Magnetic Field-Induced Strain of Metamagnetic Heusler Alloy Ni41Co9Mn31.5Ga18.5

Ni41Co9Mn31.5Ga18.5 is a re-entrant and metamagnetic Heusler alloy. In order to investigate the magnetic functionality of polycrystalline Ni41Co9Mn31.5Ga18.5, magnetic field-induced strain (MFIS) measurements were performed. A 0.12% MFIS was observed at 340 K and 10 T. Strict MFISs between 330 and 370 K were observed. These magneto-structural variances acted in concert with the metamagnetic property observed by the magnetization measurements and magneto-caloric property observed by the caloric measurements in applied magnetic fields. The MFISs were proportional to the fourth power of the magnetization, and this result is in agreement with Takahashi’s spin fluctuation theory of itinerant electron magnetism. The investigation of time response of the MFIS was performed by means of water-cooled electric magnet, zero magnetic field to 1.66 T in 8.0 s at 354 K. A 2.2×10−4 MFIS was observed, which was 80% of the MFIS in a 60-s mode. This indicates that a high-speed transition has occurred on applying magnetic fields

Predictors for the outcomes of patients with estrogen receptor-positive, HER2-negative advanced breast cancer treated using palbociclib plus endocrine therapy

　A cyclin-dependent kinase (CDK) 4/6 inhibitor, palbociclib (PAL), combined with endocrine therapy is frequently used for the treatment of patients with estrogen receptor (ER)-positive, human epidermal growth factor receptor (HER) 2-negative advanced breast cancer. However, as predictors for the outcomes remain unclear, we retrospectively investigated them.　A total of 36 patients with ER-positive, HER2-negative advanced breast cancer were treated using PAL plus endocrine therapy at our hospital. Treatment outcomes, objective response rates (ORR), progression-free survival (PFS) and post-treatment overall survival (OS) were analyzed. As possible predictive biomarkers, retinoblastoma protein (Rb), phosphorylated Rb (pRb) and different CDKs were immunohistochemically investigated using primary tumor tissues. Non-visceral metastasis, use of fulvestrant (FUL) and the 1st- or 2nd-line treatment were significant predictors for a better ORR (P = 0.0080, P = 0.0080 and P = 0.0080, respectively). No objective response (OR) was observed in patients with progesterone receptor (PR)-negative, CDK6-positive or cytosolic cyclin E1-positive tumors. Non-visceral metastasis and use of FUL were significant predictors for a better PFS (P = 0.0030 and P = 0.0443, respectively). The Cox proportional hazards model revealed that visceral metastasis (hazard ratio [HR], 4.9; P = 0.0019) and PR-negativity (HR, 3.2; P = 0.0411) were independent predictors for a poorer PFS. Nonvisceral metastasis, pRb-negativity and CDK6-negativity were significant predictors for a better OS (P = 0.0281, P = 0.0014 and P = 0.0396, respectively). The Cox proportional hazards model revealed that visceral metastasis (HR, 7.2; P = 0.0131) and pRb-positivity (HR, 18.5; P = 0.0060) were independent predictors for a poorer OS.　In conclusion, PR-negativity and pRb-positivity in primary tumors may be independent predictors for PFS and OS, respectively. CDK6-positivity and cytosolic cyclin E1-positivity may be predictors for a poorer OS and ORR, respectively. Further investigation is needed to confirm these factors

