Hepatocellular carcinoma (HCC) is one of the most prevalent types of cancer and is the third leading cause of cancer-related mortality [1]. The immunosuppressive tumor microenvironment (TME) is a pivotal factor contributing significantly to the development, progression, metastasis and treatment resistance of HCC [2,3].

Myeloid-derived suppressor cells (MDSCs) constitute major components in the microenvironment of human solid tumors. Previous studies have established that MDSCs exert immunosuppressive properties, including inhibition of T-cell proliferation/activation, and promotion of tumor angiogenesis [4]. Therefore, inhibiting MDSCs represents an attractive possibility to reshape the tumor immune microenvironment in current HCC immunotherapy efforts [5]. Currently, MDSC defines two major subtypes, including monocyte (M-MDSC) and polymorphonuclear cell (PMN-MDSC) [6]. Of note, recent studies have highlighted that the MDSCs compartment is highly dynamic and heterogeneous, lacking of distinct markers hamper the progress in understanding of the biology and clinical importance of these cells [7]. More importantly, the mechanisms responsible for acquisition of pathological activity by MDSCs in cancer remained unclear.

Colony-stimulating factor 1 receptor (CSF1R) has been identified as a gatekeeper in controlling myeloid cell homeostasis and functions. Increased expression of CSF1R has been shown to be associated with aggressive tumors characterized by immunosuppressive microenvironments and poor prognosis [8]. Upregulation of CSF1R promotes the immune-suppressive activity of tumor-associated macrophages (TAMs), supports tumor growth, angiogenesis, invasion, and metastasis [9,10]. However, clinical trials show monotherapy by CSF1R blockade have limited efficacy in impeding tumor progression [11]. The divergent role of CSF1R in myeloid cells and the impact of the local microenvironment contribute to this challenge [12]. Thus, fully elucidating the regulation and role of CSF1R in MDSCs developmental trajectories occurring within tumors may further expand the landscape of HCC immunotherapy.

Triptolide (TPT) is a major bioactive ingredient of the Chinese herb Tripterygium wilfordii Hook f (TWHf), which has been used as anti-inflammatory and immunosuppressive drugs for centuries [13]. TPT or its derivatives possess potent anti-tumor activity in a variety of human cancers, including HCC [14]. Several TPT derivatives are currently in phase I/II clinical trial for cancer therapy [15,16]. TPT exerts therapeutic effect by regulating the function of immune cells, such as polarizing macrophage toward the M2 phenotype [[17], [18], [19]]. Nonetheless, the effects and signaling pathways of TPT in modulating MDSCs differentiation and function in HCC are not clear.

Using single-cell transcriptomic profiling of liver biopsies from HCC patients, we identified CSF1R+ MDSCs as a distinct cell lineage in HCC patients. Pseudo-timing analysis of CSF1R+ MDSCs, coupled with in vitro experiments employing ER inhibitors, revealed the essential function of the ER stress pathway in MDSCs lineage differentiation. Moreover, TPT can reduce the frequencies of MDSCs and CSF1R+ PMN-MDSCs by downregulating protein kinase RNA activated -like endoplasmic reticulum kinase (PERK)pathway. Our results support that CSF1R expression regulated by ER stress contributes to HCC disease progression by promoting the immunosuppressive effects of MDSCs, and identify CSF1R as a novel immunosuppressive target of TPT in HCC.