The Seeds and the Soil: Roles of Tumor Heterogeneity and the Tumor Microenvironment in Cancer Metastasis

Lecture Summaries

week #	topics	lecture summaries
1	Introduction	During the initial session, we will introduce ourselves and outline the objectives, structure, and expectations for the course. We will discuss general strategies for accessing and reading scientific literature. The instructors will provide a brief overview of the general principles of cancer biology and metastasis, followed by an introduction to the topic for the following week.
2	Epithelial-Mesenchymal Transition and Dissemination from the Primary Tumor	Most cancers arise in epithelial tissues (carcinomas), consisting of tightly interconnected, non-motile cells that form protective barriers that line the external surfaces and inner cavities of organs, including breast cancer, lung cancer, colorectal cancer, pancreatic cancer, prostate cancer and others. Carcinomas generally maintain key cellular features of their epithelial origins, including extensive cell-cell contacts. How are some cancer cells (known as neoplastic cells) able to break free of these contacts and escape from the primary tumor? In many cases, and as we will see from the first paper (Yang et al., 2004), cancer cells co-opt a latent developmental program called the epithelial-mesenchymal transition (EMT), which normally functions during embryonic development, to enable their dissemination from the primary tumor and subsequent metastatic growth. The second paper (Janda et al., 2002) was one of the earliest studies to identify the signaling pathways utilized by carcinoma cells to activate the EMT program and facilitate their invasion and metastasis.
3	Cancer in Transit: Dynamics and Behaviors of Circulating Tumor Cells (CTCs)	Once neoplastic cells have escaped from the primary tumor, they can enter and travel through the vasculature as circulating tumor cells (CTCs), thereby gaining access to new tissues at distant anatomical sites. Although their residence in circulation might be only a few minutes, the interactions that CTCs experience during this time can be critical determinants of whether or not they will succeed in founding a metastatic colony. Although the total number of CTCs is often exceedingly low, technologies have been developed that allow for their capture and analysis. The first paper (Yu et al., 2013) describes how the analysis of CTCs from human patients can be used to understand both the dynamic clinical response to chemotherapy and the underlying biology of cancer dissemination. The second paper (Cheung et al., 2016) describes how tumor cells can disseminate and travel together in the circulation as multicellular clusters that are highly metastatic.
4	Stromal Support of Dissemination	Interactions of cancer cells with stromal cells – various non-neoplastic mesenchymal cells found in within the tumor (e.g., fibroblasts, endothelial cells, immune cells) – are important determinants of cancer progression toward metastasis. Various tumor-associated stromal cell types have been shown to support cancer cell dissemination, survival in transition, and colonization at the distant sites. This week we will discuss two papers exploring the interactions of cancer cells with stromal cells in the primary tumor site and during intravascular transit to the site of metastasis. The first paper (Lin et al., 2001) demonstrates how a certain type of macrophage (a phagocytic cell of the innate immune system), recruited to the tumor by cancer cells that secrete the protein CSF-1, can promote the progression to malignancy. The second paper (Labelle et al., 2011) explores the interactions between circulating tumor cells and blood-clotting platelets, specifically how platelets can prime tumor cells for subsequent metastasis via activation of signaling by the protein TGFβ and EMT.
5	EMT Plasticity and Metastatic Colonization	In Session 2, we discussed the importance of the epithelial-mesenchymal transition (EMT) program in carcinoma cell dissemination. Interestingly, however, the EMT program is not a stereotypical, unidirectional cell state change: there are multiple intermediate cell states that exist between the extreme epithelial and the extreme mesenchymal poles, and the residence in these states is often transient as cells exhibit E-M plasticity. This plastic nature of the EMT program contributes to the robust outgrowth of metastatic colonies. Indeed, in certain organ sites, some degree of mesenchymal-epithelial (MET) transition is required for successful colonization. The two papers this week discuss the topic of EMT plasticity and the requirement of a MET process during metastatic colonization. The first paper (Ocana et al., 2012) demonstrates that repression of an EMT-inducing transcription factor, Prrx1, which is commonly associated with the full mesenchymal state, facilitates metastatic colonization in breast and kidney cancer models. The second manuscript (Pastushenko et al., 2018) presents a more recent view of the EMT program, which in fact can generate a spectrum of mesenchymal cell types possessing important functional differences.
6	Cancer Stem Cells	The concept of cancer stem cells (CSCs) in carcinomas is based on the observation that phenotypically distinct subpopulations of cancer cells coexist within a single tumor, with a small number of cancer cells showing certain similarities to non-neoplastic stem cells. These stem-like properties include enhanced self-renewal and an ability to regenerate the heterogeneity of the original neoplastic tumor tissue, including more differentiated non-CSC derivatives. In this paradigm, CSCs are hypothesized to play essential roles in sustaining tumor growth and metastasis. In this session, we will discuss the concepts and experimental evidence regarding CSCs and their role in metastasis. The first paper (de Sousa e Melo et al., 2017) describes a population of Lgr5+ CSCs (a stem cell population defined by surface expression of the Lgr5 protein) in colon cancer models, comparing their different functions in primary tumor growth and formation of metastases. The second paper (Famagalli et al., 2020) provides recent evidence that in a colon cancer model, metastatic nodules are commonly initiated from non-CSCs that are plastic and convert into CSCs upon dissemination to distant organs.