Renal Distribution of Vasohibin-1 in Patients with Chronic Kidney Disease

Experimental studies have demonstrated the involvement of angiogenesis-related factors in the progression of chronic kidney disease (CKD). There have so far been no reports investigating the distribution and clinical roles of Vasohibin-1 (VASH-1), a negative feedback regulator of angiogenesis, in CKD. We recruited 54 Japanese CKD patients and 6 patients who had normal renal tissues excised due to localized renal cell carcinoma. We evaluated the correlations between the renal expression level of VASH-1 and the clinical/histological parameters. VASH-1 was observed in renal endothelial/mesangial cells, crescentic lesions and interstitial inflammatory cells. Significant positive correlations were observed between 1) crescent formation and the number of VASH-1+ cells in the glomerulus (r＝0.48, p＝0.001) or cortex (r＝0.64, p＜0.0001), 2) interstitial cell infiltration and the number of VASH-1+ cells in the cortex (r＝0.34, p＝0.02), 3) the glomerular VEGFR-2+ area and the number of VASH-1+ cells in the glomerulus (r＝0.44, p＝0.01) or medulla (r＝0.63, p＝0.01). These results suggest that the renal levels of VASH-1 may be affected by local inflammation, crescentic lesions and VEGFR-2

Anti-cancer stem cell activity of the Src inhibitor dasatinib in thyroid cancer cells

Although the prognosis of differentiated thyroid cancer (DTC) is good, those of poorly-differentiated and undifferentiated thyroid cancers (PDTC and UDTC) are poor. Recent preclinical studies have suggested that the Src inhibitor dasatinib is active in thyroid cancer cell lines. We conducted the present study in an attempt to clarify the antitumor activity of dasatinib in PDTC and UDTC. The expression levels of c-Src, phosphorylated Srcs (p-SrcY416 and p-SrcY527), focal adhesion kinase (FAK), and phosphorylated FAK (p-FAKY861) were immunohistochemically investigated in a case-control series (15 cases of PDTC or UDTC vs. 29 control cases of DTC). The PDTC cell line KTC-1 and UDTC cell line KTC-2 were used to investigate the anticell growth and anti-cancer stem cell (CSC) activities of dasatinib. The combined effects of dasatinib and the taxane paclitaxel on anti-cell growth and anti-CSC activities were also tested. c-Src and p-FAKY861 expression levels were significantly higher, while those of p-SrcY416 were slightly higher in PDTC and UDTC than in DTC. Dasatinib inhibited cell growth in association with G1-S cell cycle retardation and increased apoptosis in both cell lines. Dasatinib significantly decreased the proportion of CSCs and more than additively enhanced the anti-cell growth activity of paclitaxel. The results of this study suggest that the Src signaling pathway is activated more in PDTC and UDTC than in DTC. The Src inhibitor dasatinib exhibited anti-cell growth and anti-CSC activities. Furthermore, it more than additively enhanced the anti-cell growth activity of paclitaxel

Immunohistochemistry of Vasohibin-2 in Human Kidney Disease : Implications in Impaired Glucose Tolerance and Reduced Renal Function

Several angiogenesis-related factors are known to play important roles in the pathogenesis of kidney disease. Vasohibin-2 (VASH-2) was recently reported as a novel proangiogenic factor. Although VASH-2 was demonstrated to accelerate tumor angiogenesis, its roles in non-tumor processes including renal disease have not been well elucidated yet. Here, we performed a retrospective study including an immunohistochemical analysis of human kidney biopsy specimens from 82 Japanese patients with a variety of kidney diseases, and we evaluated the correlations between the immunoreactivity of VASH-2 and the patients’ clinicopathological parameters. VASH-2 immunoreactivity was detected in varying degrees in renal tubules as well as in peritubular capillaries and vasa recta. The cortical and medullary tubule VASH-2+ scores were correlated with the presence of hypertension, and the medullary tubule VASH-2+ score was significantly correlated with the blood glucose (p=0.029, r=0.35) and hemoglobin A1c levels (p=0.0066, r=0.39). Moreover, decreased VASH-2+ scores in the vasa recta were associated with reduced renal function (p=0.0003). These results suggest that VASH-2 could play an important role in the pathogenesis of renal diseases, and that VASH-2 is closely associated with hypertension and impaired glucose tolerance

