Stepwise prolongation of overall survival from first to third generation EGFR-TKIs for EGFR mutation-positive non-small-cell lung cancer: the Tokushukai REAl-world Data project (TREAD 01).

OBJECTIVE: The introduction of new-generation epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) has afforded promising overall survival outcomes in clinical trials for non-small-cell lung cancer. We aim to investigate the current adoption rate of these agents and the real-world impact on overall survival among institutions. METHODS: In a nationwide retrospective cohort study of 46 Tokushukai Medical Group hospitals in Japan, we analyzed clinical data of consecutive patients with non-small-cell lung cancer receiving EGFR-TKIs between April 2010 and March 2020. Univariate and multivariate Cox regression analyses examined the associations between overall survival and patient/tumor-related factors and first-line EGFR-TKIs. RESULTS: A total of 758 patients (58.5% females; median age, 73 years) were included. Of 40 patients diagnosed in 2010, 72.5% received gefitinib, whereas 81.3% of 107 patients diagnosed in 2019 received osimertinib as the first-line EGFR-TKI. With a median follow-up of 15.8 months, the median overall survival was 28.4 months (95% confidence interval, 15.3-31.0). In a multivariate Cox regression analysis, age, body mass index, disease status, EGFR mutational status and first-line epidermal growth factor receptor tyrosine kinase inhibitor were identified as significant prognostic factors after adjusting for background factors including study period, hospital volume and hospital type. The estimated 2-year overall survival rates for gefitinib, erlotinib, afatinib and osimertinib were 70.1% (95% confidence interval 59.7-82.4), 67.8% (95% confidence interval 55.3-83.2), 75.5% (95% confidence interval 64.7-88.0) and 90.8% (95% confidence interval 84.8-97.3), respectively. The median time to treatment failure of gefitinib, erlotinib, afatinib and osimertinib were 12.8, 8.8, 12.0 and 16.9 months or more, respectively. CONCLUSIONS: Our real-world data revealed that the swift and widespread utilization of newer-generation EGFR-TKIs in patients with EGFR mutation-positive non-small-cell lung cancer, and that these newer-generation EGFR-TKIs can prolong overall survival regardless of hospital volume or type. Therefore, osimertinib could be a reasonable first choice treatment for these patients across various clinical practice settings.

Internal doses in experimental mice and rats following exposure to neutron-activated 56MnO2 powder: results of an international, multicenter study.

The experiment was performed in support of a Japanese initiative to investigate the biological effects of irradiation from residual neutron-activated radioactivity that resulted from the A-bombing. Radionuclide 56Mn (T1/2 = 2.58 h) is one of the main neutron-activated emitters during the first hours after neutron activation of soil dust particles. In our previous studies (2016-2017) related to irradiation of male Wistar rats after dispersion of 56MnO2 powder, the internal doses in rats were found to be very inhomogeneous: distribution of doses among different organs ranged from 1.3 Gy in small intestine to less than 0.0015 Gy in some of the other organs. Internal doses in the lungs ranged from 0.03 to 0.1 Gy. The essential pathological changes were found in lung tissue of rats despite a low level of irradiation. In the present study, the dosimetry investigations were extended: internal doses in experimental mice and rats were estimated for various activity levels of dispersed neutron-activated 56MnO2 powder. The following findings were noted: (a) internal radiation doses in mice were several times higher in comparison with rats under similar conditions of exposure to 56MnO2 powder. (b) When 2.74 × 108 Bq of 56MnO2 powder was dispersed over mice, doses of internal irradiation ranged from 0.81 to 4.5 Gy in the gastrointestinal tract (small intestine, stomach, large intestine), from 0.096 to 0.14 Gy in lungs, and doses in skin and eyes ranged from 0.29 to 0.42 Gy and from 0.12 to 0.16 Gy, respectively. Internal radiation doses in other organs of mice were much lower. (c) Internal radiation doses were significantly lower in organs of rats with the same activity of exposure to 56MnO2 powder (2.74 × 108 Bq): 0.09, 0.17, 0.29, and 0.025 Gy in stomach, small intestine, large intestine, and lungs, respectively. (d) Doses of internal irradiation in organs of rats and mice were two to four times higher when they were exposed to 8.0 × 108 Bq of 56MnO2 (in comparison with exposure to 2.74 × 108 Bq of 56MnO2). (e) Internal radiation doses in organs of mice were 7-14 times lower with the lowest 56MnO2 amount (8.0 × 107 Bq) in comparison with the highest amount, 8.0 × 108 Bq, of dispersed 56MnO2 powder. The data obtained will be used for interpretation of biological effects in experimental mice and rats that result from dispersion of various levels of neutron-activated 56MnO2 powder, which is the subject of separate studies.

