Establishing an appropriate preclinical model is crucial for translational cancer research. The most common way that has been adopted by far is grafting cancer cell lines, derived from patients. Although this xenograft model is easy to generate, but has several limitations because this cancer model could not represent the unique features of each cancer patient sufficiently. Moreover, accumulating evidences demonstrate cancer is a highly heterogeneous disease so that a tumor is comprised of cancer cells with diverse characteristics. In attempt to avoid these discrepancies between xenograft model and patients' tumor, a patient-derived xenograft (PDX) model has been actively generated and applied. The PDX model can be developed by the implantation of cancerous tissue from a patient's tumor into an immune-deficient mouse directly, thereby it preserves both cell-cell interactions and tumor microenvironment. In addition, the PDX model has shown advantages as a preclinical model in drug screening, biomarker development and co-clinical trial. In this review, we will summarize the methodology and applications of PDX in detail, and cover critical issues for the development of this model for preclinical research.

The mouse Period2 ( mPer2 ) locus is an essential negative-feedback element of the mammalian circadian-clock mechanism. Recent work has shown that mPer2 circadian gene expression persists in both central and peripheral tissues. Here, we analyze the mouse mPer2 promoter and identify a circadian enhancer (E2) with a noncanonical 5′-CACGTT-3′ E-box located 20 bp upstream of the mPer2 transcription start site. The E2 enhancer accounts for most circadian transcriptional drive of the mPer2 locus by CLOCK:BMAL1, is a major site of DNaseI hypersensitivity in this region, and is constitutively bound by a transcriptional complex containing the CLOCK protein. Importantly, the E2 enhancer is sufficient to drive self-sustained circadian rhythms of luciferase activity in central and peripheral tissues from mPer2-E2 :: Luciferase transgenic mice with tissue-specific phase and period characteristics. Last, genetic analysis with mutations in Clock and Bmal1 shows that the E2 enhancer is a target of CLOCK and BMAL1 in vivo .

MicroRNA is an endogenous, small RNA controlling multiple target genes and playing roles in various biological processes including tumorigenesis. Here, we addressed the function of miR-155 using LC-MS/MS-based metabolic profiling of miR-155 deficient breast cancer cells. Our results revealed the loss of miR-155 hampers glucose uptake and glycolysis, via the down-regulation of glucose transporters and metabolic enzymes including HK2, PKM2, and LDHA. We showed this is due to the down-regulation of cMYC, controlled through phosphoinositide-3-kinase regulatory subunit alpha (PIK3R1)-PDK1/AKT-FOXO3a pathway. UTR analysis of the PIK3R1 and FOXO3a indicated miR-155 directly represses these genes. A stable expression of miR-155 in patient-derived cells (PDCs) showed activated glucose metabolism whereas a stable inhibition of miR-155 reduced in vivo tumor growth with retarded glucose metabolism. Furthermore, analysis of 50 triple-negative breast cancer (TNBC) specimens and specific uptake value (SUV) of PET images revealed a positive correlation between miR-155 level and glucose usage in human breast tumors via PIK3R1-PDK/AKT-FOXO3a-cMYC axis. Collectively, these data demonstrate the miR-155 is a key regulator of glucose metabolism in breast cancer.

Interactions between cancer cells and cancer-associated fibroblasts (CAFs) within the tumor microenvironment (TME) play an important role in promoting the profibrotic microenvironment and epithelial-mesenchymal transition (EMT), resulting in tumor progression and drug resistance in hepatocellular carcinoma (HCC). In the present study, we developed a mixed-cell spheroid model using Huh-7 HCC cells and LX-2 stellate cells to simulate the in vivo tumor environment with respect to tumor-CAF interactions. Spheroids were cultured from cancer cells alone (monospheroids) or as a mixture (mixed-cell spheroids) in ultra-low-attachment plates. Compact, well-mixed, and stroma-rich mixed-cell spheroids were successfully established with heterotypic cell-cell contacts shown by the presence of gap junctions and desmosomes. Mixed-cell spheroids showed enhanced expression of collagen type-I (Col-I) and pro-fibrotic factors such as, transforming growth factor beta1 (TGF-β1), and connective tissue growth factor (CTGF) compared to the levels expressed in mono-spheroids. The EMT phenotype was evident in mixed-cell spheroids as shown by the altered expression of E-cadherin and vimentin. Differential drug sensitivity was observed in mixed-cell spheroids, and only sorafenib and oxaliplatin showed dose-dependent antiproliferative effects. Simultaneous treatment with TGF-β inhibitors further improved sorafenib efficacy in the mixed-cell spheroids, indicating the involvement of TGF-β in the mechanism of sorafenib resistance. In 3D matrix invasion assay, mixed-cell spheroids exhibited fibroblast-led collective cell movement. Overall, our results provide evidence that mixed-cell spheroids formed with Huh-7 and LX-2 cells well represent HCC tumors and their TME in vivo and hence are useful in studying tumor-stroma interactions as mechanisms associated with drug resistance and increased cell motility.