PO-507 Deregulated mechanisms downstream of PLCG1 promoting cutaneous T cell lymphomaprogression

Abstract

IntroductionCutaneous T Cell Lymphoma (CTCL) is a heterogeneous group of non-Hodgkin lymphomas with unfavourable prognosis at advanced stages. The malignant network that control the development of CTCL is now starting to be characterised. NGS-derived studies have identified multiple molecular alterations in specific genes and signalling pathways including PLCG1 and JAK-STAT. It has been shown that up to 50% of CTCL cases develop deregulated PLCG1-downstream signalling as shown by activating PLCG1 mutations and PRKCQ amplifications. Also, frequent mutations in JAK1 and JAK3 and in members of the STAT family have been identified, thus revealing a potentially important role of these signalling pathways in CTCL. We aimed at exploring the interactions between deregulated PLCG1 and JAK-STAT activity in CTCL using pre-clinical models and in samples from CTCL patients at different stages of the disease.Material and methodsHEK-BlueTM IL-6 and CTCL cell lines were used to study PLCG1 downstream signalling towards NFAT and STAT3 activation. Immunohistochemistry (IHC) of NFAT, pSTAT1, pSTAT3, pSTAT5, p50, p52, p65 and RelB wasperformed in 71 patient samples. The status of PRKCQ was analysed in 45 samples by fluorescent in situhybridization (FISH).Results and discussionsDeregulated PLCG1 activity elicited NFAT and STAT3 activation ex vivo. Downstream of PLCG1, pharmacological and genetic inhibition of PRKCQ negatively affected NFAT and STAT3 activation whereas a constitutively active PRKCQ mutant (A148E) was sufficient to trigger their transcriptional activity. Our data shows that downstream of PLCG1, PRKCQ can activate transcription mediated by both NFAT and STAT proteins. We also analysed nuclear accumulation of NFAT, STAT and NF-κB family proteins by IHC in patient samples. Interestingly positive P-STAT3 staining correlated with advanced diseases. Moreover, most CTCL-tumour stage cases studied were positive for PRKCQ amplification and/or 10q23 polysomy.ConclusionOur results support PLCG1 playing a mayor role at controlling CTCL progression towards advanced stages. Downstream of PLCG1 or due to genetic amplification, PRKCQ can mediate the activation of NFAT and STAT3. Thus, mutations in PLCG1, PRKCQ amplifications and/or nuclear NFAT and STAT3 accumulation can serve as diagnostic markers for CTCL. Moreover, these can also provide rational to develop specific therapies targeting PRKCQ alone or in combination with calcineurin or JAK-STAT inhibitors.