Internal exposure to neutron-activated 56Mn dioxide powder in Wistar rats: part 1: dosimetry.

There were two sources of ionizing irradiation after the atomic bombings of Hiroshima and Nagasaki: (1) initial gamma-neutron irradiation at the moment of detonation and (2) residual radioactivity. Residual radioactivity consisted of two components: radioactive fallout containing fission products, including radioactive fissile materials from nuclear device, and neutron-activated radioisotopes from materials on the ground. The dosimetry systems DS86 and DS02 were mainly devoted to the assessment of initial radiation exposure to neutrons and gamma rays, while only brief considerations were given for the estimation of doses caused by residual radiation exposure. Currently, estimation of internal exposure of atomic bomb survivors due to dispersed radioactivity and neutron-activated radioisotopes from materials on the ground is a matter of some interest, in Japan. The main neutron-activated radionuclides in soil dust were 24Na, 28Al, 31Si, 32P, 38Cl, 42K, 45Ca, 46Sc, 56Mn, 59Fe, 60Co, and 134Cs. The radionuclide 56Mn (T 1/2 = 2.58 h) is known as one of the dominant beta- and gamma emitters during the first few hours after neutron irradiation of soil and other materials on ground, dispersed in the form of dust after a nuclear explosion in the atmosphere. To investigate the peculiarities of biological effects of internal exposure to 56Mn in comparison with external gamma irradiation, a dedicated experiment with Wistar rats exposed to neutron-activated 56Mn dioxide powder was performed recently by Shichijo and coworkers. The dosimetry required for this experiment is described here. Assessment of internal radiation doses was performed on the basis of measured 56Mn activity in the organs and tissues of the rats and of absorbed fractions of internal exposure to photons and electrons calculated with the MCNP-4C Monte Carlo using a mathematical rat phantom. The first results of this international multicenter study show that the internal irradiation due to incorporated 56Mn powder is highly inhomogeneous, and that the most irradiated organs of the experimental animals are: large intestine, small intestine, stomach, and lungs. Accumulated absorbed organ doses were 1.65, 1.33, 0.24, 0.10 Gy for large intestine, small intestine, stomach, and lungs, respectively. Other organs were irradiated at lower dose levels. These results will be useful for interpretation of the biological effects of internal exposure of experimental rats to powdered 56Mn as observed by Shichijo and coworkers.

High Excess Risk of Heart Disease Mortality among Hiroshima Atomic Bomb Male Survivors Exposed Near the Hypocenter.

Heart disease (HD) mortality is the second leading cause of death in Japan. The HD mortality risk among Atomic bomb survivors is slightly positive but shows a statistically significant dose-response relationship with initial radiation dose, as reported by the Radiation Effects Research Foundation. In that report, dosimetry was based on initial radiation only, with the effect of indirect radiation dose not taken into consideration. The atomic bomb radiation, however, consisted of both initial and residual radiation. We reevaluated the dose-response relationship for HD mortality using exposure distance (ground distance between the location where exposed and the hypocenter) as a surrogate indicator of radiation dose. At Hiroshima University, a cohort study has been conducted with Hiroshima Atomic Bomb Survivors (ABS) since 1970. We selected 29605 subjects from the ABS who were exposed at 3.5 km or less from the hypocenter and alive on January 1, 1970. These subjects, referred to as "Hiroshima hibakusha" in this paper, were followed until December 31, 2010. We stratified the cohort data with respect to sex and age at the time of bombing (ATB) into 10-year age groups. For each stratum, by applying an extended Cox regression model with time-dependent covariates, we analyzed the risk of HD mortality using either initial radiation dose or exposure distance as an explanatory variable. The results indicate a high excess risk in males and older age ATB females who were exposed near the hypocenter. This difference may be explained by the effect of female sex hormone on the circulatory system among young age ATB females. Some unknown risk factor related to exposure distance was also implicated in the elevated risk of HD among the Hiroshima hibakusha, especially in males. This necessitates further study.

High Initial-dose Dependency of Cerebrovascular Disease Mortality among Female Survivors of the Hiroshima Atomic Bomb Exposed in Teens: A Cohort Study, 1970-2010.

