Single cell RNA sequencing (scRNAseq) of transplanted mT3 tumors

Background & Aims: Pancreatic ductal adenocarcinomas
(PDAC) are characterized by fibrosis and an abundance of cancer-associated
fibroblasts (CAFs). We investigated strategies to disrupt interactions
among CAFs, the immune system, and cancer cells, focusing on adhesion
molecule cadherin 11 (CDH11), which has been associated with other
fibrotic disorders and is expressed by activated fibroblasts.
Methods: We compared levels of CDH11mRNA in human
pancreatitis and pancreatic cancer tissues and cells, compared with normal
pancreas, and measured levels of CDH11 protein in human and mouse
pancreatic lesions and normal tissues. We crossed
p48-Cre;LSL-KrasG12D/+;LSL-Trp53R172H/+(KPC) mice with CDH11-knockout mice
and measured survival times of offspring. Pancreata were collected and
analyzed by histology, immunohistochemistry, and (single-cell) RNA
sequencing; RNA and proteins were identified by imaging mass cytometry.
Some mice were given injections of PD1 antibody or gemcitabine and
survival was monitored. Pancreatic cancer cells from KPC mice were
subcutaneously injected
into Cdh11+/+ and Cdh11–/– mice and
tumor growth was monitored. Pancreatic cancer cells (mT3) from KPC mice
(C57BL/6), were subcutaneously injected
into Cdh11+/+ (C57BL/6J) mice and mice were given
injections of antibody against CDH11, gemcitabine, or small molecule
inhibitor of CDH11 (SD133) and tumor growth was monitored.
Results: Levels of CDH11mRNA and protein were
significantly higher in CAFs than in pancreatic cancer epithelial cells,
human or mouse pancreatic cancer cell lines, or immune cells.
KPC/Cdh11+/– and KPC/Cdh11–/– mice survived
significantly longer than KPC/Cdh11+/+ mice. Markers of stromal
activation entirely surrounded pancreatic intraepithelial neoplasias in
KPC/Cdh11+/+ mice and incompletely in KPC/Cdh11+/– and
KPC/Cdh11–/– mice, whose lesions also contained fewer FOXP3+cells
in the tumor center. Compared with pancreatic tumors
inKPC/Cdh11+/+ mice, tumors of KPC/Cdh11+/– mice had
increased markers of antigen processing and presentation; more lymphocytes
and associated cytokines; decreased extracellular matrix components; and
reductions in markers and cytokines associated with immunosuppression.
Administration of the PD1 antibody did not prolong survival of KPC mice
with 0, 1, or 2 alleles of Cdh11. Gemcitabine extended survival
only of KPC/Cdh11+/– and KPC/Cdh11–/– mice or reduced
subcutaneous tumor growth in mT3 engrafted Cdh11+/+ mice
given in combination with the CDH11 antibody. A small molecule inhibitor
of CDH11 reduced growth of pre-established mT3 subcutaneous tumors only if
T and B cells were present in mice. Conclusions: Knockout or
inhibition of CDH11, which is expressed by CAFs in the pancreatic tumor
stroma, reduces growth of pancreatic tumors, increases their response to
gemcitabine, and significantly extends survival of mice. CDH11 promotes
immunosuppression and extracellular matrix deposition, and might be
developed as a therapeutic target for pancreatic cancer