Immuno-reactive cancer organoid model to assess effects of the microbiome on cancer immunotherapy.

Immune checkpoint blockade (ICB) therapy has demonstrated good efficacy in many cancer types. In cancers such as non-resectable advanced or metastatic triple-negative breast cancer (TNBC), it has recently been approved as a promising treatment. However, clinical data shows overall response rates (ORRs) from ~ 3-40% in breast cancer patients, depending on subtype, previous treatments, and mutation status. Composition of the host-microbiome has a significant role in cancer development and therapeutic responsiveness. Some bacterial families are conducive to oncogenesis and progression, while others aid innate and therapeutically induced anti-tumor immunity. Modeling microbiome effects on anti-tumor immunity in ex vivo systems is challenging, forcing the use of in vivo models, making it difficult to dissect direct effects on immune cells from combined effects on tumor and immune cells. We developed a novel immune-enhanced tumor organoid (iTO) system to study factors affecting ICB response. Using the 4T1 TNBC murine cell line and matched splenocytes, we demonstrated ICB-induced response. Further administration of bacterial-derived metabolites from species found in the immunomodulatory host-microbiome significantly increased ICB-induced apoptosis of tumor cells and altered immune cell receptor expression. These outcomes represent a method to isolate individual factors that alter ICB response and streamline the study of microbiome effects on ICB efficacy.

Organoid Platform in Preclinical Investigation of Personalized Immunotherapy Efficacy in Appendiceal Cancer: Feasibility Study.

PURPOSE: Immunotherapy efficacy data on appendiceal cancer from clinical trials does not exist, due to appendiceal cancer incidence of 0.97 per 100,000. The goal of this study was to preclinically explore the application of immunotherapy in treating appendiceal cancer in a personalized organoid model. EXPERIMENTAL DESIGN: Patient tumor organoids (PTO) were fabricated using unsorted tumor cells with and without enrichment with patient-matched immune components derived from peripheral blood leukocytes, spleen, or lymph nodes [immune-enhanced PTOs (iPTO)]. Organoids were cultured for 7 days, followed by treatment with immunotherapy (pembrolizumab, ipilimumab, nivolumab), and assessed for treatment efficacy. RESULTS: Between September 2019 and May 2021, 26 patients were enrolled in the study. Successful testing was conducted in 19 of 26 (73.1%) patients, with 13 of 19 (68.4%) and 6 of 19 (31.6%) patients having low-grade appendiceal (LGA) and high-grade appendiceal (HGA) primaries, respectively. Immunotherapy response, with increased expression of granzyme B and cleaved caspase 3 and decreased expression of CK20 and ATP activity, was exhibited in 4 of 19 (21.1%) pembrolizumab-treated and 2 of 19 (10.5%) nivolumab-treated iPTOs. Post-immunotherapy cellular viability, in responding HGA organoids to pembrolizumab, decreased to less than 15% (P < 0.05). LGA iPTO treatment responses were observed in pembrolizumab and nivolumab, with an 8%-47.4% (P < 0.05) viability compared with controls. Ipilimumab showed no efficacy in the examined cohort. CONCLUSIONS: Immunotherapy shows measurable efficacy in appendiceal cancer organoids. Information derived from immunocompetent organoids may be applied in selecting patients for clinical trial enrollment in rare diseases where preclinical models of disease are lacking.

Simulating the human colorectal cancer microenvironment in 3D tumor-stroma co-cultures in vitro and in vivo.

The tumor microenvironment (TME) plays a significant role in cancer progression and thus modeling it will advance our understanding of cancer growth dynamics and response to therapies. Most in vitro models are not exposed to intact body physiology, and at the same time, fail to recapitulate the extensive features of the tumor stroma. Conversely, animal models do not accurately capture the human tumor architecture. We address these deficiencies with biofabricated colorectal cancer (CRC) tissue equivalents, which are built to replicate architectural features of biopsied CRC tissue. Our data shows that tumor-stroma co-cultures consisting of aligned extracellular matrix (ECM) fibers and ordered micro-architecture induced an epithelial phenotype in CRC cells while disordered ECM drove a mesenchymal phenotype, similar to well and poorly differentiated tumors, respectively. Importantly, co-cultures studied in vitro, and upon implantation in mice, revealed similar tumor growth dynamics and retention of architectural features for 28 days. Altogether, these results are the first demonstration of replicating human tumor ECM architecture in ex vivo and in vivo cultures.

In Vitro Modeling of the Tumor Microenvironment in Tumor Organoids.

BACKGROUND: The tumor microenvironment (TME) represents the many components occupying the space within and surrounding a tumor, including cells, signaling factors, extracellular matrix, and vasculature. Each component has the potential to assume many forms and functions which in turn contribute to the overall state of the TME, and further contribute to the progression and disposition of the tumor itself. The sum of these components can drive a tumor towards progression, keep a migratory tumor at bay, or even control chemotherapeutic response. The wide potential for interaction that the TME is an integral part of a tumor's ecosystem, and it is imperative to include it when studying and modeling cancer in vitro. Fortunately, the development of tissue engineering and biofabrication technologies and methodologies have allowed widespread inclusion of TME-based factors into in vitro tissue-equivalent models. METHODS: In this review, we compiled contemporary literature sources to provide an overview of the field of TME models, ranging from simple to complex. RESULTS: We have identified important components of the TME, how they can be included in in vitro study, and cover examples across a range of cancer types. CONCLUSION: Our goal with this text is to provide a foundation for prospective research into the TME.

Bioengineered Tumor Organoids.

Recent advances in biofabrication technologies, such as cell culture systems, and biomaterials have led to the development of three-dimensional (3D) cell culture platforms, such as tumor organoids. Tumor organoids are more physiologically accurate to the in vivo system, which they are intended to model, compared with traditional 2D cancer cell culture systems. Tumor organoids can mimic pathological and physical characteristics of tumors as well as maintain genetic stability of the cancer cells. Furthermore tumor organoids have advantage over animal models, being made from human cells and easily controlled in the laboratory to attain the desired tissue characteristics. In this section, we describe general tumor organoid technologies, the importance of the tumor microenvironment (TME) in model culture systems, and the use of tumor organoids in drug development and precision medicine. Organoid technologies continue to develop rapidly for applications in academic, clinical, and pharmaceutical settings.

Oncogenic Addiction to ERBB2 Signaling Predicts Response to Trastuzumab in Urothelial Cancer.

Urothelial carcinoma (UC) is a common and frequently lethal cancer. Despite the presence of genomic alterations creating dependency on particular signaling pathways, the use of targeted therapies in advanced and metastatic UC has been limited. We performed an integrated analysis of whole-exome and RNA sequencing of primary and metastatic tumors in a patient with platinum-resistant UC. We found a strikingly high ERBB2 mRNA expression and enrichment of downstream oncogenic ERBB2 signaling in this patient's tumors compared with tumors from an unselected group of patients with UC (N=17). This patient had an exceptional sustained response to trastuzumab. Our findings show that oncogenic addiction to ERBB2 signaling potentially predicts response to ERBB2-directed therapy of UC.