Several studies have been conducted on cerebrovascular disease mortality in Atomic bomb survivors. Previous studies have investigated the relationship between mortality and initial radiation dose after adjusting for the effects of sex and age at the time of the bombing (ATB), and detected a weak (but statistically significant) dose-response relationship was detected. The objective of the present study was to examine whether the sex- and age ATB-specific cerebrovascular disease mortality among Hiroshima atomic bomb survivors can be explained by the initial radiation dose. At Hiroshima University, a cohort study has been conducted with Hiroshima Atomic Bomb Survivors (ABS) since 1970. We selected 30,378 subjects from the ABS who were exposed at 3.5 km or less from the hypocenter and still alive on January 1, 1970. These subjects were followed up until December 31, 2010. The cohort data were stratified with respect to sex and age ATB into 10-year age groups. For each stratum, using Cox regression, we performed survival analyses of the risk of cerebrovascular mortality using the initial radiation dose and the exposure distance (the ground distance between the exposure location and the hypocenter) as explanatory variables. The results indicated that the risks to females exposed at 10 to 19 years old were highly dependent on the initial radiation dose (hazard ratio: 1.51, p < 0.001), while the risks to males were not. There might exist some radiation exposure effects limited to women who were in their teens at the time of exposure. However, the background mechanisms remain unclear, necessitating further study.

Contribution of radioactive particles to the post-explosion exposure of atomic bomb survivors implied from their stable chromosome aberration rates.

Even today when nearly 80 years have passed after the atomic bomb (A-bomb) was dropped, there are still debates about the exact doses received by the A-bomb survivors. While initial airborne kerma radiation (or energy spectrum of emitted radiation) can be measured with sufficient accuracy to assess the radiation dose to A-bomb survivors, it is not easy to accurately assess the neutron dose including appropriate weighting of neutron absorbed dose. Particularly, possible post-explosion exposure due to the radioactive particles generated through neutron activation have been almost neglected so far, mainly because of a large uncertainty associated to the behavior of those particles. However, it has been supposed that contribution of such non-initial radiation exposure from the neutron-induced radioactive particles could be significant, according to the findings that the stable chromosomal aberration rates which indicate average whole-body radiation doses were found to be more than 30% higher for those exposed indoors than for those outdoors even at the same initial dose estimated for the Life Span Study. In this Mini Review article, the authors explain that such apparently controversial observations can be reasonably explained by assuming a higher production rate of neutron-induced radioactive particles in the indoor environment near the hypocenter.

Prognostic impact of concomitant pH-regulating drugs in patients with non-small cell lung cancer receiving epidermal growth factor receptor tyrosine kinase inhibitors: the Tokushukai REAl-world Data project 01-S1.

PURPOSE: This study aimed to examine the prognostic impact of concomitant pH-regulating drug use in patients with epidermal growth factor receptor (EGFR)-mutation-positive non-small-cell lung cancer (NSCLC) receiving EGFR-tyrosine kinase inhibitors (TKIs). METHODS: We conducted a nationwide retrospective cohort study and reviewed clinical data of consecutive patients with NSCLC treated with the first-line EGFR-TKIs in 46 hospitals between April 2010 and March 2020. Cox regression analyses were conducted to examine the differences in overall survival (OS) between patients treated with and without concomitant pH-regulating drugs, including potassium-competitive acid blockers (P-CABs), proton pump inhibitors (PPIs), and H2-receptor antagonists (H2RAs). RESULTS: A total of 758 patients were included in the final dataset, of which 307 (40%) were administered concomitant pH-regulating drugs while receiving frontline EGFR-TKIs. After adjusting for basic patient characteristics, patients administered gefitinib, erlotinib, afatinib, and osimertinib with concomitant pH-regulating drugs had lower OS than those without concomitant pH-regulating drugs, with hazard ratios of 1.74 (with a 95% confidence interval of 1.34-2.27), 1.33 (0.80-2.22), 1.73 (0.89-3.36), and 5.04 (1.38-18.44), respectively. The 2-year OS rates of patients receiving gefitinib with or without concomitant pH-regulating drugs were 65.4 and 77.5%, those for erlotinib were 55.8 and 66.6%, and those for afatinib were 63.2 and 76.9%, respectively. The 1-year OS rates of patients receiving osimertinib with or without concomitant pH-regulating drugs were 88.1% and 96.9%, respectively. CONCLUSION: In addition to the first-generation EGFR-TKIs, the second- and third-generation EGFR-TKIs also resulted in OS deterioration in patients with EGFR mutation-positive NSCLC when used concurrently with pH-regulating drugs.

TMEM158 and FBLP1 as novel marker genes of cisplatin sensitivity in non-small cell lung cancer cells.