エストロゲン受容体陽性乳癌細胞における抗エストロゲン薬とmTOR阻害薬エベロリムスの細胞増殖及び癌幹細胞制御に対する効果

乳癌を含め多くの固形腫瘍において,癌幹細胞(CSC)が治療抵抗性や再発の原因となることが示唆されている.一方, エストロゲン感受性乳癌におけるCSCの制御機構に関する研究は少ない.そこで,エストロゲン受容体(ER)陽性乳癌細胞におけるエストロゲンや抗エストロゲン薬(抗E薬)の細胞増殖やCSC制御に与える影響について検討した.さらに,内分泌療法抵抗性乳癌に有効性が期待されているmTOR阻害薬エベロリムス(EVE)と抗E薬との併用効果も検討した.ER陽性乳癌の実験モデルとして,エストロゲン高感受性(HS)のMCF-7,T-47D乳癌細胞株,エストロゲン低感受性(LS)のKPL-1,KPL-3C乳癌細胞株を用いた.薬剤は,17β-estradiol(E2),4-hydroxytamoxifen(4-OHT),fulvestrant(FUL),EVEを用い,細胞増殖,細胞周期,アポトーシス,CSC比率に与える影響を検討した.CSCの同定には,CD44/CD24/EpCAM抗体を用いたフローサイトメトリー法及びmammosphere assayを用いた.ER-α,PgRおよびER関連転写因子(GATA3等)の発現は免疫細胞化学的に検討した.結果として(1)LS細胞株では,PgRの発現が認められなかった.それ以外のER関連因子は,HS細胞株,LS細胞株ともに高発現が認められた.(2)HS細胞株はLS細胞株に比べ,E2による細胞増殖の促進効果,CSC比率の増加効果が,ともにより顕著であった.(3)HS細胞株はLS細胞株に比べ,抗E薬による細胞増殖の抑制効果,CSC比率の低下効果がより顕著であった.(4)EVEと抗E薬の併用は,LS細胞株において相加的な細胞増殖抑制効果を示した.両薬の併用により,一部の細胞株ではCSC比率の減少効果が増強された.以上の結果は,LS乳癌において,抗E薬の増殖抑制効果ばかりでなく,CSC比率の低下効果も減弱していることを示唆している.抗E薬抵抗性獲得のメカニズムの一つとして,CSC制御機構の異常が関わっている可能性がある.また,一部の乳癌細胞株において,抗E薬とEVE併用の結果から,内分泌抵抗性乳癌におけるEVEの有用性が示唆された.Recent studies have revealed the existence of cancer stem cells (CSCs) in breast cancer. CSCs may play critical roles in resistance to anticancer agents and radiation, and in the development of metastases. However, the roles of CSCs in estrogen-responsive breast cancer remain to be elucidated. In this study, we investigated the effects of estrogen and antiestrogens (AEs) as well as an inhibitor of mammalian target of rapamycin (mTOR), everolimus (EVE) on the cell growth and regulation of the CSC population in estrogen-responsive breast cancer cell lines (BCCLs). Two high estrogen-sensitivity (HS) BCCLs, MCF-7 and T-47D, and two low estrogen-sensitivity (LS) BCCLs, KPL-1 and KPL-3C, were used. 4-hydroxytamoxifen (4-OHT) and a steroidal AE fulvestrant (FUL) and EVE were studied. Expression levels of ER-related proteins, ER-α, progesterone receptor (PgR), GATA3, FOXA1 and XBP1 were immunocytochemically investigated. The effects of the agents on the cell growth, cell cycle progression, apoptosis and regulation of the CSC population, defined as EpCAM-positive, CD44-high, CD24-low or negative cells using flow cytometery or mammosphere assay, were examined. Experimental results are as follows: (1) The LS BCCLs were negative for PgR but positive for the other ER-related proteins examined. The HS BCCLs were positive for all the ER-related proteins. The LS BCCLs were tumorigenic in female nude mice without estrogen supplementation, but the HS BCCLs were not. (2) The increase in cell growth induced by 17β-estradiol (E2) was more pronounced in the HS BCCLs. The decrease in cell growth induced by 4-OHT or FUL was more pronounced in the HS BCCLs. The AEs inhibited the G1-S cell cycle progression and induced a slight apoptosis. (3) E2 significantly increased the proportion of CSCs in all the cell lines tested. The increase in the CSC proportion by E2 was more pronounced in the HS BCCLs. The decrease in the CSC proportion by both 4-OHT and FUL was more pronounced in the HS BCCLs. (4) The decrease in the cell growth by EVE was more pronounced in LS BCCLs. Combined treatment of 4-OHT or FUL with EVE showed an additive effect in the LS BCCLs but not in the HS BCCLs. (5) Although EVE alone slightly increased the proportion of CSCs, the combined treatment of 4-OHT or FUL with EVE did not increased it. In conclusions, the HS BCCLs were more sensitive to estrogen and AEs than the LS BCCLs with regard to the cell growth response and CSC regulation. Thus, the LS BCCLs were more resistant to AE. It should be noted that EVE was more active in the LS BCCLs. These findings suggest that the combined use of EVE with AEs may be more useful in patients with breast cancer resistant to AEs

