Search

In additive manufacturing (AM), the powder properties and laser powder bed fusion (LPBF) process parameters influence the quality of materials and building parts. However, the relationship between the size of the powder, LPBF process parameters, and mechanical properties is not well-established. In addition, Hastelloy X (HX) is attracting attention for its excellent high-temperature properties, but it is difficult to process, such as by cutting and milling, because of its high hardness and high ductility. This can be overcome by applying the AM process. We compared the LPBF window process maps for two HX powders of different sizes. Despite their small difference of 19.7% in particle size, it was confirmed that the difference in laser power was more than 40 W, the difference in scan speed was more than 100 mm/s, and the difference in energy density was more than 20% under the optimal process conditions. The as-built specimen had a larger molten-pool size as the energy density was higher, which resulted in the differences in mechanical properties at room temperature and high temperature (816 °C). We considered the control of the size of the powder to obtain the properties required for each temperature condition. The microstructures and mechanical properties of the as-built LPBF specimens were also investigated and compared with those of cast HX. Because of the rapid melting and solidification processes in LPBF, the as-built HX exhibited nano-sized dendrite structures and large internal strain energy. This resulted in the as-built LPBF exhibiting a higher room-temperature tensile strength than the cast material. Under high-temperature conditions, the grain boundary of the as-built LPBF acts as a sliding path, and the as-built LPBF HX showed significantly better high-temperature tensile strength characteristics than the cast HX.

Cbl‑associated protein (CAP) is encoded by the sorbin and SH3 domain‑containing 1 (SORBS1) gene. CAP has been reported to be associated with the actin cytoskeleton, receptor tyrosine kinase signaling and cell adhesion through interactions with various proteins. It may be hypothesized that SORBS1 has numerous unknown functions, which may include providing a favorable condition for metastasis. Although CAP has been demonstrated to possess a number of functions, the role of this protein has only been reported in metabolic signaling pathways and its function in cancer remains to be elucidated. In the present study, SORBS1 expression was detected in colorectal cancer cell lines divided into the primary group and the metastatic group by reverse transcription‑quantitative PCR and western blot analysis. In addition, SORBS1 expression was manipulated by vector transfection and lentivirus transduction. The metastatic role of SORBS1, as determined by assessing its effects on cell proliferation and migration, was determined by colony formation assay, cell cycle analysis and Boyden chamber assay. To elucidate the SORBS1‑binding protein, immunoprecipitation was performed. Co‑localization of SORBS1 and AHNAK nucleoprotein (AHNAK) was identified by confocal microscopy. Notably, the protein expression levels of CAP were higher in SNU‑769A and SW480 cells than in SNU‑769B and SW620 cells. In addition, the number of colonies in the SORBS1‑overexpressing group was significantly increased compared with that of the control group, as determined using the colony formation assay; the SORBS1 overexpression group formed >8‑fold more colonies than the control group. The proliferative ability of the SORBS1 overexpression group was also significantly increased compared with the control group over the entire incubation period. Cell migration assays revealed that the number of migrated SORBS1‑knockdown cells was reduced compared with the control in both HCT‑116 and SNU‑C4 cell lines; migration area was decreased to 31 and 26% in HCT‑116 and SNU‑C4 cell lines, respectively. Consequently, it was confirmed that SORBS1 could form a complex with AHNAK, which functions as a tumor suppressor through inhibition of phosphorylated‑ERK and Rho‑associated coiled‑coil containing protein kinase 1. In conclusion, SORBS1 may serve a crucial role in cancer growth and migration via inhibition of AHNAK expression.

