CBP501 induces immunogenic tumor cell death and CD8 T cell infiltration into tumors in combination with platinum, and increases the efficacy of immune checkpoint inhibitors against tumors in mice

CBP501, a calmodulin-binding peptide, is an anti-cancer drug candidate and functions as an enhancer of platinum uptake into cancer cells. Here we show that CBP501 promotes immunogenic cell death (ICD) in combination with platinum agents. CBP501 enhanced a clinically relevant low dose of cisplatin (CDDP) in vitro as evidenced by upregulation of ICD markers, including cell surface calreticulin exposure and release of high-mobility group protein box-1. Synergistic induction of ICD by CDDP plus CBP501 as compared to CDDP alone was confirmed in the well-established vaccination assay. Furthermore, cotreatment of CDDP plus CBP501 significantly reduced the tumor growth and upregulated the percentage of tumor infiltrating CD8+ T cell in vivo. Importantly, the antitumor effect of CDDP plus CBP501 was significantly reduced by anti-CD8 antibody treatment. Based on this novel effect of CBP501, we analyzed the combination treatment with immune checkpoint inhibitors in vivo. Mice treated with CBP501 in combination with CDDP and anti-PD-1 or anti-PD-L1 showed an additive antitumor effect. These results support the conclusion that CBP501 enhances CDDP-induced ICD in vitro and in vivo. The findings also support the further clinical development of the CBP501 for enhancing the antitumor activity of immune checkpoint inhibitors in combination with CDDP

Molecular and cellular characteristics of hybrid vigour in a commercial hybrid of Chinese cabbage

Abstract Background Heterosis or hybrid vigour is a phenomenon in which hybrid progeny exhibit superior performance compared to their parental inbred lines. Most commercial Chinese cabbage cultivars are F1 hybrids and their level of hybrid vigour is of critical importance and is a key selection criterion in the breeding system. Results We have characterized the heterotic phenotype of one F1 hybrid cultivar of Chinese cabbage and its parental lines from early- to late-developmental stages of the plants. Hybrid cotyledons are larger than those of the parents at 4 days after sowing and biomass in the hybrid, determined by the fresh weight of leaves, is greater than that of the larger parent line by approximately 20 % at 14 days after sowing. The final yield of the hybrid harvested at 63 days after sowing is 25 % greater than the yield of the better parent. The larger leaves of the hybrid are a consequence of increased cell size and number of the photosynthetic palisade mesophyll cells and other leaf cells. The accumulation of plant hormones in the F1 was within the range of the parental levels at both 2 and 10 days after sowing. Two days after sowing, the expression levels of chloroplast-targeted genes in the cotyledon cells were upregulated in the F1 hybrid relative to their mid parent values. Shutdown of chlorophyll biosynthesis in the cotyledon by norflurazon prevented the increased leaf area in the F1 hybrid. Conclusions In the cotyledons of F1 hybrids, chloroplast-targeted genes were upregulated at 2 days after sowing. The increased activity levels of this group of genes suggested that their differential transcription levels could be important for establishing early heterosis but the increased transcription levels were transient. Inhibition of the photosynthetic process in the cotyledon reduced heterosis in later seedling stages. These observations suggest early developmental events in the germinating seedling of the hybrid may be important for later developmental vigour and yield advantage.This work was supported in part by a grant-in-aid for Scientific Research on Innovative Areas (24113509) (JSPS), by the Sasakawa Scientific Research Grant (24–517) from The Japan Science Society, by Grant for Promotion of Niigata University Research Projects (23C024) and by PREST (12101066) (JST) to R. Fujimoto