7	Genetic Heterogeneity and Evolution of Metastatic Disease	The phenotypic diversity of neoplastic cells within a tumor, also known as tumor heterogeneity, is increasingly considered a major obstacle to the success of cancer therapies. Both genetic and epigenetic mechanisms contribute to phenotypic heterogeneity within individual tumors. With recent advances in next-generation sequencing technologies, the impact and influence of genetic heterogeneity has been widely recognized and, for many tumor types, tumor evolution can be mapped by studying the patterns of acquired somatic mutations that occur as cancer cells in their primary tumor site progress to later stages of malignancy. However, it remains unclear exactly how metastatic disease evolves over time or if any metastasis-specific mutations exist. This week we will read two papers that explore this question using clinical samples from different types of human cancer. The first paper (Turajlic et al., 2018) uses sequencing of matched primary and metastatic tumors from patients with renal cell carcinoma to track the the evolution of metastatic disease, pinpointing chromosome complexity rather than new driver mutations as key determinants of metastatic competence. In the second paper (Reiter et al., 2018), the authors sequence metastases from untreated patients and find that the functional driver mutations are largely shared by the primary tumor and all metastases in a given patient.
8	Field Trip	This week we will virtually visit scientists working at Agios, a Cambridge-based biotechnology company that is developing new therapeutic agents for patients with cancer. Their approach is based on understanding and targeting various aspects of tumor metabolism implicated in facilitating cancer progression in addition to the fueling primary tumor growth. Their initial program involved patients with a mutation in isocitrate deyhydrogenase (IDH), but they have since expanded their research portfolio and clinical pipeline to include metabolic immuno-oncology and rare metabolic disorders. This visit will allow students to see firsthand how basic biological insights can be turned into new drugs that benefit patients with cancer and will offer a perspective about possible future career options. The paper for this week (Dang et al., 2009) describes a seminal discovery concerning how mutant IDH contributes to cancer through the production of an oncometabolite.
9	Determinants of Metastatic Tropism	Primary tumors often have a characteristic pattern of metastatic spread that differs across cancer types. This tendency to colonize certain organs is referred to as metastatic tropism. For example, while prostate cancer preferentially metastasizes to the bone, colon cancer most often spawns metastases in the liver. Metastatic tropism is determined by the frequency and number of carcinoma cells that seed an organ and the ease with which these cells can adapt to their new environment. This week we will learn about the types of programs and mechanisms that facilitate organ-specific metastasis. The first paper (Valiente et al., 2014) describes how cancer cells that infiltrate the brain are able to survive and subsequently initiate brain metastases. The second paper (Reichert et al., 2018) demonstrates that for pancreatic cancer cells there are different requirements for colonization in the liver or the lung. Pancreatic cancer cells that are able to complete a MET tend to metastasize to the liver, whereas cells retaining more mesenchymal features tend to colonize in the lung.
10	The Metastatic Niche	After extravasation from the bloodstream and entry into a distant organ, disseminated tumor cells (DTCs) may occupy specialized niches, which support their survival and subsequent outgrowth. This week we will explore two different types of niches – one in the bone, the other in the lung – and two different strategies that carcinoma cells use to “find” these niches. The first paper (Shiozawa et al., 2011) describes how disseminated prostate cancer cells home to the hematopoietic stem cell niche, where they directly compete with hematopoietic stem cells for residence in this favored location. The second paper (Malanchi et al., 2011) describes an alternative strategy, whereby breast cancer cells that have disseminated to the lung are able to create their own supportive niche.
11	Systemic Effects and the Pre-metastatic Niche	For several types of cancer, accumulating evidence suggests the existence of metastasis-supportive microenvironments termed pre-metastatic niches, which form at the sites of future metastases to allow the colonization and growth of disseminated tumor cells. The formation of pre-metastatic niches is induced by signals secreted by specific subsets of primary tumor cells and usually through modulating the distribution and function of immune cells, either systemically or at the sites of future metastases. Two papers exploring such process will be discussed this week. The first paper (Costa-Silva et al., 2015) demonstrates that exosomes, one type of tiny membrane-bound extracellular vesicles, secreted by malignant pancreatic cancer cells elicit a pre-metastatic niche in the liver through a multi-step process. The second paper (Elkabets et al., 2011) reports that certain malignant breast tumors can stimulate the growth of a distant otherwise quiescent tumor in an experimental animal by activating and mobilizing a specific subtype of bone-marrow cells into the stroma of the once-quiescent tumor.
12	Interactions Between Dormant DTCs and the Immune System	Some disseminated tumor cells (DTCs), upon reaching a distant organ, do not outgrow or die, but instead persist in an indolent state referred to as dormancy. Such dormant DTCs represent ticking time bombs; indeed, metastases can appear numerous years – sometimes decades – after an ostensibly successful treatment of the primary tumor. How do these dormant DTCs persist within tissues for extended periods of time? And what is preventing them from actively growing into a metastatic lesion? Recent work indicates that the growth of DTCs might be held in check by components of the immune system and that DTCs that persist in the tissue are somehow spared from complete elimination. The first paper (Malladi et al, 2016) describes how slow-cycling dormant DTCs downregulate receptors that would normally trigger their elimination by natural killer cells of the innate immune system.  The second paper (Pommier et al., 2018) finds that quiescent DTCs exhibit chronic endoplasmic reticulum stress, which allows them to avoid detection by T cells of the adaptive immune system.
13	Oral Presentations and Closing Remarks	Students will deliver their oral presentations, allowing for questions and discussion with the class. We will reflect on what we have learned in the class and discuss where the field may be going next. Finally, students will complete course and instructor evaluations.