Preferential antitumor effect of the Src inhibitor dasatinib associated with a decreased proportion of aldehyde dehydrogenase 1-positive cells in breast cancer cells of the basal B subtype

<p>Abstract</p> <p>Background</p> <p>Recent studies have suggested that the Src inhibitor dasatinib preferentially inhibits the growth of breast cancer cells of the basal-like subtype. To clarify this finding and further investigate combined antitumor effects of dasatinib with cytotoxic agents, a panel of breast cancer cell lines of various subtypes was treated with dasatinib and/or chemotherapeutic agents.</p> <p>Methods</p> <p>Seven human breast cancer cell lines were treated with dasatinib and/or seven chemotherapeutic agents. Effects of the treatments on c-Src activation, cell growth, cell cycle, apoptosis and the proportion of aldehyde dehydrogenase (ALDH) 1-positive cells were examined.</p> <p>Results</p> <p>The 50%-growth inhibitory concentrations (IC₅₀s) of dasatinib were much lower in two basal B cell lines than those in the other cell lines. The IC₅₀s of chemotherapeutic agents were not substantially different among the cell lines. Dasatinib enhanced antitumor activity of etoposide in the basal B cell lines. Dasatinib induced a G1-S blockade with a slight apoptosis, and a combined treatment of dasatinib with etoposide also induced a G1-S blockade in the basal B cell lines. Dasatinib decreased the expression levels of phosphorylated Src in all cell lines. Interestingly, dasatinib significantly decreased the proportion of ALDH1-positive cells in the basal B cell lines but not in the other cell lines.</p> <p>Conclusions</p> <p>The present study indicates that dasatinib preferentially inhibits the growth of breast cancer cells of the basal B subtype associated with a significant loss of putative cancer stem cell population. A combined use of dasatinib with etoposide additively inhibits their growth. Further studies targeting breast cancers of the basal B subtype using dasatinib with cytotoxic agents are warranted.</p

