sensitivity to 5H4. A significant increase in CSF-1 secretion was observed in osteoclast precursors after treatment with the highest concentration of Den. Further research is ongoing to evaluate the efficacy of 5H4 combination with Den.

Bone metastasis is a complex process that needs to be better understood in order to help clinicians prevent and treat it. Xenografts using patient-derived material (PDX) rather than cancer cell lines are a novel approach that guarantees more clinically realistic results. A primary culture of bone metastasis derived from a 67-year-old patient with breast cancer was cultured and then injected into zebrafish (ZF) embryos to study its metastatic potential. In vivo behavior and results of gene expression analyses of the primary culture were compared with those of cancer cell lines with different metastatic potential (MCF7 and MDA-MB-231). The MCF7 cell line, which has the same hormonal receptor status as the bone metastasis primary culture, did not survive in the in vivo model. Conversely, MDA-MB-231 disseminated and colonized different parts of the ZF, including caudal hematopoietic tissues (CHT), revealing a migratory phenotype. Primary culture cells disseminated and in later stages extravasated from the vessels, engrafting into ZF tissues and reaching the CHT. Primary cell behavior reflected the clinical course of the patient’s medical history. Our results underline the potential for using PDX models in bone metastasis research and outline new methods for the clinical application of this in vivo model.

e23190Background: Patient-derived specimens are an invaluable resource to study cancer in particular in rare tumors as Soft Tissue Sarcomas, for which several subtypes exist, that are not all cover...

11128 Background: Bone Metastases (BM) are responsible of the high morbidity, low quality of life and poor prognosis of patients with BC. The evaluation of bone response to treatment is still contr...

e22248 Background: The debate on the association between Vit D deficiency and breast cancer risk is still open. Some studies indicate that ethnic differences play a role in the circulating levels of serum Vit D. Moreover, single nucleotide polymorphisms (SNPs) in the Vit D receptor (VDR) have been associated with various types of cancer.The study evaluates Vit D and Binding Protein (DBP) levels, VDR and DBP polymorphisms in Tanzanian and Italian healthy donors (HD) and breast cancer patients (BCP). Methods: The study provides 200 adult women: 100 Tanzanian and 100 Italians (for each population: 50 donors, 25 primary and 25 advanced breast cancer patients). At first, analysis on serum Vit D concentrations has been performed until now on 25 HD and 25 BCP from Italy. The analysis on the overall population is ongoing. All bio-molecular characterization are performed at IRST IRCCS in Italy. The association between Vit D serum concentration and osteoporosis risk factors was evaluated. Results: From Sept to Dec ...

Background Leiomyosarcoma (LMS) is an aggressive soft tissue sarcoma derived from smooth muscle cells typically of uterine, gastrointestinal or soft tissue origin. The prognosis for this tumor is poor, with survival rates among the lowest of all soft tissue sarcomas. Surgery is the best approach for localized disease. The principal role of chemotherapy is prevalently in the treatment of metastatic disease. Trabectedin, a promising new DNA-damaging agent with a mechanism of action that differs from that of traditional alkylating agents, has been approved in Europe for the treatment of patients with advanced soft tissue sarcoma after failure of anthracyclines and ifosfamide, Case presentation We report the case of a 53-year-old woman with metastatic well differentiated uterine leiomyosarcoma refractory to multiple treatments who underwent 22 cycles of trabectedin over 30 months, obtaining a partial response according to RECIST (Response Evaluation Criteria in Solid Tumors) criteria, with good tolerability, and maintaining the response for 10 months after trebectedin withdrawal. Conclusion This very prolonged response, which persisted after drug discontinuation, suggests that trabectedin exerts an oncostatic effect rather than the cytotoxic one produced by other chemotherapeutic agents. Our experience also raises the question of the best way to evaluate trabectedin efficacy.