F-FDG PET/CT has been used as an indicator of chemotherapy effects, but cancer cells can remain even when no FDG uptake is detected, indicating the importance of exploring other metabolomic pathways. ...Therefore, we explored the amino acid metabolism, including L-type amino acid transporter-1 (LAT1), in breast cancer tissues and clarified the role of LAT1 in therapeutic resistance and clinical outcomes of patients. We evaluated LAT1 expression before and after neoadjuvant chemotherapy and examined the correlation of glucose uptake using FDG-PET with the pathological response of patients. It revealed that LAT1 levels correlated with proliferation after chemotherapy, and amino acid and glucose metabolism were closely correlated. In addition, LAT1 was considered to be involved in treatment resistance and sensitivity only in luminal type breast cancer. Results of in vitro analyses revealed that LAT1 promoted amino acid uptake, which contributed to energy production by supplying amino acids to the TCA cycle. However, in MCF-7 cells treated with chemotherapeutic agents, oncometabolites and branched-chain amino acids also played a pivotal role in energy production and drug resistance, despite decreased glucose metabolism. In conclusion, LAT1 was involved in drug resistance and could be a novel therapeutic target against chemotherapy resistance in luminal type breast cancer.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
•A simple, highly sensitive, and specific LC/ESI-MS/MS method was established.•CDK4/6 inhibitors, aromatase inhibitors, and an estrogen receptor antagonist were quantitated ...simultaneously.•Palbociclib, abemaciclib, letrozole, and fulvestrant in plasma of breast cancer patients were analyzed.
Cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors (palbociclib, abemaciclib, and ribociclib) are used to treat human epithelial growth factor receptor (HER)-2 negative and hormone receptor (HR) positive advanced breast cancer in combination with aromatase inhibitors (letrozole, anastrozole) or an estrogen receptor antagonist (fulvestrant). Administration of these drugs frequently causes severe side effects, such as neutropenia and diarrhea. Therefore, therapeutic drug monitoring (TDM) of CDK4/6 inhibitors, aromatase inhibitors, and the estrogen receptor antagonist is considered important for ensuring the efficacy and safety of these drugs. In this study, we describe a simple, highly sensitive, and specific liquid chromatography/electrospray ionization tandem mass spectrometry method for simultaneous quantitation of the concentrations of palbociclib, abemaciclib, ribociclib, letrozole, anastrozole, and fulvestrant. In addition, we analyzed plasma samples from patients with HER2-negative and HR-positive advanced breast cancer treated with these compounds using this novel method. In our method, the intra-assay relative error (RE) values ranged from −12.8% to 12.9%, the inter-assay RE values ranged from −4.8% to 6.2%, and the coefficient of variation (CV) values for intra- and inter-assay were ≤8.6% and ≤13.3%, respectively. The analytes showed good stability with RE values ranging from −13.5% to 13.6% and CV values <10.4%. Moreover, all the samples from patients were successfully quantified, and were within the range of measurement. This method can be used for TDM of routine anticancer drugs in clinical practice and for pharmacokinetics/pharmacodynamics research in future studies.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Homologous recombination (HR) repairs DNA damage including DNA double-stranded breaks and alterations in HR-related genes results in HR deficiency. Germline alteration of HR-related genes, such as ...BRCA1 and BRCA2, causes hereditary breast and ovarian cancer (HBOC). Cancer cells with HR deficiency are sensitive to poly (ADP-ribose) polymerase (PARP) inhibitors and DNA-damaging agents. Thus, accurately evaluating HR activity is useful for diagnosing HBOC and predicting the therapeutic effects of anti-cancer agents. Previously, we developed an assay for site-specific HR activity (ASHRA) that can quantitatively evaluate HR activity and detect moderate HR deficiency. HR activity in cells measured by ASHRA correlates with sensitivity to the PARP inhibitor, olaparib. In this study, we applied ASHRA to lymphoblastoid cells and xenograft tumor tissues, which simulate peripheral blood lymphocytes and tumor tissues, respectively, as clinically available samples. We showed that ASHRA could be used to detect HR deficiency in lymphoblastoid cells derived from a BRCA1 pathogenic variant carrier. Furthermore, ASHRA could quantitatively measure the HR activity in xenograft tumor tissues with HR activity that was gradually suppressed by inducible BRCA1 knockdown. The HR activity of xenograft tumor tissues quantitatively correlated with the effect of olaparib. Our data suggest that ASHRA could be a useful assay for diagnosing HBOC and predicting the efficacy of PARP inhibitors.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Aim: The aim of this study was to examine the effects of evolocumab on favorable limb events in patients with chronic limb-threatening ischemia (CLTI). Methods: A single-center, prospective ...observational study was performed on 30 patients with CLTI. The subjects were divided into 2 groups based on evolocumab administration: evolocumab-treated (E) group ( n=14) and evolocumab non-treated (non-E) group (n=16). The primary outcome was 12-month freedom from major amputation. The secondary outcomes were 12-month amputation-free survival (AFS), overall survival (OS), and wound-free limb salvage. The mean follow-up period was 18±11 months. Results: No significant difference was detected between the two groups for the 12-month freedom from major amputation (log-rank p=0.15), while the 12-month AFS rate was significantly higher in the E group than that in the non-E group (log-rank p=0.02). The 12-month OS rate in the E group was shown a tendency for improvement, as compared with that in the non-E group (log-rank p=0.056). Evolocumab administration was not associated with a significant change in freedom from major amputation (HR, 0.23, 95% CI, 0.03-2.07, p=0.19). However, evolocumab administration was related to a tendency for improvement of AFS and OS (HR, 0.13, 95% CI, 0.02-1.06, p=0.056; HR, 0.16, 95% CI, 0.02-1.37, p=0.09, respectively). Moreover, The E group had a higher proportion of wound-free limb salvage at 12 months (92% vs. 42%, p=0.03). Conclusion: Evolocumab administration was associated with a better AFS outcome in patients with CLTI. Long-term administration of evolocumab over 12 months contributed to improving proportion of wound-free limb salvage.