ウメ ハフチエソビョウ ビョウゲン ウイルス ノ ジュウエキ セッシュ ニ ヨル ウメ ヒンシュ ナンコウ ニ オケル ビョウチョウ サイゲン ノ カクニン

わが国有数のウメの産地,和歌山県紀南地方のウメ品種南高に発生している"ウメ葉縁えそ病"(茶がす症)がCucumber mosaic virus(CMV-Um)およびPrunus necrotic ringspot virus類似ウイルス(PNRLV)の重複感染によることを証明する目的で,り病樹から分離された,これら2ウイルスをウメ品種南高に混合接種して病徴の推移を調べた。その結果,萌芽の遅延,不完全花の高率発生,葉脈間の黄化による濃淡斑,全葉の黄化,葉縁部のえそ,早期の落葉など本病の病徴がおおよそ再現された。PNRLV単独接種では病徴は軽く,CMV-Um単独接種では健全対照樹と大差はなかった。以上から本病はこの両ウイルスの重複感染によるものと結論された。なお,花器の異常として雌蕊の発育不全が不稔の主因と思われることを示唆する結果が得られた。Mume leaf margin necrosis (mume leaf edge necrosis) disease, which has occurred in the southern district of Wakayama Prefecture, a major area for mume fruits production in Japan, was tested for symptoms expression by sap inoculation of causal viruses to original mume plantlets cv.Nankou grafted onto Nankou seedlings. Tobacco leaf tissues infected with Cucumber mosaic virus (CMV-Um) and Prunus necrotic ringspot-like virus (PNRLV) isolated from diseased mume plants in Wakayama were macerated, respectively, in phosphate buffer added with nicotine and sodium DIECA. Inoculations were made by slashing the surface of green shoots with a surgical knife dipped into inoculum. Eaqualy mixed saps of CMV-Um and PNRLV and both of single virus were inoculated to each 5 potted plantlets, respectively. Inoculated plants with mixed sap showed the symptoms of delayed sprouting, leaf interveinal chrolosis, leaf rolling, leaf margin necrosis, earlier yellowing of leaves and defoliation, and higher percentage occurrence of deformed flowers. These symptoms are similar to the naturally infected plants in the fields. The symptoms in the PNRLV plot indicated slighter symptoms than the mixed one, the CMV-Um plot showed almost symptomless and the buffer-inoculated control plot remained healthy. These results were evidence that the disease is caused by multiple infection with two viruses described above. Infertility of pistil in deformed flower suggests the major cause of barrenness in the diseased plant

Global Landscape of a Co-Expressed Gene Network in Barley and its Application to Gene Discovery in Triticeae Crops

Accumulated transcriptome data can be used to investigate regulatory networks of genes involved in various biological systems. Co-expression analysis data sets generated from comprehensively collected transcriptome data sets now represent efficient resources that are capable of facilitating the discovery of genes with closely correlated expression patterns. In order to construct a co-expression network for barley, we analyzed 45 publicly available experimental series, which are composed of 1,347 sets of GeneChip data for barley. On the basis of a gene-to-gene weighted correlation coefficient, we constructed a global barley co-expression network and classified it into clusters of subnetwork modules. The resulting clusters are candidates for functional regulatory modules in the barley transcriptome. To annotate each of the modules, we performed comparative annotation using genes in Arabidopsis and Brachypodium distachyon. On the basis of a comparative analysis between barley and two model species, we investigated functional properties from the representative distributions of the gene ontology (GO) terms. Modules putatively involved in drought stress response and cellulose biogenesis have been identified. These modules are discussed to demonstrate the effectiveness of the co-expression analysis. Furthermore, we applied the data set of co-expressed genes coupled with comparative analysis in attempts to discover potentially Triticeae-specific network modules. These results demonstrate that analysis of the co-expression network of the barley transcriptome together with comparative analysis should promote the process of gene discovery in barley. Furthermore, the insights obtained should be transferable to investigations of Triticeae plants. The associated data set generated in this analysis is publicly accessible at http://coexpression.psc.riken.jp/barley/

CBP501 suppresses macrophage induced cancer stem cell like features and metastases

CBP501 is an anti-cancer drug candidate which has been shown to increase cis-diamminedichloro-platinum (II) (CDDP) uptake into cancer cell through calmodulin (CaM) inhibition. However, the effects of CBP501 on the cells in the tumor microenvironment have not been addressed. Here, we investigated new aspects of the potential anti-tumor mechanism of action of CBP501 by examining its effects on the macrophages. Macrophages contribute to cancer-related inflammation and sequential production of cytokines such as IL-6 and TNF-α which cause various biological processes that promote tumor initiation, growth and metastasis (1). These processes include the epithelial to mesenchymal transition (EMT) and cancer stem cell (CSC) formation, which are well-known, key events for metastasis. The present work demonstrates that CBP501 suppresses lipopolysaccharide (LPS)-induced production of IL-6, IL-10 and TNF-α by macrophages. CBP501 also suppressed formation of the tumor spheroids by culturing with conditioned medium from the LPS-stimulated macrophage cell line RAW264.7. Moreover, CBP501 suppressed expression of ABCG2, a marker for CSCs, by inhibiting the interaction between cancer cells expressing VCAM-1 and macrophages expressing VLA-4. Consistently with these results, CBP501 in vivo suppressed metastases of a tumor cell line, 4T1, one which is insensitive to combination treatment of CBP501 and CDDP in vitro. Taken together, these results offer potential new, unanticipated advantages of CBP501 treatment in anti-tumor therapy through a mechanism that entails the suppression of interactions between macrophages and cancer cells with suppression of sequential CSC-like cell formation in the tumor microenvironment