Detailed phylogenies of tumor populations can recount the history and chronology of critical events during cancer progression, such as metastatic dissemination. We applied a Cas9-based, single-cell lineage tracer to study the rates, routes, and drivers of metastasis in a lung cancer xenograft mouse model. We report deeply resolved phylogenies for tens of thousands of cancer cells traced over months of growth and dissemination. This revealed stark heterogeneity in metastatic capacity, arising from pre-existing and heritable differences in gene expression. We demonstrate that these identified genes can drive invasiveness, and uncovered an unanticipated suppressive role for KRT17. We also show that metastases disseminated via multidirectional tissue routes and complex seeding topologies. Overall, we demonstrate the power of tracing cancer progression at subclonal resolution and vast scale.

https://science.sciencemag.org/content/sci/early/2021/01/21/science.abc1944.full.pdf

Single-cell lineages reveal the rates, routes, and drivers of metastasis in cancer xenografts.

Cancer progression is characterized by rare, transient events which are nonetheless highly consequential to disease etiology and mortality. Detailed cell phylogenies can recount the history and chronology of these critical events - including metastatic seeding. Here, we applied our Cas9-based lineage tracer to study the subclonal dynamics of metastasis in a lung cancer xenograft mouse model, revealing the underlying rates, routes, and drivers of metastasis. We report deeply resolved phylogenies for tens of thousands of metastatically disseminated cancer cells. We observe surprisingly diverse metastatic phenotypes, ranging from metastasis-incompetent to aggressive populations. These phenotypic distinctions result from pre-existing, heritable, and characteristic differences in gene expression, and we demonstrate that these differentially expressed genes can drive invasiveness. Furthermore, metastases transit via diverse, multidirectional tissue routes and seeding topologies. Our work demonstrates the power of tracing cancer progression at unprecedented resolution and scale.

Single-cell lineages reveal the rates, routes, and drivers of metastasis in cancer xenografts

The regenerating islet-derived (REG) gene family encodes a group of proteins highly expressed in several human pathologies, many of which are associated with epithelial inflammation. All human family members, namely REG1A, REG1B, REG3A and REG4, are closely related in genomic sequence and all are part of the c-type lectin superfamily. REGs are highly expressed during inflammatory bowel disease (IBD)-related colonic inflammation and the in vivo expression pattern of REG1A and REG4 has been localised by using immunohistochemistry. However, the function of the encoded proteins is largely unknown and the cellular localisation of REG expression during colonic inflammation has not been described. Therefore, we have used in situ hybridisation to demonstrate the cellular localisation of REG expression in healthy and diseased colonic mucosa. Samples drawn from an IBD cohort including both inflamed and un-inflamed colonic mucosa are described, as are samples from non-IBD inflammation and healthy controls. Immunohistochemistry against known cell-type markers on serial sections has localised the expression of REGs to metaplastic Paneth cells (REG1A, REG1B and REG3A) and enteroendocrine cells (REG4), with a marked expansion of expression during inflammation. The group of REGs can, based on gene expression patterns, be divided into at least two groups; REG1A, REG1B and REG3A with their expression focused in the crypt base spreading from Paneth cells and REG4 being more highly expressed towards the luminal face. This exploration of expression pattern forms provides the background for further exploration of REG function in the intestine.

/pdf/reg-gene-expression-in-inflamed-and-healthy-colon-mucosa-2ay7owbt89.pdf

REG gene expression in inflamed and healthy colon mucosa explored by in situ hybridisation

Background

This study examined the clinical significance of regenerating islet-derived family member 4 (REGIV) in surgically resected gastric tumors The potential of REGIV as a biomarker in gastric cancer was also assessed including its predictive value for prognosis and recurrence after surgery



Methods

Immunohistochemistry was performed to assess the clinical significance of REGIV expression status in surgically resected specimens The quantitative genetic diagnostic method, transcription-reverse transcription concerted reaction (TRC) that targeted REGIV mRNA was applied for prediction of peritoneal recurrence in gastric cancer



Results

Positive immunostaining for REGIV was observed in 85 cases (525%), and correlated significantly with diffuse type histopathology (P = 0001), advanced T stage (P = 0022), and frequent peritoneal recurrence (P = 0009) Multivariate analysis identified advanced T stage (P < 0001) and REGIV expression (P = 0034) as independent prognostic factors for peritoneal recurrence-free survival Overexpression of REGIV protein was evident in the majority of peritoneal tumors (938%) REGIV mRNA assessed by TRC could be a predictive marker for peritoneal recurrence after curative operation



Conclusions

REGIV overexpression is common in primary gastric tumors and a potentially suitable marker of diffuse type histopathology and peritoneal dissemination Overexpression of REGIV mRNA, assessed by the TRC method, is a potentially suitable marker of peritoneal recurrence after curative resection J Surg Oncol 2012; 105:189–194 © 2011 Wiley Periodicals, Inc

REGIV as a potential biomarker for peritoneal dissemination in gastric adenocarcinoma

Regenerating islet-derived family member, 4 (Reg IV) is a secreted protein and member of the C-type lectin superfamily. Expression analyses have characterized Reg IV as a prognostic marker for certain cancers; however, the functional role of Reg IV in cancer, including downstream signaling, has only begun to be elucidated. To investigate the biological role of Reg IV in cancer, phosphorylation events were studied in cancer cell lines in the context of either Reg IV stimulation (HCT116 cells) or knockdown of endogenous Reg IV (PC3 and KM12 cells). In addition to the previously observed impact on epidermal growth factor receptor and Akt phosphorylation, we observed modulation in the phosphorylation of multiple additional receptor tyrosine kinases (RTKs), including insulin receptor, insulin-like growth factor receptor as well as their downstream effectors, mitogen-activated protein kinase and phosphatidylinositol-3-kinase pathways. Furthermore, knockdown of Reg IV impacted the ability of insulin and EGF to stimulate downstream tyrosine phosphorylation. Knockdown of Reg IV in cancer cell lines inhibited anchorage-dependent and anchorage-independent (both soft-agar and spheroid assays) cell growth and induced cell cycle arrest. This was accompanied by upregulation of p21 and p27. Transiently silencing Reg IV in cancer cells induced apoptosis and downregulated Bcl-2. Conversely, stimulation of HCT116 cells with recombinant Reg IV induced Bcl-2. Hsp27, a molecule implicated in drug resistance, was similarly modulated by Reg IV. Consistent with our observations with Reg IV siRNA-mediated knockdown, monoclonal antibodies directed against Reg IV inhibited PC3 and KM12 cell growth. Collectively, Reg IV plays an important role in cancer by modulation of key signaling molecules including Hsp27, Bcl-2 and multiple RTKs.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/ijc.26089

Nan Li

Papers

COMMD10 inhibits HIF1α/CP loop to enhance ferroptosis and radiosensitivity by disrupting Cu-Fe balance in hepatocellular carcinoma.

RegIV potentiates colorectal carcinoma cell migration and invasion via its CRD domain.

Exploring prognostic value and regulation network of PPP1R1A in hepatocellular carcinoma