Ovarian granulosa cell tumor characterization identifies FOXL2 as an immunotherapeutic target
Stefano Pierini, Janos L. Tanyi, Fiona Simpkins, Erin George, Mireia Uribe-Herranz, Ronny Drapkin, Robert Burger, Mark A. Morgan, Andrea Facciabene
Stefano Pierini, Janos L. Tanyi, Fiona Simpkins, Erin George, Mireia Uribe-Herranz, Ronny Drapkin, Robert Burger, Mark A. Morgan, Andrea Facciabene
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Research Article Immunology Oncology

Ovarian granulosa cell tumor characterization identifies FOXL2 as an immunotherapeutic target

Abstract

Granulosa cell tumors (GCT) are rare ovarian malignancies. Due to the lack of effective treatment in late relapse, there is a clear unmet need for novel therapies. Forkhead Box L2 (FOXL2) is a protein mainly expressed in granulosa cells (GC) and therefore is a rational therapeutic target. Since we identified tumor infiltrating lymphocytes (TILs) as the main immune population within GCT, TILs from 11 GCT patients were expanded, and their phenotypes were interrogated to determine that T cells acquired late antigen-experienced phenotypes and lower levels of PD1 expression. Importantly, TILs maintained their functionality after ex vivo expansion as they vigorously reacted against autologous tumors (100% of patients) and against FOXL2 peptides (57.1% of patients). To validate the relevance of FOXL2 as a target for immune therapy, we developed a plasmid DNA vaccine (FoxL2–tetanus toxin; FoxL2-TT) by fusing Foxl2 cDNA with the immune-enhancing domain of TT. Mice immunization with FoxL2-TT controlled growth of FOXL2-expressing ovarian (BR5) and breast (4T1) cancers in a T cell–mediated manner. Combination of anti–PD-L1 with FoxL2-TT vaccination further reduced tumor progression and improved mouse survival without affecting the female reproductive system and pregnancy. Together, our results suggest that FOXL2 immune targeting can produce substantial long-term clinical benefits. Our study can serve as a foundation for trials testing immunotherapeutic approaches in patients with ovarian GCT.

Authors

Stefano Pierini, Janos L. Tanyi, Fiona Simpkins, Erin George, Mireia Uribe-Herranz, Ronny Drapkin, Robert Burger, Mark A. Morgan, Andrea Facciabene

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Figure 6

Vaccination with FoxL2-TT reduced tumor progression.

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Vaccination with FoxL2-TT reduced tumor progression. (A) Healthy organs ...
(A) Healthy organs and tissues were collected from 3 mice, pooled together. RNA was extracted using Trizol, and foxl2 fold expression was calculated by real-time PCR. Mean ± SEM is shown. (B and C) T cells from FVB and BALB/c vaccinated mice were cocultured with BR5-FOXL2 and BR5 WT (B) and with IFN-γ–pretreated 4T1-FOXL2 and 4T1 WT (C). (D and F) FVB and BALB/c were injected s.c. with 1 × 106 BR5-FOXL2 (D) and 2.5 × 105 4T1-FOXL2 (F) and, 3 days later, injected 3 times with FoxL2-TT or empty pVAX (CTRL) vaccines followed by electroporation. Data are shown as mean ± SEM (n = 8–10 mice per group) from 1 of 3 experiments. Two-way ANOVA analyses were performed for tumor growth experiments. Percentage of tumor-infiltrating CD8+ and CD4+ T cells were calculated by flow cytometry at day 25. (E and G) Each dot represents a mouse, n = 6–3 mice per group (E) and n = 5 mice per group (G). Mean ± SEM is shown. Two-tailed t test analyses were performed. *P < 0.05, **P < 0.01.