PO-300 Unveiling and exploiting cancer stem cell editing and immunogenicity for precision medicine

Introduction Immunogenic chemotherapy (IC) induces immunogenic cell death (ICD), which, similar to viral infection, leads to a cancer-cell autonomous Type-I-Interferon (IFN-I) signalling. This immunological signature is crucial for effective antitumor responses but may paradoxically promote the emergence of a rare population of cancer stem cells (CSCs) acting as a chemoresistant niche within the tumour and roots for metastasis and relapse. In this study, we have investigated the role of IFN-I during IC in inducing a cancer editing program resulting in the appearance of poor immunogenic CSCs. Material and methods Human and murine tumour cell lines were treated in vitro with ICD-inducers or IFN-I as control and the induction of CSC were analysed by cytofluorometry, quantitative real time (qRT)-PCR, 3D culture and functional assays. Free and vesicle-mediated nucleic acid transfer during ICD has been characterised by co-culture experiments. IC-induced CSC immunogenicity has been studied through cytofluorometry, microfluidic devices and in vivo experiments. All experiments have been done in triplicate and statistical significance evaluated by two-tailed Student's t test and two-way ANOVA. Results and discussions The transient/acute induction of IFN-I during ICD is followed by the appearance of a rare population of CSCs. Both free nucleic acids and extracellular vesicles are released during tumour ICD constituting the upstream inducers of IFN-I-mediated reprogramming of neighbouring cells. IC-induced CSCs display epithelial-to-mesenchymal transition traits, multidrug resistance and regenerative properties, and a significant tumorigenic potential when inoculated in immunodeficient and immunocompetent mice. As expected, tumour growth and size are reduced in the presence of an intact immune system. Experiments on microfluidic devices reveal a poor immunogenic potential of CSCs, further confirmed by the expression of immune checkpoint blockers. Conclusion Our results pinpoint a surprising link between ICD, IFN-I and CSCs. Elucidating the mechanisms of CSC editing together with a deep characterisation of CSC (immune) properties could be crucial to prevent tumour relapse. This could undoubtedly have dramatic implications for the clinical management of cancer in an era of terrific development of precision combined chemo-immune therapy

Hyperthermic intraperitoneal chemotherapy in interval debulking surgery for advanced epithelial ovarian cancer: A single-center, real-life experience.

Background: An improvement in survival without increasing perioperative morbidity in patients with advanced epithelial ovarian cancer treated with hyperthermic intraperitoneal chemotherapy (HIPEC) after interval debulking surgery (IDS) has been recently demonstrated in a randomized controlled trial. This study was aimed at assessing the feasibility and perioperative outcomes of the use of HIPEC after IDS at a referral cancer center. Methods: Over the study period, 149 IDSs were performed. Patients who had at least International Federation of Gynecology and Obstetrics stage III disease, with <2.5 mm of residual disease (RD) at the end of surgery and were not participating in clinical trials received HIPEC. Moreover, specific exclusion criteria were considered. These patients were compared with 51 patients with similar clinical characteristics at the same institution and within the same timeframe who did not receive HIPEC. Results: No differences in patient or disease characteristics with the exception of the type of neoadjuvant chemotherapy (P =.002) were found between the 2 groups. As for surgical characteristics, significant differences were found in RD after IDS (P =.007) and in the duration of surgery (P <.001), whereas the bowel resection and diversion rates (P =.583 and P =.213, respectively) and the postoperative intensive care unit and hospital stays (P =.567 and P =.727, respectively) were comparable. The times to start adjuvant chemotherapy were also similar (P =.998). Equally, the rates of any grade of both intraoperative complications (P =.189) and early postoperative complications (P =.238) were superimposable. Conclusions: In the authors' experience, the addition of HIPEC to IDS is feasible in 35% for the population. This value might increase with changes in the inclusion/exclusion criteria. HIPEC does not increase perioperative complications and does not affect a patient's recovery or time to start adjuvant chemotherapy. HIPEC should be offered to select patients listed for IDS

Prevalence and severity of airway obstruction in an Italian adult population

Background. This study sets out to estimate the prevalence and the degree of severity of bronchial obstruction in an adult population with three different diagnostic criteria: the European Respiratory Society (ERS), the American Thoracic Society (ATS), and the World Health Organization (WHO) defined as Global Obstructive Lung Disease (GOLD). Methods. 1514 subjects underwent complete medical evaluation and spirometry. Results. The prevalence of bronchial obstruction was respectively 27.5% (ERS), 33% (GOLD), and 47.3% (ATS). The prevalence of bronchial obstruction in the smoker group was 33.4% (ERS), 38.1% (GOLD), and 52.3% (ATS). The prevalence of obstruction in the ex-smoker group was 33% (ERS), 41.4% (GOLD), and 57.1% (ATS). The prevalence of obstruction in the non-smoker group was 21.1% (ERS), 24.9% (GOLD), and 38.6% (ATS). Conclusions. The results show that the prevalence of airway obstruction increases proportionally with age; the cigarette smoking represents an important conditioning factor. These observations warrant the necessity of a more complete and multi-parametric analysis in the evaluation of patients with airway obstruction using methodologies that explore the functional state and the risk factors that cause the airway obstruction