Even after development of molecular targeting therapies, platinum-based chemotherapy is still a standard care for treatment of locally advanced non-small cell lung cancer (NSCLC). So far, critical molecular markers capable to predict the therapeutic response in NSCLC patients remain undetermined. We here attempted to identify novel biomarker genes for cisplatin (CDDP) for a tailored therapy. Initial screening to explorer association of IC(50) values of CDDP obtained by MTT assay and gene expression levels measured with oligonucleotide microarray and real-time RT-PCR provided 6 candidate genes, namely, NUBPL, C9orf30, ZNF12, TMEM158, GSK3B, and FBLP1 using 9 lung cancer cells consisting of 3 small and 6 NSCLC cells. These 6 genes together with 5 reported biomarkers, i.e., GSTP1, ERCC1, BRCA1, FRAP1, and RRM1, were subjected to a linear regression analysis using 12 NSCLC cell lines including 6 additional NSCLC cells: only FBLP1 and TMEM158 genes showed positive associations with statistical significances (P = .016 and .026, respectively). The biological significance of these genes was explored by in vitro experiments: Knockdown experiments in PC-9/CDDP cells revealed that the reduced expression of TMEM158 significantly decreased the chemo-resistance against CDDP (P <.0001), while 2 transformants of PC-6 cells stably over-expressing FBLP1 resulted in an enhanced resistance to CDDP (P = .004 and P = .001). Furthermore, a stepwise multiple regression analysis demonstrated the best prediction formula could be fixed when we used expression data of TMEM158 and FBLP1 (R(2) = 0.755, P = .0018). TMEM158 and FBLP1 may be powerful predictive biomarkers for CDDP therapy in NSCLC.

Effect of age on the fractional anisotropy (FA) value of peripheral nerves and clinical significance of the age-corrected FA value for evaluating polyneuropathies.

INTRODUCTION: We investigated the correlation between age and the fractional anisotropy (FA) values of peripheral nerves in healthy adults and compared the age-corrected FA values of peripheral nerves in healthy subjects and patients with polyneuropathy. METHODS: The institutional review board approved this study and informed consent was obtained from all participants before entry into the study. We optimized diffusion tensor imaging using a 3-T magnetic resonance scanner and an extremity coil for scanning tibial nerves. The effect of age and sex on the FA values of tibial nerves in healthy volunteers was investigated and the age-corrected FA values of tibial nerves in healthy volunteers and patients with polyneuropathy were compared. RESULTS: The maximum FA values of the tibial nerves remained constant until age 45 (approximately 0.516); they subsequently decreased by 0.004/year in healthy volunteers. After removing the effect of age with an age-adjusted equation, the median maximum FA values in the volunteers and patients were 0.518 (range, 0.406-0.616) and 0.442 (range, 0.376-0.530), respectively. The age-corrected FA values were significantly lower in the patients than the healthy volunteers (p < 0.001). There was no significant gender-related difference in the maximum FA values of the tibial nerves (p = 0.416). CONCLUSION: The age-corrected FA value of the peripheral nerves helps to differentiate between age-related peripheral nerve degeneration and polyneuropathies.

Investigation on circular asymmetry of geographical distribution in cancer mortality of Hiroshima atomic bomb survivors based on risk maps: analysis of spatial survival data.

While there is a considerable number of studies on the relationship between the risk of disease or death and direct exposure from the atomic bomb in Hiroshima, the risk for indirect exposure caused by residual radioactivity has not yet been fully evaluated. One of the reasons is that risk assessments have utilized estimated radiation doses, but that it is difficult to estimate indirect exposure. To evaluate risks for other causes, including indirect radiation exposure, as well as direct exposure, a statistical method is described here that evaluates risk with respect to individual location at the time of atomic bomb exposure instead of radiation dose. In addition, it is also considered to split the risks into separate risks due to direct exposure and other causes using radiation dose. The proposed method is applied to a cohort study of Hiroshima atomic bomb survivors. The resultant contour map suggests that the region west to the hypocenter has a higher risk compared to other areas. This in turn suggests that there exists an impact on risk that cannot be explained by direct exposure.

Solid cancer mortality risk among a cohort of Hiroshima early entrants after the atomic bombing, 1970-2010: implications regarding health effects of residual radiation.

