Tumor tissue samples, excised from mice or human subjects, are integrated into a surrounding supportive tissue matrix, including an extensive network of stroma and blood vessels. The methodology is significantly more representative than tissue culture assays and considerably faster than patient-derived xenograft models. It's easily implementable, compatible with high-throughput procedures, and is not burdened by the ethical or financial costs associated with animal studies. High-throughput drug screening can be efficiently performed using our physiologically relevant model.

To investigate organ physiology and to create models of diseases, like cancer, renewable and scalable human liver tissue platforms prove to be a powerful instrument. Models originating from stem cells stand as a replacement for cell lines, potentially demonstrating less applicability to the nature of primary cells and their tissues. Two-dimensional (2D) models of liver function have been common historically, as they lend themselves well to scaling and deployment. 2D liver models exhibit inadequate functional diversity and phenotypic stability within prolonged culture settings. To mitigate these problems, protocols for generating three-dimensional (3D) tissue structures were developed. We outline a method for creating three-dimensional liver spheres using pluripotent stem cells in this report. Liver spheres, containing hepatic progenitor cells, endothelial cells, and hepatic stellate cells, have enabled significant advancements in the study of human cancer cell metastasis patterns in humans.

In diagnostic investigations of blood cancer patients, peripheral blood and bone marrow aspirates are obtained, yielding readily accessible specimens of patient-specific cancer cells and non-malignant cells suitable for research projects. By employing density gradient centrifugation, this method, easily replicable and simple, facilitates the isolation of viable mononuclear cells, including malignant cells, from fresh peripheral blood or bone marrow aspirates. Further purification of the cells obtained using the outlined protocol is possible to facilitate various cellular, immunological, molecular, and functional studies. Furthermore, these cells are capable of being cryopreserved and stored in a biobank for future research initiatives.

Applications of three-dimensional (3D) tumor spheroids and tumoroids extend to the study of lung cancer, encompassing aspects of tumor growth, proliferation, invasion, and the screening of novel therapies. While 3D tumor spheroids and tumoroids are valuable tools, they fail to completely reproduce the structural complexity of human lung adenocarcinoma tissue, particularly the direct cellular contact with air, as they lack polarity. Our approach circumvents this constraint by facilitating the growth of lung adenocarcinoma tumoroids and healthy lung fibroblasts at the air-liquid interface (ALI). The ability to easily access both the apical and basal surfaces of the cancer cell culture contributes several advantages to drug screening applications.

In the context of cancer research, the human lung adenocarcinoma cell line A549 is a standard model for mimicking malignant alveolar type II epithelial cells. In the cultivation of A549 cells, Ham's F12K (Kaighn's) or Dulbecco's Modified Eagle's Medium (DMEM) is typically supplemented with 10% fetal bovine serum (FBS) and glutamine. However, the implementation of FBS raises important scientific doubts regarding the indeterminacy of its constituents and inconsistencies between batches, which may jeopardize the reproducibility of experiments and the accuracy of results. Selleck AZD0095 In this chapter, the process of switching A549 cells to a FBS-free medium is described, accompanied by recommendations for further characterization and functional assays to validate the cultured cells' properties.

Despite the emergence of improved therapies for specific subsets of non-small cell lung cancer (NSCLC), the chemotherapy agent cisplatin remains a standard treatment for advanced NSCLC patients lacking oncogenic driver mutations or immune checkpoint activity. Unfortunately, acquired drug resistance, a common issue in solid tumors, is also prevalent in non-small cell lung cancer (NSCLC), creating a significant clinical challenge for oncology specialists. To investigate the cellular and molecular mechanisms underlying cancer drug resistance, isogenic models offer a valuable in vitro platform for exploring novel biomarkers and pinpointing potential druggable pathways in drug-resistant cancers.