Prevalence and severity of airway obstruction in an Italian adult population

Background. This study sets out to estimate the prevalence and the degree of severity of bronchial obstruction in an adult population with three different diagnostic criteria: the European Respiratory Society (ERS), the American Thoracic Society (ATS), and the World Health Organization (WHO) defined as Global Obstructive Lung Disease (GOLD). Methods. 1514 subjects underwent complete medical evaluation and spirometry. Results. The prevalence of bronchial obstruction was respectively 27.5% (ERS), 33% (GOLD), and 47.3% (ATS). The prevalence of bronchial obstruction in the smoker group was 33.4% (ERS), 38.1% (GOLD), and 52.3% (ATS). The prevalence of obstruction in the ex-smoker group was 33% (ERS), 41.4% (GOLD), and 57.1% (ATS). The prevalence of obstruction in the non-smoker group was 21.1% (ERS), 24.9% (GOLD), and 38.6% (ATS). Conclusions. The results show that the prevalence of airway obstruction increases proportionally with age; the cigarette smoking represents an important conditioning factor. These observations warrant the necessity of a more complete and multi-parametric analysis in the evaluation of patients with airway obstruction using methodologies that explore the functional state and the risk factors that cause the airway obstruction

miR-205-5p-mediated downregulation of ErbB/HER receptors in breast cancer stem cells results in targeted therapy resistance

The ErbB tyrosine kinase receptor family has been shown to have an important role in tumorigenesis, and the expression of its receptor members is frequently deregulated in many types of solid tumors. Various drugs targeting these receptors have been approved for cancer treatment. Particularly, in breast cancer, anti-Her2/EGFR molecules represent the standard therapy for Her2-positive malignancies. However, in a number of cases, the tumor relapses or progresses thus suggesting that not all cancer cells have been targeted. One possibility is that a subset of cells capable of regenerating the tumor, such as cancer stem cells (CSCs), may not respond to these therapeutic agents. Accumulating evidences indicate that miR-205-5p is significantly downregulated in breast tumors compared with normal breast tissue and acts as a tumor suppressor directly targeting oncogenes such as Zeb1 and ErbB3. In this study, we report that miR-205-5p is highly expressed in BCSCs and represses directly ERBB2 and indirectly EGFR leading to resistance to targeted therapy. Furthermore, we show that miR-205-5p directly regulates the expression of p63 which is in turn involved in the EGFR expression suggesting a miR-205/p63/EGFR regulation

Produção de sementes sintéticas de maracujazeiro silvestre com potencial ornamental

