Changes in the way that tumor cells use nutrients to produce energy is a hallmark of cancer (Hanahan and Weinberg, 2011). Yet, the relentless search for metabolic features specific to cancer cells, which could then become therapeutic targets, has faced many challenges (Martínez-Reyes and Chandel, 2021) – including concerns around toxicity and the ability for these cells to easily alter their metabolism to resist therapy.
The fact that the metabolism of lab-grown tumor cells may differ from those growing inside the body has also presented a serious hurdle. What food type is available in the immediate surrounding, or ‘microenvironment’, of a cell can have an immense impact on which nutrients it takes up and metabolizes, and how. Therefore, studying isolated tumor cells exposed to nutrient levels that do not match what they normally encounter in their microenvironment could lead to misleading results that hinder how scientists can both understand and treat cancer.
Some researchers have attempted to optimize the media cancer cells are cultured in so that it mimics the composition of nutrients circulating in human blood (Cantor et al., 2017; Vande Voorde et al., 2019). In 2019, a group – led by Matthew Vander Heiden and Alexander Muir – decided to take this one step further and profiled the metabolic content in the interstitial fluid immediately surrounding tumor cells in mice genetically modified to have pancreatic ductal adenocarcinoma (PDA), the most common form of pancreatic cancer (Sullivan et al., 2019). This revealed that some nutrients are present at different levels than in blood. Now, in eLife, two teams in the United States – one led by Alexander Muir with Juan Apiz-Saab and Lindsey Dzierozynski as co-first authors, and one led by Costas Lyssiotis and Marina Pasca di Magliano with Rosa Menjivar as first author – report how the nutrients available in the microenvironment of pancreatic tumors shape the metabolism of cancer cells and protect their growth (Apiz Saab et al., 2023; Menjivar et al., 2023).
Apiz-Saab, Dzierozynski et al. (who are based at the University of Chicago and University of Michigan) used the nutrient composition of the interstitial fluid of PDA tumors as a template to generate a customized medium for cancer cells named TFIM (short for Tumor Interstitial Fluid Medium). The medium contained plenty of glutamine, an amino acid preferred by tumors which is also abundant in blood, but was significantly depleted in arginine compared to blood plasma. While some differences remained (for instance, tumors in vivo contain a greater range of cell types), PDA cells cultured in TIFM adopted a cellular state similar to that observed in tumors growing inside a host.
Further investigations revealed that TIFM-grown cells were able to activate the urea cycle, a series of biochemical reactions that normally occur in the liver. This metabolic process allowed the cancer cells to produce their own arginine, which may explain why PDA cells are able to survive in a tumor microenvironment lacking this amino acid (Figure 1A). The cells relied on the presence of the intermediary molecule citrulline, which is present in TIFM but absent in standard cancer cell media, to be able to complete the urea cycle. This result was further supported by in vivo experiments conducted in mice bearing PDA tumors in their pancreas.
By closely mimicking the nutrient composition of the tumor microenvironment, TIFM allows researchers to better understand how PDA cells overcome metabolic challenges, like low levels of arginine. It also highlights metabolic vulnerabilities which would be missed when using standard media, such as the fact that cancer cells may depend on the urea cycle to synthesize the arginine they need to grow. However, TIFM does not explain why arginine is depleted inside the PDA tumor microenvironment.
Menjivar et al. (who are based at the University of Michigan, Henry Ford Pancreatic Cancer Center, Rogel Cancer Center, and University of California Irvine) were able to provide an answer to this question by building on previous work they had done on myeloid cells, a group of immune cells which support cancer progression (Zhang et al., 2017). Their 2017 study showed that early malignant cells with a specific mutation could induce the expression of the enzyme arginase 1 (ARG1) in myeloid cells surrounding them. However, whether the enzyme’s ability to breakdown arginine influences tumor growth had not been fully explored.
Menjivar et al. found that a high number of myeloid cells expressing the gene for ARG1 were present in the microenvironment of human and mice pancreatic tumors. Depleting this gene from the myeloid cells of mice resulted in higher levels of arginine, an effect known to alter the fitness of immune cells (Geiger et al., 2016). This helped to recruit anti-cancer CD8+ T cells to the tumor microenvironment and delayed disease progression.
However, many of these mice still developed invasive pancreatic cancer, which Menjivar et al. attribute to two factors. First, a compensatory mechanism which allows myeloid cells to produce the mitochondrial form of arginase known as ARG2, and pancreatic cells to synthetize ARG1. Second, a large portion of the CD8+ T cells infiltrating the PDA tumors are in fact ‘exhausted’ and not fully functioning. The compensatory expression of ARG1 and ARG2 could be counteracted by pharmacologically inhibiting all extracellular arginases; however, tumor reduction was most pronounced when the arginase inhibitor CB-1158 was combined with a drug that could reactivate exhausted CD8+ T cells and boost their anti-tumor role (Figure 1B).