Radiation therapy's role in cancer treatment is paramount across the world. Unfortunately, the control of tumor growth is frequently absent, and treatment resistance is a common characteristic of many tumors. Extensive research has been conducted on the molecular pathways that underlie cancer's resistance to treatment. The investigation of the molecular underpinnings of radioresistance in cancer research is greatly enhanced by the use of isogenic cell lines with varying radiosensitivities. These lines curtail the significant genetic variation present in patient samples and cell lines of different origins, thereby enabling the discovery of the molecular determinants of radiation response. The procedure for generating an in vitro model of radioresistant esophageal adenocarcinoma, which involves chronic X-ray irradiation of esophageal adenocarcinoma cells at clinically relevant doses, is detailed. In this model, we also investigate the underlying molecular mechanisms of radioresistance in esophageal adenocarcinoma, characterizing cell cycle, apoptosis, reactive oxygen species (ROS) production, DNA damage, and repair.

Investigating mechanisms of radioresistance in cancer cells has seen an increase in the use of in vitro isogenic models generated through fractionated radiation exposures. Because of the intricate biological response to ionizing radiation, a precise understanding of radiation exposure protocols and cellular endpoints is crucial to the creation and confirmation of these models. Bioelectronic medicine Within this chapter, we describe a protocol for the development and assessment of an isogenic model for radioresistant prostate cancer cells. Other cancer cell lines might find this protocol useful.

While non-animal methodologies (NAMs) experience a surge in adoption and development, alongside validation, animal models continue to be employed in cancer research. Animals serve multiple roles in research, encompassing molecular trait and pathway investigation, mimicking clinical tumor development, and evaluating drug responses. AIT Allergy immunotherapy In vivo studies are multifaceted and require expertise across diverse fields, including animal biology, physiology, genetics, pathology, and animal welfare. The goal of this chapter is not to provide an exhaustive catalog of all cancer research animal models. Rather, the authors aim to furnish experimenters with the strategies for in vivo experimental procedures, encompassing the selection of cancer animal models, during both the planning and execution phases.

Cellular growth outside of an organism, cultivated in a laboratory setting, is a crucial instrument in expanding our comprehension of a plethora of biological concepts, including protein production, the intricate pathways of drug action, the potential of tissue engineering, and the intricacies of cellular biology in its entirety. Decades of cancer research have been heavily reliant on conventional two-dimensional (2D) monolayer culture methods for evaluating a multitude of cancer characteristics, encompassing everything from the cytotoxic effects of anti-tumor medications to the toxicity profiles of diagnostic stains and contact tracers. Yet, many potentially effective cancer therapies display limited or no efficacy in clinical practice, thereby delaying or preventing their actual application to patients. The 2D cultures used for testing these substances, in part, contribute to the discrepancies in results. They lack the necessary cell-cell interactions, exhibit altered signaling mechanisms, fail to mimic the natural tumor microenvironment, and show different responses to treatment compared to the reduced malignant phenotype seen in in vivo tumors. Cancer research has undergone a transition to 3-dimensional biological investigations, thanks to recent progress. The relatively low cost and scientific accuracy of 3D cancer cell cultures make them a valuable tool for studying cancer, effectively reproducing the in vivo environment more accurately than their 2D counterparts. This chapter emphasizes the significance of 3D culture, particularly 3D spheroid culture, by reviewing key spheroid formation methodologies, examining the instrumental tools compatible with 3D spheroids, and concluding with their applications in oncology.

Air-liquid interface (ALI) cell cultures are a valid and valuable method for replacing animals in biomedical research applications. ALI cell cultures, in mimicking the essential features of human in vivo epithelial barriers (specifically the lung, intestine, and skin), enable the development of appropriate structural architectures and functional differentiation in normal and diseased tissue barriers. As a result, ALI models closely resemble tissue conditions, generating responses comparable to those seen within a living system. Upon their implementation, these methods have seen widespread adoption in various applications, from toxicity screening to cancer investigations, receiving a substantial degree of acceptance (and sometimes regulatory endorsement) as an appealing alternative to animal testing. This chapter will provide an overview of ALI cell cultures, explaining their application in cancer cell culture, and elaborating on both the positive and negative aspects of this model.

In spite of substantial advancements in both investigating and treating cancer, the practice of 2D cell culture remains indispensable and undergoes continuous improvement within the industry's rapid progression. 2D cell culture, from fundamental monolayer cultures and functional assays to innovative cell-based cancer treatments, is indispensable for cancer diagnosis, prognosis, and therapy. While optimization of research and development is paramount in this field, cancer's diverse nature compels the need for precision medicine approaches adapted for individual patients.