Passiflora cincinnata Mast. é uma espécie silvestre de maracujazeiro de ampla distribuição. Possui hábito trepador e crescimento vigoroso, com flores muito vistosas e perfumadas. O trabalho teve como objetivo a produção de sementes sintéticas da espécie P. cincinnata, utilizando-se de embriões somáticos e zigóticos em diferentes formas de cultivo. Os embriões somáticos em estágio de desenvolvimento pré cotiledonar e cotiledonar foram obtidos a partir de embriões zigóticos cultivados em meio de MS na presença de 18,1 μM de 2,4-Ácido-dichlorophenoxiacetico (2,4-D) e 4,5 μM Benziladenine (BA). Embriões zigóticos e embriões somáticos foram encapsulados em matriz de alginato de sódio a 2,5% (p v -1 ) e complexados em solução estéril de CaCl 2 .2H 2 O a 0,1M. Embriões somáticos e zigóticos foram encapsulados na Matriz (I) com alginato de sódio, Matriz (II) com alginato de sódio + endosperma artificial e Matriz (III) - alginato de sódio + endosperma artificial e suplementado com 0,15% (p v -1 ) de carvão ativado. Embriões zigóticos cultivados em frascos encapsulados na Matriz (I) 79% germinaram, 76% Matriz (II) e na Matriz (III) 86%. Os embriões somáticos cotiledonares encapsulados nos três tipos de matrizes responderam com maior percentual de germinação quando cultivados em plugs de celulose com mais de 50% de embriões convertidos. Os embriões somáticos pre cotiledonares encapsulados nos três tipos de matrizes e nas diferentes formas de cultivo não responderam. No cultivo ex vitro nos dois tipos de substratos PlantMax e Florialite o número de embriões convertidos foi baixo, sendo o melhor resultado com 12,67 % no Florialite e encapsulados na Matriz (I). Palavras-Chave: Passiflora cincinnata, alginato de sódio, sementes artificiais, cultivo in vitroPassiflora cincinnata is a wild species of passion fruit with a wide geographical distribution. It has vigorous growth, climbing habit and very showy and fragrant flowers. The aim of the present investigation was to obtain synthetic seeds from encapsulated zygotic and somatic embryos of P. cincinnata, cultivated under different conditions. Precotyledonary and cotyledonary stage embryos were obtained from zygotic embryos cultivated on MS medium supplemented with 18.1 μM of 2,4-Acid-dichlorophenoxyacetic (2,4-D) and 4.5 μM of Benzyladenine (BA). Zygotic embryos and somatic embryos stages were encapsulated using sodium alginate (2.5% w v-1) and CaCl2.2H2O (1 mM) as complexing agent. The zygotic and somatic embryos were encapsulated in a matrix containing (I) sodium alginate, (II) sodium alginate + artificial endosperm and (III) sodium alginate + artificial endosperm supplemented with activated charcoal (0.15% w/v). Zygotic embryos encapsulated in the matrix (I), matrix (II) and matrix (III) and cultivated in flasks, germinated at rates of 79%, 76% and 86% respectively. The cotyledonary somatic embryos encapsulated in the 3 different matrices showed better germination rates when cultivated on cellulose plugs, with more than 50% of embryos converted into plants. Precotyledonary somatic embryos did not germinated regardless the matrix and cultivation. When cultivating the alginate beads ex vitro, both substrate Plantmax and Florialite showed low number of germinated embryos, and the best result (12.67%) were obtained using Florialite and embryos encapsulated in the matrix (I)

The Effects of Aging on the Molecular and Cellular Composition of the Prostate Microenvironment

Advancing age is associated with substantial increases in the incidence rates of common diseases affecting the prostate gland including benign prostatic hyperplasia (BPH) and prostate carcinoma. The prostate is comprised of a functional secretory epithelium, a basal epithelium, and a supporting stroma comprised of structural elements, and a spectrum of cell types that includes smooth muscle cells, fibroblasts, and inflammatory cells. As reciprocal interactions between epithelium and stromal constituents are essential for normal organogenesis and serve to maintain normal functions, discordance within the stroma could permit or promote disease processes. In this study we sought to identify aging-associated alterations in the mouse prostate microenvironment that could influence pathology.We quantitated transcript levels in microdissected glandular-adjacent stroma from young (age 4 months) and old (age 20-24 months) C57BL/6 mice, and identified a significant change in the expression of 1259 genes (p<0.05). These included increases in transcripts encoding proteins associated with inflammation (e.g., Ccl8, Ccl12), genotoxic/oxidative stress (e.g., Apod, Serpinb5) and other paracrine-acting effects (e.g., Cyr61). The expression of several collagen genes (e.g., Col1a1 and Col3a1) exhibited age-associated declines. By histology, immunofluorescence, and electron microscopy we determined that the collagen matrix is abundant and disorganized, smooth muscle cell orientation is disordered, and inflammatory infiltrates are significantly increased, and are comprised of macrophages, T cells and, to a lesser extent, B cells.These findings demonstrate that during normal aging the prostate stroma exhibits phenotypic and molecular characteristics plausibly contributing to the striking age associated pathologies affecting the prostate