Radiofrequency catheter ablation (RFCA) to treat ventricular arrhythmias (VAs) originating below the His bundle (HB) region of the right ventricular (RV) septum could impair the atrioventricular node ...conduction. This study aimed to clarify the parameters of the 12-lead electrocardiography that predict successful RFCA of VAs originating from this region. This study included 20 consecutive patients (13 men; mean age, 68 ± 7 years) with monomorphic VAs in whom the earliest ventricular activation during the VA was below the HB region of the RV septum. According to the ablation results, the patients were divided into two groups: successful ablation (S-group; n = 10) and failed ablation groups (F-group; n = 10). The electrocardiographic parameters during the VAs and RFCA results were assessed. The R wave amplitudes in leads aVL (P = 0.001) and I (P = 0.010) in the S-group were both smaller than those in the F-group. In addition, the S-group had smaller negative deflection amplitudes in leads III (P = 0.002) and aVF (P = 0.003) than the F-group. According to the receiver operating characteristic curve analysis, the most useful electrocardiographic parameter for predicting successful ablation was the R wave amplitude in lead aVL (area under the curve, 0.895; P < 0.001); a cutoff value of < 1.3 mV predicted a successful RFCA with the highest accuracy (sensitivity, 90%; specificity, 80%; positive predictive value, 82%; negative predictive value, 89%). The R wave amplitude in lead aVL was the most useful parameter for predicting a successful RFCA to treat VAs originating below the HB region of the RV septum.
Cultured cells easily develop resistance to kinesin-5 inhibitors (K5Is) often by overexpressing a related motor protein, kinesin-12/KIF15, or by acquiring mutations in the N-terminal motor domain of ...kinesin-5/KIF11 itself. We aimed to identify novel mechanisms responsible for resistance to S-trityl L-cysteine (STLC), one of the K5Is, using human osteosarcoma cell lines. Among six lines examined, U-2OS and HOS survived chronic STLC treatment and gave rise to resistant cells with IC50s at least 10-fold higher than those of the respective parental lines. Depletion of KIF15 largely eliminated the acquired K5I resistance in both cases, consistent with the proposed notion that KIF15 is indispensable for it. In contrast to the KIF11-independent property of the cells derived from HOS, those derived from U-2OS still required KIF11 for their growth and, intriguingly, expressed a C-terminal truncated variant of KIF11 resulting from a frame shift mutation (S1017fs). All of the isolated clones harbored the same mutation, suggesting its clonal expansion in the cell population due to the growth advantage during chronic STLC treatment. Transgenic expression of KIF11S1017fs in the parental U-2OS cells, as well as in HeLa cells, conferred a moderate but reproducible STLC resistance, probably owing to STLC-resistant localization of the mutant KIF11 on mitotic spindle. Our observations indicate that both KIF15 and the C-terminal-truncated KIF11 contributes to the STLC resistance of the U-2OS derived cells.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
BRCA‐related breast carcinoma can be prevented through prophylactic surgery and an intensive follow‐up regimen. However, BRCA genetic tests cannot be routinely performed, and some BRCA mutations ...could not be defined as deleterious mutations or normal variants. Therefore, an easy functional assay of BRCA will be useful to evaluate BRCA status. As it has been reported that BRCA functions in the regulation of centrosome number, we focused on centrosome number in cancer tissues. Here, 70 breast cancer specimens with known BRCA status were analyzed using immunofluorescence of γ‐tubulin (a marker of centrosome) foci. The number of foci per cell was higher in cases with BRCA mutation compared to wild‐type cases, that is, 1.9 (95% confidence interval CI, 1.5‐2.3) vs 0.5 (95% CI, 0.2‐0.8) (P < .001). Specifically, foci numbers per cell in BRCA1 and BRCA2 mutation cases were 1.2 (95% CI, 0.6‐1.8) and 2.2 (95% CI, 1.7‐2.6), respectively, both higher than those in wild‐type cases (P = .042 and P < .0001, respectively). The predictive value of γ‐tubulin foci as determined by area under the curve (AUC = 0.86) for BRCA status was superior to BRCAPRO (AUC = 0.69), Myriad Table (AUC = 0.61), and KOHBRA BRCA risk calculator (AUC = 0.65) pretest values. The use of γ‐tubulin foci to predict BRCA status had sensitivity = 83% (19/23), specificity = 89% (42/47), and positive predictive value = 77% (20/26). Thus, γ‐tubulin immunofluorescence, a functional assessment of BRCA, can be used as a new prospective test of BRCA status.
