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Objective: Colorectal Cancer (CRC) is the third most commonly diagnosed cancer in males and the second in females globally. One strategy to reduce mortality in CRC is development of personalized therapies to treat patients in the more advanced stages of the disease. The development of up-to-date in vitro and in vivo preclinical models, expressing the patient-specific cancer lineage and genetic diversity is urgently needed. Three-dimensional models of cancer are emerging: patientderived tumor organoids (PDTOs) and Multicellular Tumor Spheroids (MTS), as well as the role of exosomes as specific biomarkers in CRC prediction and screening. We studied the exosome secretion pattern by TEM and SEM in patientderived MTS and in mouse xenografts, to find possible differences between "in vivo" and "ex vivo" models. Methods: MTS (authorization no. CE5ISS 09/282 of the ethics committee on human experimentation of the Istituto Superiore di Sanità) originated from CRC of a 63 year old man who was then transplanted in an immunodeficient mouse. Both MTS and xenograft were prepared and analyzed by electron microscopy. Morphology, exosome and multivesicular body size were compared and statistically analyzed. Results: In MTS exosome secretion increases proportionately to spheroid complexity, and is less intense than in xenograft. Size of exosome and multivesicular bodies is the same (about 70 nm and 240 nm) in both spheroid and xenograft. Conclusion: MTS are a useful 3D preclinical model and reproduce several aspects of tumor biology. MTS-derived xenografts represent a further improved model of cancer behavior and show sustained exosome production/secretion, possibly due to the influence of the in vivo tumor microenvironment. [ABSTRACT FROM AUTHOR]

Background and aim: Colorectal Cancer (CRC) represents 10% of the global cancer incidence CRC is the third leading cause of cancer-related deaths in both genders worldwide. CRC survival rate is related to the time of diagnosis, when the patients show the symptoms of CRC, such as rectal bleeding, anemia, or abdominal pain, they are in advanced CRC stages when the cancer is aggressive, malignant, and metastatic. To reduce mortality in CRC the development of personalized therapies to treat patients in the more advanced stages of the CRC is required. To achieve this goal a simultaneous development of up-to-date in vitro and in vivo preclinical models, that express the patient-specific cancer lineage and genetic diversity has to be carried on. In the last years are emerging new preclinical models of cancer, such as the three-dimensional patient-derived tumor organoids (PDTOs) and the multicellular tumor spheroids (MTS), and the role of exosomes as specific bio- markers in CRC prediction and screening is growing in importance. In our study we focused on the exosome secretion pattern in patient-derived MTS and in mouse xenografts, to fi nd possible differences between “in vivo” and “ex vivo” models. Methods: MTS originated from CRC of a 63 year old man were then transplanted in immunodeficient mice. Both MTS and xenograft biopsies were analyzed by scanning and transmis- sion electron microscopy. Exosome and multivesicular body size and secretion patterns were compared and statistically analyzed. Results: Exosome secretion pattern in MTS and xenograft is different, in MTS exosome secretion increases as spheroid complexity does and is generally less intense than what is observed in xenograft. The size of exosomes is about 70 nm in both samples as well as for multivesicular bodies (about 240 nm). Conclusions: MTS can be considered a useful 3D preclinical model, they reproduce several aspects of CRC biology. MTS-derived xenografts appear as a further improvement of the cancer behavior model, they show a well-sustained exosome production/secretion, which could be possibly due to the influence of factors present in the microenvironment of the in vivo model. [ABSTRACT FROM AUTHOR]

Simple Summary: In recent years, colorectal cancer stem cells (cCSCs) have been the object of intense investigation for their promise to disclose new aspects of colorectal cancer cell biology, as well as to devise new treatment strategies for colorectal cancer (CRC). However, accumulating studies on cCSCs by complementary technologies have progressively disclosed their plastic nature, i.e., their capability to acquire different phenotypes and/or functions under different circumstances in response to both intrinsic and extrinsic signals. In this review, we aim to recapitulate how a progressive methodological development has contributed to deepening and remodeling the concept of cCSCs over time, up to the present. Colorectal cancer (CRC) represents one of the most deadly cancers worldwide. Colorectal cancer stem cells (cCSCs) are the driving units of CRC initiation and development. After the concept of cCSC was first formulated in 2007, a huge bulk of research has contributed to expanding its definition, from a cell subpopulation defined by a fixed phenotype in a plastic entity modulated by complex interactions with the tumor microenvironment, in which cell position and niche-driven signals hold a prominent role. The wide development of cellular and molecular technologies recent years has been a main driver of advancements in cCSCs research. Here, we will give an overview of the parallel role of technological progress and of theoretical evolution in shaping the concept of cCSCs. [ABSTRACT FROM AUTHOR]

Cancer stem cells (CSCs) are responsible for the initiation of primary tumors and for metastasis seeding at distant organs. Therefore, they represent crucial targets for the study and preclinical testing of new antimetastatic approaches. We recently generated a molecularly characterized biobank of colorectal CSCs, isolated from individual patients and cultured in serum-free medium as multicellular spheroids. Here, we describe in detail the generation of a metastatic model of colorectal cancer based on the orthotopic injection of CSCs into the cecum serosa of immunodeficient mice. Such a model represents an excellent experimental system to investigate the cellular and molecular mechanisms involved in colorectal cancer metastasis, to analyze rare premetastatic elements such as circulating and disseminated tumor cells, and for the preclinical testing of new agents with potential antimetastatic activity., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Breast cancer is the most frequent cancer among women worldwide. Therapeutic strategies to prevent or treat metastatic disease are still inadequate although great progress has been made in treating early-stage breast cancer. Cancer stem-like cells (CSCs) that are endowed with high plasticity and self-renewal properties have been shown to play a key role in breast cancer development, progression, and metastasis. A subpopulation of CSCs that combines tumor-initiating capacity and a dormant/quiescent/slow cycling status is present throughout the clinical history of breast cancer patients. Dormant/quiescent/slow cycling CSCs are a key component of tumor heterogeneity and they are responsible for chemoresistance, tumor migration, and metastatic dormancy, defined as the ability of CSCs to survive in target organs and generate metastasis up to two decades after diagnosis. Understanding the strategies that are used by CSCs to resist conventional and targeted therapies, to interact with their niche, to escape immune surveillance, and finally to awaken from dormancy is of key importance to prevent and treat metastatic cancer. This review summarizes the current understanding of mechanisms involved in CSCs chemoresistance, dissemination, and metastasis in breast cancer, with a particular focus on dormant cells. Finally, we discuss how advancements in the detection, molecular understanding, and targeting of dormant CSCs will likely open new therapeutic avenues for breast cancer treatment., Competing Interests: The authors declare no conflicts of interest.

Banks of genetically characterized cancer stem cells (CSCs) isolated from individual patients and grown as spheroids offer an invaluable approach to identify genetic determinants of drug resistance versus sensitivity, and to study new stem cell-directed therapies. Here, we describe our standardized procedure for in vitro drug screening on colorectal CSCs, taking irinotecan as an example.