Plasma cells in human pancreatic ductal adenocarcinoma secrete antibodies against self-antigens.

Intratumoral B cell responses are associated with more favorable clinical outcomes in human pancreatic ductal adenocarcinoma (PDAC). However, the antigens driving these B cell responses are largely unknown. We sought to discover these antigens by using single-cell RNA sequencing (scRNA-Seq) and immunoglobulin (Ig) sequencing of tumor-infiltrating immune cells from 7 primary PDAC samples. We identified activated T and B cell responses and evidence of germinal center reactions. Ig sequencing identified plasma cell (PC) clones expressing isotype-switched and hypermutated Igs, suggesting the occurrence of T cell-dependent B cell responses. We assessed the reactivity of 41 recombinant antibodies that represented the products of 235 PCs and 12 B cells toward multiple cell lines and PDAC tissues and observed frequent staining of intracellular self-antigens. Three of these antigens were identified: the filamentous actin (F-actin), the nucleic protein RuvB like AAA ATPase 2 (RUVBL2), and the mitochondrial protein heat shock protein family D (Hsp60) member 1 (HSPD1). Antibody titers against F-actin and HSPD1 were substantially elevated in the plasma of patients with PDAC compared with healthy donors. Thus, PCs in PDAC produce autoantibodies reacting with intracellular self-antigens, which may result from promotion of preexisting, autoreactive B cell responses. These observations indicate the chronic inflammatory microenvironment of PDAC can support the adaptive immune response.

T cell-Mediated Development of Stromal Fibroblasts with an Immune-Enhancing Chemokine Profile.

Stromal fibroblasts reside in inflammatory tissues that are characterized by either immune suppression or activation. Whether and how fibroblasts adapt to these contrasting microenvironments remains unknown. Cancer-associated fibroblasts (CAF) mediate immune quiescence by producing the chemokine CXCL12, which coats cancer cells to suppress T-cell infiltration. We examined whether CAFs can also adopt an immune-promoting chemokine profile. Single-cell RNA sequencing of CAFs from mouse pancreatic adenocarcinomas identified a subpopulation of CAFs with decreased expression of Cxcl12 and increased expression of the T cell-attracting chemokine Cxcl9 in association with T-cell infiltration. TNFα and IFNγ containing conditioned media from activated CD8+ T cells converted stromal fibroblasts from a CXCL12+/CXCL9- immune-suppressive phenotype into a CXCL12-/CXCL9+ immune-activating phenotype. Recombinant IFNγ and TNFα acted together to augment CXCL9 expression, whereas TNFα alone suppressed CXCL12 expression. This coordinated chemokine switch led to increased T-cell infiltration in an in vitro chemotaxis assay. Our study demonstrates that CAFs have a phenotypic plasticity that allows their adaptation to contrasting immune tissue microenvironments.

T cell-mediated development of stromal fibroblasts with an immune-enhancing chemokine profile.

Stromal fibroblasts reside in inflammatory tissues that are characterized by either immune suppression or activation. Whether and how fibroblasts adapt to these contrasting microenvironments remains unknown. Cancer-associated fibroblasts (CAFs) mediate immune quiescence by producing the chemokine CXCL12, which coats cancer cells to suppress T-cell infiltration. We examined whether CAFs can also adopt an immune-promoting chemokine profile. Single-cell RNA-sequencing of CAFs from mouse pancreatic adenocarcinomas identified a sub-population of CAFs with decreased expression of Cxcl12 and increased expression of the T cell-attracting chemokine Cxcl9 in association with T-cell infiltration. TNFα and IFNγ containing conditioned media from activated CD8+ T cells converted stromal fibroblasts from a CXCL12+/CXCL9- immune suppressive phenotype into a CXCL12-/CXCL9+ immune-activating phenotype. Recombinant IFNγ and TNFα acted together to augment CXCL9 expression, whereas TNFα alone suppressed CXCL12 expression. This coordinated chemokine switch led to increased T-cell infiltration in an in vitro chemotaxis assay. Our study demonstrates that CAFs have a phenotypic plasticity that allows their adaptation to contrasting immune tissue microenvironments.

Carcinomas assemble a filamentous CXCL12-keratin-19 coating that suppresses T cell-mediated immune attack.

Cancer immunotherapy frequently fails because most carcinomas have few T cells, suggesting that cancers can suppress T cell infiltration. Here, we show that cancer cells of human pancreatic ductal adenocarcinoma (PDA), colorectal cancer, and breast cancer are coated with transglutaminase-2 (TGM2)-dependent covalent CXCL12-keratin-19 (KRT19) heterodimers that are organized as filamentous networks. Since a dimeric form of CXCL12 suppresses the motility of human T cells, we determined whether this polymeric CXCL12-KRT19 coating mediated T cell exclusion. Mouse tumors containing control PDA cells exhibited the CXCL12-KRT19 coating, excluded T cells, and did not respond to treatment with anti-PD-1 antibody. Tumors containing PDA cells not expressing either KRT19 or TGM2 lacked the CXCL12-KRT19 coating, were infiltrated with activated CD8+ T cells, and growth was suppressed with anti-PD-1 antibody treatment. Thus, carcinomas assemble a CXCL12-KRT19 coating to evade cancer immune attack.

Bacterial Peptidoglycan Traverses the Placenta to Induce Fetal Neuroproliferation and Aberrant Postnatal Behavior

Maternal infection during pregnancy is associated with adverse outcomes for the fetus, including postnatal cognitive disorders. However, the underlying mechanisms are obscure. We find that bacterial cell wall peptidoglycan (CW), a universal PAMP for TLR2, traverses the murine placenta into the developing fetal brain. In contrast to adults, CW-exposed fetal brains did not show any signs of inflammation or neuronal death. Instead, the neuronal transcription factor FoxG1 was induced, and neuroproliferation leading to a 50% greater density of neurons in the cortical plate was observed. Bacterial infection of pregnant dams, followed by antibiotic treatment, which releases CW, yielded the same result. Neuroproliferation required TLR2 and was recapitulated in vitro with fetal neuronal precursor cells and TLR2/6, but not TLR2/1, ligands. The fetal neuroproliferative response correlated with abnormal cognitive behavior in CW-exposed pups following birth. Thus, the bacterial CW-TLR2 signaling axis affects fetal neurodevelopment and may underlie postnatal cognitive disorders.