甲状腺低分化・未分化癌細胞に対するヘッジホッグ阻害薬GANT61の抗腫瘍効果と癌幹細胞制御作用

　甲状腺分化癌は予後良好であるが低分化・未分化癌は予後不良であり，新規治療薬の開発が急務である．多くの悪性腫瘍でヘッジホッグ（Hh）経路の異常な活性化が起こっており，Hh 経路を標的とした治療戦略が有望視されている．Hh 経路の活性化は，腫瘍の生存・増殖・血管新生の促進ばかりでなく，癌幹細胞の制御との関与が示されている．我々は，甲状腺癌細胞を用いてHh経路を阻害するGANT61の抗腫瘍効果並びに癌幹細胞に与える影響を検討した．また，進行甲状腺癌の治療薬として用いられているタキサン系抗癌化学療法薬パクリタキセルとの併用効果も検討した．当教室で樹立された甲状腺低分化癌細胞株KTC-1及び甲状腺未分化細胞株KTC-2，KTC-3を用いてGANT61の細胞増殖，細胞周期，アポトーシス，癌幹細胞比率に与える影響を検討した．また，Hh 経路のeffector であるglioma-associated oncogene （Gli） 1，その下流にある癌幹細胞制御因子（aldehyde dehydrogenase [ALDH], Snail, Slug）や抗アポトーシス分子（survivin,Bcl-2）発現に与えるGANT61の効果を調べた．GANT61は，すべての甲状腺癌細胞株で細胞増殖を用量依存性に抑制した（50% 阻止濃度の平均値: KTC-1細胞は17.2 μM; KTC-2細胞は13.6 μM;KTC-3細胞は13.3 μM）．GANT61は，KTC-1及びKTC-2細胞のsub-G1分画を増加したが，G1-Sブロックは起こさなかった．GANT61は，全ての細胞株において用量依存性にアポトーシス分画を増加し，survivin やBcl-2の発現を低下させた．GANT61は，すべての細胞株でGli1, ALDH, Slugの発現を低下し，癌幹細胞比率を低下させた．以上の結果は，GANT61が甲状腺低分化・未分化癌細胞のsurvivin やBcl-2発現低下を介してアポトーシス誘導し，細胞増殖を抑制し，さらに，Hhシグナル標的因子Gli1, ALDH, Slug の発現低下により癌幹細胞の自己再生能を抑制することを示唆している．さらにGANT61は，すべての甲状腺癌細胞株においてパクリタキセルの細胞増殖抑制効果を増強した．これらの基礎研究の結果は，GANT61が甲状腺低分化・未分化癌の新規治療薬として有望なことを示唆している．　Patients with differentiated thyroid cancer have a good prognosis, but those with poorly-differentiated or undifferentiated thyroid cancer (PDTC or UDTC) do not. Thus, new therapeutics are urgently needed for PDTC and UDTC. As abnormal activation of the hedgehog (Hh) signaling pathway is observed in several malignancies, it is a promising therapeutic target. Activation of the Hh pathway is suggested to promote not only tumor survival, growth and angiogenesis, but also the growth of cancer stem cells (CSC). Therefore, we investigated the anti-cell growth and anti-CSC effects of the Hh inhibitor GANT61 in thyroid cancer cells. In addition, the combined anti-cell growth activity of GANT61 with an anti-thyroid cancer agent, paclitaxel, was evaluated. The effects of GANT61 on cell growth, cell cycle progression, apoptosis and CSC proportion using the Aldefluor and Thyrosphere assays were measured in the KTC-1 PDTC cell line, and KTC-2 and KTC-3 UDTC cell lines. All cell lines were established at our institute. We also examined the influence of GANT61 on the expression levels of the Hh effector glioma-associated oncogene (Gli) 1, its down-stream CSC-related molecules, aldehyde dehydrogenase (ALDH), Snail and Slug, and anti-apoptotic molecules, Bcl-2 and survivin. GANT61 dose-dependently inhibited the growth of all cell lines (mean 50% inhibitory concentrations: 17.2 μM for KTC-1 cells, 13.6 μM for KTC-2 cells and 13.3 μM for KTC-3 cells) in association with increased apoptosis, and decreased expression of survivin and Bcl-2. Furthermore, the proportion of surviving CSC cells decreased with decreased expression of Gli1, ALDH and Slug. These results demonstrate that GANT61 induced apoptosis via decreased expression levels of survivin and Bcl-2, and inhibited cell growth in thyroid cancer cells. Moreover, GANT61 reduced the expression levels of Hh target genes, such as Gli1, ALDH and Slug, and inhibited CSC self-renewal. GANT61 also enhanced the anti-cell growth effects of paclitaxel in all three thyroid cancer cell lines. Therefore, GANT61 may be a promising antitumor therapy for patients with PDTC or UTC