Purpose This study was designed to identify novel fusion transcripts (FTs) and their functional significance in colorectal cancer (CRC) lines. Materials and methods We performed paired-end RNA sequencing of 28 CRC cell lines. FT candidates were identified using TopHat-fusion, ChimeraScan, and FusionMap tools and further experimental validation was conducted through reverse transcription-polymerase chain reaction and Sanger sequencing. FT was depleted in human CRC line and the effects on cell proliferation, cell migration, and cell invasion were analyzed. Results One thousand three hundred eighty FT candidates were detected through bioinformatics filtering. We selected six candidate FTs, including four inter-chromosomal and two intrachromosomal FTs and each FT was found in at least one of the 28 cell lines. Moreover, when we tested 19 pairs of CRC tumor and adjacent normal tissue samples, NFATC3-PLA2G15 FT was found in two. Knockdown of NFATC3-PLA2G15 using siRNA reduced mRNA expression of epithelial-mesenchymal transition (EMT) markers such as vimentin, twist, and fibronectin and increased mesenchymal-epithelial transition markers of E-cadherin, claudin-1, and FOXC2 in colo-320 cell line harboring NFATC3-PLA2G15 FT. The NFATC3-PLA2G15 knockdown also inhibited invasion, colony formation capacity, and cell proliferation. Conclusion These results suggest that that NFATC3-PLA2G15 FTs may contribute to tumor progression by enhancing invasion by EMT and proliferation.

Although MEK blockade has been highlighted as a promising antitumor drug, it has poor clinical efficacy in KRAS mutant colorectal cancer (CRC). Several feedback systems have been described in which inhibition of one intracellular pathway leads to activation of a parallel signaling pathway, thereby decreasing the effectiveness of single-MEK targeted therapies. Here, we investigated a bypass mechanism of resistance to MEK inhibition in KRAS CRC. We found that KRAS mutant CRC cells with refametinib, MEK inhibitor, induced MIF secretion and resulted in activation of STAT3 and MAPK. MIF knockdown by siRNA restored sensitivity to refametinib in KRAS mutant cells. In addition, combination with refametinib and 4-IPP, a MIF inhibitor, effectively reduced the activity of STAT3 and MAPK, more than single-agent treatment. As a result, combined therapy was found to exhibit a synergistic growth inhibitory effect against refametinib-resistant cells by inhibition of MIF activation. These results reveal that MIF-induced STAT3 and MAPK activation evoked an intrinsic resistance to refametinib. Our results provide the basis for a rational combination strategy against KRAS mutant colorectal cancers, predicated on the understanding of cross talk between the MEK and MIF pathways.

The tumor microenvironment is recognized as developing crosstalk between different cells by secretion of growth factors and cytokines, thus providing oncogenic signals enhancing tumor progression and drug resistance. Here, we hypothesized that tumor cells carrying KRAS mutation-induced cytokines may have a critical role in intercellular communication between microenvironment and tumor cells. We first investigated biologic evidence of secreted cytokines in KRAS mutant type (KRAS MT) cell lines. We explored the roles of cytokine that cell morphology, colony-formation ability, and wound-healing ability were enhanced when KRAS wild-type (KRAS WT) cell lines were exposed to conditioned media (CM) from KRAS MT cells, and we found macrophage migration inhibitory factor (MIF) was highly expressed in KRAS MT cells by analysis of proteomics. We also observed secreted MIF level between KRAS WT and MT cell lines, and it was highly secreted in KRAS MT cell lines. In addition, compared with patients whose KRAS mutation was not harbored, MIF expression was higher in patients who have KRAS mutation. The predictive marker of KRAS mutation in colorectal cancer patients is associated with resistance to antiepidermal growth factor receptor (EGFR) antibody cetuximab. Following the hypothesis, we investigated that secreted cytokines have an effect on the cetuximab resistance to surrounding cells including KRAS WT cells. Treatment of CM from KRAS MT cells led to cetuximab resistance in KRAS WT cells and MIF blockade prevented cetuximab resistance. Also, CM from knockout in KRAS MT cells by using CRISPR/Cas9 system resulted in sensitizing to cetuximab resistance compared with CM from KRAS MT cells. In conclusion, we demonstrated for the first time that MIF promotes the cetuximab resistance of KRAS WT colorectal cancer cells, and this effect is mediated by paracrine and autocrine signaling-induced activation of the intracellular AKT signaling pathways and regulated by NF-kb transcription factor through oncogenic KRAS mutation and MIF overexpression, respectively. These findings suggest that MIF may be a promising predictive biomarker for the cetuximab resistance of KRAS wild-type colorectal cancer patients. Citation Format: Jee Eun Jang, Hwang-Phill Kim, Sae Won Han, Tae You Kim. Mutant KRAS-induced macrophage migration inhibitory factor (MIF) secretion promotes resistance to cetuximab in colorectal cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B100.