CREB3L1 Overexpression Can Reliably Discriminate Ph-MPNs from Reactive Cases
Authors: Morishita Soji , Yasuda Hajime , Yamawaki Saya , Kawaji Hideya , Itoh Masayoshi , Edahiro Yoko , Imai Misa , Kogo Yasushi , Tsuneda Satoshi , Ohsaka Akimichi , Hayashizaki Yoshihide , Araki Marito , Komatsu Norio 
Published: February 2021

Discrimination of Philadelphia-negative myeloproliferative neoplasms (Ph-MPNs) from reactive hypercytosis and myelofibrosis is imperative because treatment strategies differ greatly, and an exhaustive search for the underlying cause becomes mandatory in reactive cases. However, discrimination is not necessarily easy in the real-world setting, and a simple and universally utilizable method that can efficiently differentiate Ph-MPNs from reactive cases is awaited.
We extracted platelet rich plasma (PRP) derived RNAs from 9 essential thrombocythemia (ET) patients (3 patients with JAK2V617F, 3 patients with MPLW515L/K, and 3 patients with CALR exon 9 frameshift mutation) and 6 patients with reactive thrombocytosis (3 cases due to chronic inflammation, and 3 cases due to rebound thrombocytosis) and performed RNA-seq to identify a gene expressed specifically in ET. RNA-seq analysis followed by differential expression and principal component analysis revealed that CREB3L1 had the highest impact in discriminating ET from reactive cases. Subsequently, expression levels of CREB3L1 in PRP were quantified by reverse transcription quantitative PCR and compared within patients with various Ph-MPNs harboring either JAK2, MPL, or CALR mutations, chronic myeloid leukemia (CML), and reactive cases, and found that CREB3L1 expression levels were significantly higher in 66 ET compared to 33 reactive thrombocytosis (p &lt; 0.0001), 26 polycythemia vera (PV) compared to 23 reactive erythrocytosis (p &lt; 0.0001), 22 primary myelofibrosis and 15 post-PV/ET myelofibrosis (MF) compared to 3 reactive MF (p &lt; 0.001, and p &lt; 0.01, respectively), and the entire cohort of 129 Ph-MPN compared to 5 CML patients (p &lt; 0.001). A clear cut-off value discriminating Ph-MPNs and non-Ph-MPNs was determined, and sensitivity and specificity were both 1.0000. Furthermore, when we tested CREB3L1 expression levels of triple-negative cases with thrombocytosis, all patients with CREB3L1 overexpression were pathologically diagnosed as ET by bone marrow biopsy.
We demonstrate that CREB3L1 overexpression can single-handedly and reliably discriminate Ph-MPNs from reactive hypercytosis, reactive myelofibrosis, and CML. Early utilization of this method in the diagnostic process can guide patients to an efficient diagnosis and free many patients from unnecessary testing.

Disclosures
Komatsu: Otsuka Pharmaceutical Co., Ltd., PharmaEssentia Japan KK, AbbVie GK, Celgene KK, Novartis Pharma KK, Shire Japan KK, Japan Tobacco Inc: Consultancy; Takeda Pharmaceutical Co., Ltd, Novartis Pharma KK, Shire Japan KK: Speakers Bureau; AbbVie: Other: member of safety assessment committee in M13-834 clinical trial.; PPMX: Consultancy, Research Funding; Otsuka Pharmaceutical Co., Ltd., Shire Japan KK, Novartis Pharma KK, PharmaEssentia Japan KK, Fuso Pharmaceutical Industries, Ltd., Fujifilm Wako Pure Chemical Corporation, Chugai Pharmaceutical Co., Ltd., Kyowa Hakko Kirin Co., Ltd., Takeda Pharmaceutica: Research Funding; Meiji Seika Pharma Co., Ltd.: Patents & Royalties: PCT/JP2020/008434, Research Funding.