Polymerase ε (POLE) ultra-mutated tumors induce robust tumor-specific CD4+ T cell responses in endometrial cancer patients.
OBJECTIVE
Around 7-10% of endometrial carcinomas are characterized by polymerase-ε-(POLE) exonuclease-domain-mutations, an ultra-mutated-phenotype and a favorable prognosis. It is currently unknown whether POLE ultra-mutated-tumors are more immunogenic when compared to the other groups of endometrial cancers.


METHODS
We used autologous-dendritic-cells (DC) pulsed with whole-tumor-extracts to assess the level of CD8+ and CD4+ T-cell-activation induced by POLE-ultramutated (+) and POLE wild-type (-) endometrial cancer cells in vitro. T-lymphocyte-proliferations were evaluated using CFSE and/or ([3H])thymidine-incorporation-assays while the ability to specifically kill autologous-tumor-cells by cytotoxic-T-lymphocyte (CTL) was tested in standard 4-h-(51)Cr-cytotoxicity-assays. In order to correlate cytotoxic activity and proliferation by CD4+ and CD8+ T-lymphocytes, respectively, with a particular lymphoid subset, two-color-flow-cytometric analysis of intracellular-cytokine-expression (IFN-γ vs IL-4) at the single cell level was also performed.


RESULTS
DC-pulsed with tumor extracts were able to induce CTL-responses against autologous-tumor-cells in both POLE (+) and POLE (-) cancer patients (P=0.305). These CD8+ T-cell-populations were cytotoxic against tumor-cells but they did not lyse PHA-stimulated-autologous-lymphocytes or autologous-EBV-transformed-lymphoblastoid-control-cell-lines. In contrast, only POLE (+) tumor-lysate-pulsed-DC were able to induce significant proliferation and high IFN-γ expression (i.e., Th1-cytokine-bias) in autologous in vitro DC-stimulated CD4+ T-cells as well as naïve CD4+ and CD8+ T-cells from patients-peripheral-blood (P<0.05).


CONCLUSIONS
POLE ultra-mutated-tumors are significantly more immunogenic when compared to POLE (-) tumors, in particular to the helper arm of the immune system. These data lend support to the hypothesis that the better prognosis of patients with POLE (+) tumors may at least in part be linked to their enhanced immunogenicity.
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Stefania
S
Bellone
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Floriana
F
Centritto
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Jonathan
J
Black
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Carlton
C
Schwab
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Diana
D
English
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Emiliano
E
Cocco
Division of Gynecologic Oncology, University Campus Bio-Medico of Roma, Roma, Italy.
Salvatore
S
Lopez
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Elena
E
Bonazzoli
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Federica
F
Predolini
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Francesca
F
Ferrari
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Dan-Arin
DA
Silasi
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Elena
E
Ratner
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Masoud
M
Azodi
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA.
Peter E
PE
Schwartz
Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, 333 Cedar Street, PO Box 208063, New Haven, CT 06520-8063, USA. Electronic address: alessandro.santin@yale.edu.
Alessandro D
AD
Santin
D000328:Adult; D000368:Aged; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D045744:Cell Line, Tumor; D004257:DNA Polymerase II / Q000235:genetics* / Q000276:immunology; D003713:Dendritic Cells / Q000276:immunology; D016889:Endometrial Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000276:immunology*; D005260:Female; D006801:Humans; D008213:Lymphocyte Activation; D008875:Middle Aged; D009154:Mutation; D000075223:Poly-ADP-Ribose Binding Proteins; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology
D000075223:Poly-ADP-Ribose Binding Proteins; D004257:DNA Polymerase II; C119525:POLE protein, human
Endometrial carcinoma; Polymerase ε; Uterine serous papillary cancer; Whole exome sequencing