Catecholamines released by sympathetic nerves can activate adrenergic receptors present on nearly every cell type, including myeloid derived suppressor cells (MDSCs). Using in vitro systems and murine tumor models, in wild-type mice and genetically modified (β2-AR^–/–) mice, as well adoptive transfer approaches, we found that the degree of β2-AR signaling significantly influences MDSC frequency and survival in tumors and other tissues, modulates their expression of immunosuppressive molecules such as arginase-I and PDL-1 and alters their ability to suppress the proliferation of T cells. The regulatory functions of β-AR signaling in MDSCs were found to be dependent upon STAT3 phosphorylation. Moreover, we observed that the β2-AR-mediated increase in survival of MDSCs is dependent upon Fas-FasL interactions, and this is consistent with gene expression analyses which reveal a greater expression of apoptosis-related genes in β2-AR^–/– MDSCs. Our data reveals the potential of β2-AR signaling to increase the generation of MDSCs from both murine and human peripheral blood cells and that the immunosuppressive function of MDSCs could be mitigated by treatment with β-AR antagonists, or enhanced by β-AR agonists, strongly supporting the possibility that reducing stress-induced activation of β2-ARs could help to overcome immune suppression and enhance the efficacy of immunotherapy and other cancer therapies.

Metabolic dysfunction and exhaustion in tumor-infiltrating T cells have been linked to ineffectual antitumor immunity and the failure of immune checkpoint inhibitor therapy. We report here that chronic stress plays a previously unrecognized role in regulating the state of T cells in the tumor microenvironment (TME). Using two mouse tumor models, we found that blocking chronic adrenergic stress signaling using the pan β-blocker propranolol or by using mice lacking the β2-adrenergic receptor (β2-AR) results in reduced tumor growth rates with significantly fewer tumor-infiltrating T cells that express markers of exhaustion, with a concomitant increase in progenitor exhausted T cells. We also report that blocking β-AR signaling in mice increases glycolysis and oxidative phosphorylation in tumor-infiltrating lymphocytes (TIL), which associated with increased expression of the costimulatory molecule CD28 and increased antitumor effector functions, including increased cytokine production. Using T cells from Nur77-GFP reporter mice to monitor T-cell activation, we observed that stress-induced β-AR signaling suppresses T-cell receptor (TCR) signaling. Together, these data suggest that chronic stress–induced adrenergic receptor signaling serves as a “checkpoint” of immune responses and contributes to immunosuppression in the TME by promoting T-cell metabolic dysfunction and exhaustion. These results also support the possibility that chronic stress, which unfortunately is increased in many patients with cancer following their diagnoses, could be exerting a major negative influence on the outcome of therapies that depend upon the status of TILs and support the use of strategies to reduce stress or β-AR signaling in combination with immunotherapy.

An immune response must be tightly controlled so that it will be commensurate with the level of response needed to protect the organism without damaging normal tissue. The roles of cytokines and chemokines in orchestrating these processes are well known, but although stress has long been thought to also affect immune responses, the underlying mechanisms were not as well understood. Recently, the role of nerves and, specifically, the sympathetic nervous system, in regulating immune responses is being revealed. Generally, an acute stress response is beneficial but chronic stress is detrimental because it suppresses the activities of effector immune cells while increasing the activities of immunosuppressive cells. In this review, we first discuss the underlying biology of adrenergic signaling in cells of both the innate and adaptive immune system. We then focus on the effects of chronic adrenergic stress in promoting tumor growth, giving examples of effects on tumor cells and immune cells, explaining the methods commonly used to induce stress in preclinical mouse models. We highlight how this relates to our observations that mandated housing conditions impose baseline chronic stress on mouse models, which is sufficient to cause chronic immunosuppression. This problem is not commonly recognized, but it has been shown to impact conclusions of several studies of mouse physiology and mouse models of disease. Moreover, the fact that preclinical mouse models are chronically immunosuppressed has critical ramifications for analysis of any experiments with an immune component. Our group has found that reducing adrenergic stress by housing mice at thermoneutrality or treating mice housed at cooler temperatures with β-blockers reverses immunosuppression and significantly improves responses to checkpoint inhibitor immunotherapy. These observations are clinically relevant because there are numerous retrospective epidemiological studies concluding that cancer patients who were taking β-blockers have better outcomes. Clinical trials testing whether β-blockers can be repurposed to improve the efficacy of traditional and immunotherapies in patients are on the horizon.