The findings of Menjivar et al. demonstrate why arginine is depleted in the pancreatic tumor environment, and how this change in nutrient availability affects immune cells and influences tumor progression. In turn, arginine depletion formed the basis of the customized medium formulated by Apiz Saab, Dzierozynski et al., which allowed them to probe the metabolic changes associated with PDA.
Intriguingly, Apiz Saab, Dzierozynski et al. found that tumor cells sustained their urea cycle activity to produce arginine even when myeloid cells could not produce arginase anymore, and the amino acid was available in the environment. Not rapidly taking advantage of this new source of arginine may seem counterintuitive and energetically costly for the cells, but maintaining their own production may be beneficial. For example, maintaining an active urea cycle could help cancer cells dispose of the nitrogen they accumulate due to their malignant activity, or allow them to better channel arginine into cancer-promoting processes (Zaytouni et al., 2017).
In addition, not metabolically re-adapting from a nutrient-starved to a nutrient-replete state could protect cancer cells from sudden fluctuations in nutrient availability that could lead to cell cycle arrest or death. Such changes could result from growing tumors experiencing rapid alterations in their three-dimensional landscape which alter how close they are to blood vessels or other cell types that deplete the environment of specific nutrients. Consistent with this idea, PDA cells were recently shown to unconventionally rely on glutamine rather than arginine to synthetize the precursors needed to build polyamines, a class of molecules important in cancer progression. This unusual glutamine-derived pathway persisted even when arginine became widely available, suggesting that the cells had been rewired to favor this biochemical process (Lee et al., 2023).
Taken together, these two studies show how investigating the behavior of pancreatic cancer cells in response to nutrient availability can lead to a deeper understanding of the biology of an aggressive, uncooperative disease. In particular, they have unveiled metabolic challenges and vulnerabilities that could become therapeutic targets. Studying the ever-changing nutrient composition of the tumor microenvironment should prove to be a promising tool to identify the distinct metabolic features that cancer cells rely on in a range of tissues and at various stages of the disease.
An extensive fibroinflammatory stroma rich in macrophages is a hallmark of pancreatic cancer. In this disease, it is well appreciated that macrophages are immunosuppressive and contribute to the poor response to immunotherapy; however, the mechanisms of immune suppression are complex and not fully understood. Immunosuppressive macrophages are classically defined by the expression of the enzyme Arginase 1 (ARG1), which we demonstrated is potently expressed in pancreatic tumor-associated macrophages from both human patients and mouse models. While routinely used as a polarization marker, ARG1 also catabolizes arginine, an amino acid required for T cell activation and proliferation. To investigate this metabolic function, we used a genetic and a pharmacologic approach to target Arg1 in pancreatic cancer. Genetic inactivation of Arg1 in macrophages, using a dual recombinase genetically engineered mouse model of pancreatic cancer, delayed formation of invasive disease, while increasing CD8+ T cell infiltration. Additionally, Arg1 deletion induced compensatory mechanisms, including Arg1 overexpression in epithelial cells, namely Tuft cells, and Arg2 overexpression in a subset of macrophages. To overcome these compensatory mechanisms, we used a pharmacological approach to inhibit arginase. Treatment of established tumors with the arginase inhibitor CB-1158 exhibited further increased CD8+ T cell infiltration, beyond that seen with the macrophage-specific knockout, and sensitized the tumors to anti-PD1 immune checkpoint blockade. Our data demonstrate that Arg1 drives immune suppression in pancreatic cancer by depleting arginine and inhibiting T cell activation.
Identification oncogenes is fundamental to revealing the molecular basis of cancer. Here, we found that FOXP2 is overexpressed in human prostate cancer cells and prostate tumors, but its expression is absent in normal prostate epithelial cells and low in benign prostatic hyperplasia. FOXP2 is a FOX transcription factor family member and tightly associated with vocal development. To date, little is known regarding the link of FOXP2 to prostate cancer. We observed that high FOXP2 expression and frequent amplification are significantly associated with high Gleason score. Ectopic expression of FOXP2 induces malignant transformation of mouse NIH3T3 fibroblasts and human prostate epithelial cell RWPE-1. Conversely, FOXP2 knockdown suppresses the proliferation of prostate cancer cells. Transgenic overexpression of FOXP2 in the mouse prostate causes prostatic intraepithelial neoplasia. Overexpression of FOXP2 aberrantly activates oncogenic MET signaling and inhibition of MET signaling effectively reverts the FOXP2-induced oncogenic phenotype. CUT&Tag assay identified FOXP2-binding sites located in MET and its associated gene HGF. Additionally, the novel recurrent FOXP2-CPED1 fusion identified in prostate tumors results in high expression of truncated FOXP2, which exhibit a similar capacity for malignant transformation. Together, our data indicate that FOXP2 is involved in tumorigenicity of prostate.