For predicting BRCA status by an immunostaining method, we focused the number of centrosome, because both of BRCA1 and 2 proteins control centrosome duplication. Our test was based on a quantitative analysis of centrosome numbers (γ‐tubulin foci) in cells in breast cancer tissue specimens. The number of foci per cell was higher in BRCA1 and/or 2 mutation as compared to wild‐type cases, i.e., 1.7 95%; confidence interval (CI): 1.3‐2.2 vs 0.7 (95% CI: 0.4‐1.0) (P < .001). The predictive value of our method for BRCA status (AUC = 0.86) was superior to that of existing pretests that are based on clinico‐pathological information, and had a sensitivity of 83%, specificity of 89%, and positive predictive value of 77%. We believe that this contribution is theoretically and practically relevant because, BRCA genetic tests have some problems, cost, privacy and counseling and cannot be routinely carried out in clinical practice; therefore, a more straightforward and broadly applicable test is needed.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Objectve To assess the impact of glycemic variability on blood pressure in hospitalized patients with cardiac disease. Methods In 40 patients with cardiovascular disease, the glucose levels were ...monitored by flash continuous glucose monitoring (FGM; Free-Style Libre™ or Free-Style Libre Pro; Abbott, Witney, UK) and self-monitoring blood glucose (SMBG) for 14 days. Blood pressure measurements were performed twice daily (morning and evening) at the same time as the glucose level measurement using SMBG. Results The detection rate of hypoglycemia using the FGM method was significantly higher than that with the 5-point SMBG method (77.5% vs. 5.0%, p<0.001). Changes in the systolic blood pressure from evening to the next morning morning - evening (ME) difference were significantly correlated with night glucose variability (r=0.63, P<0.001). A multiple regression analysis showed that night glucose variability using FGM was more closely correlated with the ME difference r=0.62 (95% confidence interval, 0.019-0.051); p<0.001 than with the age, body mass index, or smoking history. Night glucose variability was also more closely associated with the ME difference in patients with unstable angina pectoris (UAP) than in those with acute myocardial infarction (AMI) or heart failure (HF) (r=0.83, p=0.058). Conclusion Night glucose variability is associated with the ME blood pressure difference, and FGM is more accurate than the 5-point SMBG approach for detecting such variability.
The evaluation of angiogenesis inhibitors requires the analysis of the precise structure and function of tumor vessels. The anti-angiogenic agents lenvatinib and sorafenib are multi-target tyrosine ...kinase inhibitors that have been approved for the treatment of hepatocellular carcinoma (HCC). However, the different effects on tumor vasculature between lenvatinib and sorafenib are not well understood. In this study, we analyzed the effects of both drugs on vascular structure and function, including vascular normalization, and investigated whether the normalization had a positive effect on a combination therapy with the drugs and radiation using micro X-ray computed tomography with gold nanoparticles as a contrast agent, as well as immunohistochemical analysis and interstitial fluid pressure (IFP) measurement. In mice subcutaneously transplanted with mouse HCC cells, treatment with lenvatinib or sorafenib for 14 days inhibited tumor growth and reduced the tumor vessel volume density. However, analysis of integrated data on vessel density, rates of pericyte-covering and perfused vessels, tumor hypoxia, and IFP measured 4 days after drug treatment showed that treatment with 3 mg/kg of lenvatinib significantly reduced the microvessel density and normalized tumor vessels compared to treatment with 50 mg/kg of sorafenib. These results showed that lenvatinib induced vascular normalization and improved the intratumoral microenvironment in HCC tumors earlier and more effectively than sorafenib. Moreover, such changes increased the radiosensitivity of tumors and enhanced the effect of lenvatinib and radiation combination therapy, suggesting that this combination therapy is a powerful potential application against HCC.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
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