Unresolved endoplasmic reticulum stress engenders immune-resistant, latent pancreatic cancer metastases

INTRODUCTION Pancreatic ductal adenocarcinoma (PDA) is the fourth most common cause of death from cancer worldwide and has a 5-year survival rate of 6%. Patients who have had their primary PDA surgically resected often develop metastatic disease, despite intra-operative examination of the liver confirming the absence of macrometastatic lesions. These observations lead to the conclusion that latent metastases, detectable only microscopically, were present in these patients and were responsible for the postoperative development of metastatic disease. RATIONALE Latent metastases with the potential for outgrowth had been considered to be lesions in which cancer cell proliferation is balanced by immune-mediated cancer cell death, but a more recent explanation invokes quiescent, single disseminated cancer cells (DCCs). Single, nonreplicating DCCs have been observed in several cancer types, but whether quiescence is enforced by the microenvironment or is cancer cell–autonomous is not known. Immunity, both innate and adaptive, also is likely to have a role in the selection and/or maintenance of latent DCCs. This has long been suspected on the basis of the clinical observation of donor-derived cancer in immune-suppressed recipients of allografts. However, there is an unexplained paradox of immunity preventing the outgrowth of latent metastases but not eliminating them. RESULTS We studied the metastatic process in the context of an ongoing adaptive immune response because of the occurrence of cancer cell–specific immunity in human and mouse PDA. Livers from patients and mice with PDA contained single DCCs with an unusual phenotype of being negative for cytokeratin 19–negative (CK19) and major histocompatibility complex class I (MHCI). The absence of the expression of MHCI in DCCs and the occurrence of cancer-specific CD8 + T cells in the genetically engineered mouse model of PDA, and possibly in patients with PDA, suggested that DCCs may be selected by an anticancer immune response during the metastatic process. To investigate this hypothesis, we created a mouse model that would allow us to determine how DCCs develop, their relationship to metastatic latency, and the role of immunity. Intraportal injection of immunogenic PDA cells into preimmunized mice seeded livers only with single, nonreplicating DCCs lacking MHCI and CK19, whereas naive recipients of PDA cells had macrometastases. We found that a subpopulation of PDA cells with the phenotype of DCCs was present in vitro and that those cells are the precursors of DCCs in vivo. We found that T cells select DCCs by eliminating MHCI + proliferating cancer cells. To identify the cell-autonomous “switch” regulating the developmental state of the metastases, we preformed single-cell RNA sequencing of PDA cells with the DCC phenotype. This transcriptomic analysis demonstrated an endoplasmic reticulum (ER) stress response. Moreover, DCCs showed a lack of activation of the IRE1α (inositol-requiring enzyme 1α) pathway of the unfolded protein response, whereas the PERK (protein kinase RNA-like ER kinase) pathway was activated, suggesting that DCCs cannot resolve ER stress. Relieving ER stress pharmacologically with a chemical chaperone or genetically by overexpression of spliced XBP1, in combination with T cell depletion, stimulated outgrowth of macrometastatic lesions containing PDA cells expressing MHCI and CK19. CONCLUSION We find that a PDA-specific adaptive immune response selects DCCs, in which the ER stress response accounts for both quiescence and resistance to immune elimination. Accordingly, outgrowth of DCCs to macrometastases requires not only relief from the cancer cell–autonomous ER stress response, but also suppression of systemic immunity. Thus, the ER stress response is a cell-autonomous reaction that enables DCCs to escape immunity and establish latent metastases.