Consensus guidelines for the detection of immunogenic cell death

none82siApoptotic cells have long been considered as intrinsically tolerogenic or unable to elicit immune responses specific for dead cell-associated antigens. However, multiple stimuli can trigger a functionally peculiar type of apoptotic demise that does not go unnoticed by the adaptive arm of the immune system, which we named "immunogenic cell death" (ICD). ICD is preceded or accompanied by the emission of a series of immunostimulatory damage-associated molecular patterns (DAMPs) in a precise spatiotemporal configuration. Several anticancer agents that have been successfully employed in the clinic for decades, including various chemotherapeutics and radiotherapy, can elicit ICD. Moreover, defects in the components that underlie the capacity of the immune system to perceive cell death as immunogenic negatively influence disease outcome among cancer patients treated with ICD inducers. Thus, ICD has profound clinical and therapeutic implications. Unfortunately, the gold-standard approach to detect ICD relies on vaccination experiments involving immunocompetent murine models and syngeneic cancer cells, an approach that is incompatible with large screening campaigns. Here, we outline strategies conceived to detect surrogate markers of ICD in vitro and to screen large chemical libraries for putative ICD inducers, based on a high-content, high-throughput platform that we recently developed. Such a platform allows for the detection of multiple DAMPs, like cell surface-exposed calreticulin, extracellular ATP and high mobility group box 1 (HMGB1), and/or the processes that underlie their emission, such as endoplasmic reticulum stress, autophagy and necrotic plasma membrane permeabilization. We surmise that this technology will facilitate the development of next-generation anticancer regimens, which kill malignant cells and simultaneously convert them into a cancer-specific therapeutic vaccine.Kepp, Oliver; Senovilla, Laura; Vitale, Ilio; Vacchelli, Erika; Adjemian, Sandy; Agostinis, Patrizia; Apetoh, Lionel; Aranda, Fernando; Barnaba, Vincenzo; Bloy, Norma; Bracci, Laura; Breckpot, Karine; Brough, David; Buqué, Aitziber; Castro, Maria G; Cirone, Mara; Colombo, Maria I; Cremer, Isabelle; Demaria, Sandra; Dini, Luciana; Eliopoulos, Aristides G; Faggioni, Alberto; Formenti, Silvia C; Fučíková, Jitka; Gabriele, Lucia; Gaipl, Udo S; Galon, Jérôme; Garg, Abhishek; Ghiringhelli, François; Giese, Nathalia A; Guo, Zong Sheng; Hemminki, Akseli; Herrmann, Martin; Hodge, James W; Holdenrieder, Stefan; Honeychurch, Jamie; Hu, Hong-Min; Huang, Xing; Illidge, Tim M; Kono, Koji; Korbelik, Mladen; Krysko, Dmitri V; Loi, Sherene; Lowenstein, Pedro R; Lugli, Enrico; Ma, Yuting; Madeo, Frank; Manfredi, Angelo A; Martins, Isabelle; Mavilio, Domenico; Menger, Laurie; Merendino, Nicolò; Michaud, Michael; Mignot, Gregoire; Mossman, Karen L; Multhoff, Gabriele; Oehler, Rudolf; Palombo, Fabio; Panaretakis, Theocharis; Pol, Jonathan; Proietti, Enrico; Ricci, Jean-Ehrland; Riganti, Chiara; Rovere-Querini, Patrizia; Rubartelli, Anna; Sistigu, Antonella; Smyth, Mark J; Sonnemann, Juergen; Spisek, Radek; Stagg, John; Sukkurwala, Abdul Qader; Tartour, Eric; Thorburn, Andrew; Thorne, Stephen H; Vandenabeele, Peter; Velotti, Francesca; Workenhe, Samuel T; Yang, Haining; Zong, Wei-Xing; Zitvogel, Laurence; Kroemer, Guido; Galluzzi, LorenzoKepp, Oliver; Senovilla, Laura; Vitale, Ilio; Vacchelli, Erika; Adjemian, Sandy; Agostinis, Patrizia; Apetoh, Lionel; Aranda, Fernando; Barnaba, Vincenzo; Bloy, Norma; Bracci, Laura; Breckpot, Karine; Brough, David; Buqué, Aitziber; Castro, Maria G; Cirone, Mara; Colombo, Maria I; Cremer, Isabelle; Demaria, Sandra; Dini, Luciana; Eliopoulos, Aristides G; Faggioni, Alberto; Formenti, Silvia C; Fučíková, Jitka; Gabriele, Lucia; Gaipl, Udo S; Galon, Jérôme; Garg, Abhishek; Ghiringhelli, François; Giese, Nathalia A; Guo, Zong Sheng; Hemminki, Akseli; Herrmann, Martin; Hodge, James W; Holdenrieder, Stefan; Honeychurch, Jamie; Hu, Hong Min; Huang, Xing; Illidge, Tim M; Kono, Koji; Korbelik, Mladen; Krysko, Dmitri V; Loi, Sherene; Lowenstein, Pedro R; Lugli, Enrico; Ma, Yuting; Madeo, Frank; Manfredi, Angelo A; Martins, Isabelle; Mavilio, Domenico; Menger, Laurie; Merendino, Nicolò; Michaud, Michael; Mignot, Gregoire; Mossman, Karen L; Multhoff, Gabriele; Oehler, Rudolf; Palombo, Fabio; Panaretakis, Theocharis; Pol, Jonathan; Proietti, Enrico; Ricci, Jean Ehrland; Riganti, Chiara; Rovere Querini, Patrizia; Rubartelli, Anna; Sistigu, Antonella; Smyth, Mark J; Sonnemann, Juergen; Spisek, Radek; Stagg, John; Sukkurwala, Abdul Qader; Tartour, Eric; Thorburn, Andrew; Thorne, Stephen H; Vandenabeele, Peter; Velotti, Francesca; Workenhe, Samuel T; Yang, Haining; Zong, Wei Xing; Zitvogel, Laurence; Kroemer, Guido; Galluzzi, Lorenz