Endocrine Abstracts

Objectives: BRAF^V600E- anaplastic thyroid cancers (ATCs) show remarkable responses to dabrafenib and trametinib (dab/tram) an effect that may be in part immune-mediated. Murine BRAF^V600E-ATCs regress upon BRAF inhibition. We find that recurrences are frequent and associated with loss of Mhc class II (MhcII) expression. Our goal was to investigate the mechanisms of loss of antigen presentation by tumor cells and whether this contributes to disease recurrence.

Methods: We developed primary ATC cell lines from mouse dox-inducible Braf-ATCs (TPO-Cre/LSL-rtTA_GFP/tetO-mycBRAF^V600E/p53^fl/fl (BRAF/p53) as well as ATC lines derived from recurrent tumors arising in mice after dox-withdrawal. A syngeneic orthotopic model with Braf^V600E/Tp53^-/- (TBP3743) ATC cells was developed. RNAseq of BRAF/p53 ATC cells from mice treated with or without dox and from dab/tram or vehicle treated TBP3743 ATCs was performed. Primary and recurrent cell lines were treated with vehicle, interferon γ (IFNγ), tram or IFNγ+tram and analyzed by FACS, RT-PCR and western blotting.

Results: Dox withdrawal or dab/tram treatment resulted in ATC infiltration by CD4⁺ T-helper cells and increased MhcII expression in tumor cells. RNAseq of ATC cells showed activation of IFNγ transcriptional output and of the antigen presentation pathway (MhcII > MhcI). IFNγ induced MhcII expression in primary cell lines only upon MEK inhibition with trametinib but this was ineffective in recurrent cell lines. Loss of induction of MhcII in recurrences was not due to incomplete ERK pathway inhibition or to interference with upstream IFNγ signaling. Recurrent cell lines had markedly attenuated basal and IFNγ induced expression of Ciita, the master transcriptional regulator of genes in the MhcII signaling pathway. We developed homozygous CRISPR KO of Ciita in TBP3743 cells to study the impact of Ciita loss in vivo. Ciita KO cells lost expression of MhcII in vitro and in vivo, which did not impact tumor growth but rendered the Ciita^-/- TBP3743 cells completely refractory to dab/tram therapy.

Conclusions: (i) IFNγ induction of MhcII expression requires MEK inhibition in primary BRAF^V600E- ATC cells but the combination fails to induce MchII in recurrences. (ii) Absence of MhcII expression is associated with attenuated Ciita expression. (iii) In Ciita Crispr KO TBP3743 ATC cells MhcII expression is absent and Ciita^-/- ATCs are resistant to dab/tram in vivo.