Cancer research relies heavily on murine models for evaluating the anti-tumour efficacy of therapies. Here we show that the sensitivity of several pancreatic tumour models to cytotoxic therapies is significantly increased when mice are housed at a thermoneutral ambient temperature of 30 °C compared with the standard temperature of 22 °C. Further, we find that baseline levels of norepinephrine as well as the levels of several anti-apoptotic molecules are elevated in tumours from mice housed at 22 °C. The sensitivity of tumours to cytotoxic therapies is also enhanced by administering a β-adrenergic receptor antagonist to mice housed at 22 °C. These data demonstrate that standard housing causes a degree of cold stress sufficient to impact the signalling pathways related to tumour-cell survival and affect the outcome of pre-clinical experiments. Furthermore, these data highlight the significant role of host physiological factors in regulating the sensitivity of tumours to therapy.

Background: Apo2L/TRAIL has considerable promise for cancer therapy based on the fact that this member of the tumor necrosis factor family induces apoptosis in the majority of malignant cells, while normal cells are more resistant. Furthermore, in many cells, when Apo2L/TRAIL is combined with chemotherapy, the effect is synergistic. The majority of this work has been carried out using cell lines. Therefore, investigation of how patient tumors respond to Apo2L/TRAIL can validate and/or complement information obtained from cell lines and prove valuable in the design of future clinical trials. Methods: We have investigated the Apo2L/TRAIL sensitivity of patient derived pancreatic tumors using a patient tumor xenograft/ SCID mouse model. Mice bearing engrafted tumors were treated with Apo2L/TRAIL, gemcitabine or a combination of both therapies. Results: Patient tumors grown as xenografts exhibited a spectrum of sensitivity to Apo2L/TRAIL. Both Apo2L/TRAIL sensitive and resistant pancreatic tumors were found, as well as tumors that showed heterogeneity of response. Changes in apoptotic signaling molecules in a sensitive tumor were analyzed by Western blot following Apo2L/TRAIL treatment; loss of procaspase 8, Bid and procaspase 3 was observed and correlated with inhibition of tumor growth. However, in a tumor that was highly resistant to killing by Apo2L/TRAIL, although there was a partial loss of procaspase 8 and Bid in response to Apo2L/TRAIL treatment, loss of procaspase 3 was negligible. This resistant tumor also expressed a high level of the anti-apoptotic molecule Bcl-X_L that, in comparison, was not detected in a sensitive tumor. Importantly, in the majority of these tumors, addition of gemcitabine to Apo2L/TRAIL resulted in a greater anti-tumor effect than either therapy used alone. Conclusion: These data suggest that in a clinical setting we will see heterogeneity in the response of patients' tumors to Apo2L/TRAIL, including tumors that are highly sensitive as well as those that are resistant. While much more work is needed to understand the molecular basis for this heterogeneity, it is very encouraging, that Apo2L/TRAIL in combination with gemcitabine increased therapeutic efficacy in almost every case and therefore may be a highly effective strategy for controlling human pancreatic cancer validating and expanding upon what has been reported for cell lines.

Studies of primary patient tumor xenografts grown in immunodeficient mice have shown that these tumors histologically and genetically closely resemble the original tumors. These patient xenograft models are becoming widely used for therapeutic efficacy studies. Because many therapies are directed at tumor stromal components and because the tumor microenvironment also is known to influence the response of a tumor to therapy, it is important to understand the nature of the stroma and, in particular, the vascular supply of patient xenografts. Patient tumor xenografts were established by implanting undisrupted pieces of patient tumors in SCID mice. For this study, formalin fixed, paraffin embedded specimens from several types of solid tumors were selected and, using species-specific antibodies which react with formalin fixed antigens, we analyzed the species origin of the stroma and blood vessels that supported tumor growth in these models. Additionally, we investigated the kinetics of the vascularization process in a colon tumor and a mesothelioma xenograft. In mice bearing a head and neck xenograft, a perfusion study was performed to compare the functionality of the human and mouse tumor vessels. In patient tumors which successfully engrafted, the human stroma and vessels which were engrafted as part of the original tumor did not survive and were no longer detectable at the time of first passage (15–25 weeks). Uniformly, the stroma and vessels supporting the growth of these tumors were of murine origin. The results of the kinetic studies showed that the loss of the human vessels and vascularization by host vessels occurred more rapidly in a colon tumor (by 3 weeks) than in a mesothelioma (by 9 weeks). Finally, the perfusion studies revealed that while mouse vessels in the periphery of the tumor were perfused, those in the central regions were rarely perfused. No vessels of human origin were detected in this model. In the tumors we investigated, we found no evidence that the human stromal cells and vessels contained in the original implant either survived or contributed in any substantive way to the growth of these xenografts.