There are two types of exposure to atomic bomb (A-bomb) radiation: exposure to initial radiation released at the time of the detonation of the bomb, and exposure to residual radiation, which remains afterwards. Health hazards caused by exposure from residual radiation have not yet been clarified. The purpose of our study was to reveal the relationships between mortality risk from solid cancer and residual radiation based on data from the early entrants to Hiroshima. It is hard to identify the individual residual radiation doses. However, these are assumed to depend on the date of entry and the entrants' behavior. Individual behavior is thought to be closely related to gender and age at exposure. We investigated a cohort of 45 809 individuals who were living in Hiroshima Prefecture on 1 January 1970 and were registered on the Database of Atomic Bomb Survivors as entrants after the bombing. Poisson regression methods were used to estimate excess relative risks (ERR) with data cross-classified by sex, age at entry, and date of entry. In males in their 20s, 30s, and 40s at entry and in females less than 10 years old and in their 40s at entry, solid cancer mortality risks were significantly higher among persons who entered the city on the day of the bombing than those who entered three or more days later. With adjustments for the age-dependent sensitivities to radiation exposure, it was extrapolated that middle-aged people who entered the city on the day of the bombing were exposed to higher levels of residual radiation than younger people.

Effects of internal exposure to neutron-activated 56MnO2 powder on locomotor activity in rats.

At the detonation of the atomic bombing in Hiroshima and Nagasaki, a significant amount of radionuclides was produced by the neutron induced activation. The residual radiation from the explosion is crucial to the health risk of the people who entered these cities after the bombing and might have inhaled these radioactive materials. Because 56Mn is one of the major radionuclides produced in soil and have not been studied until now, we had conducted a series of experiments using rats to investigate the biological impacts of exposure of 56MnO2 particles. In these experiments, the rats' spontaneous locomotor activity was also assessed to examine the possible effects of 56Mn on their behavior. However, the locomotor activity data obtained from an individual experiment failed to identify radiation effects due to the large variation among animals and the small sample size. In the present study, all available data from our previous studies on 56MnO2 exposure (0.02-0.15 Gy of whole-body doses) as well as 60Co-γ exposure (at 2-5 Gy of whole-body doses) were pooled. Our statistical method, which takes into account individual differences and daily fluctuations, successfully identified a decrease in locomotor activity caused by 56MnO2 exposure, where the changes were gradual and reached the maximum reduction around 2 weeks after exposure. In contrast, exposure to 60Co-γ rays produced the highest decline of activity within one day. These results suggest that internal exposure to 56Mn at whole-body doses of even less than 0.15 Gy may have a long-lasting impact on locomotor activity.

Telomerase activation without shortening of telomeric 3'-overhang is a poor prognostic factor in human colorectal cancer.

Our previous report demonstrated a good correlation between high telomerase activity of cancer tissues and a poor prognosis of patients with colorectal cancers, except for several cases. To elucidate the additional factors that contribute to patient prognosis, the correlation among the expression levels of telomere binding proteins (TBP), the lengths of telomeres, the lengths of telomere 3'-overhang (3'-OH) and telomerase activity in 106 paired colorectal cancer and corresponding noncancerous mucosa (NCM) specimens were examined. The expression levels of eight TBP genes (TRF1, TRF2, TIN2, TANK1, TANK2, POT1, RAP1 and TPP1) were analyzed. Among the 106 cases, 35 cases had shortened telomeres (<7 kb), 15 had shortened 3'-OH (3'-OH length ratio of cancer/NCM <0.5) and 88 were classified as telomerase-activated cancers (activity ratio of cancer/NCM >2). Comparison between NCM and cancer in each case showed that all TBP except for POT1 were downregulated in cancers. A survival analysis using a Cox proportional hazard model showed that the survival rate of the telomerase-activated cases with shortened 3'-OH and that of telomerase-inactivated cases were significantly better than that of telomerase-activated cases without 3'-OH shortening, that is, restored or maintained 3'-OH (P = 0.018). In the telomerase-activated cancers, the length of 3'-OH was significantly correlated with the expression levels of POT1. Elongation of telomeric overhang by telomerase, which might be regulated by POT1, may contribute to the increase of malignant potential in colorectal cancers.

A robust method for estimating gene expression states using Affymetrix microarray probe level data.

BACKGROUND: Microarray technology is a high-throughput method for measuring the expression levels of thousand of genes simultaneously. The observed intensities combine a non-specific binding, which is a major disadvantage with microarray data. The Affymetrix GeneChip assigned a mismatch (MM) probe with the intention of measuring non-specific binding, but various opinions exist regarding usefulness of MM measures. It should be noted that not all observed intensities are associated with expressed genes and many of those are associated with unexpressed genes, of which measured values express mere noise due to non-specific binding, cross-hybridization, or stray signals. The implicit assumption that all genes are expressed leads to poor performance of microarray data analyses. We assume two functional states of a gene - expressed or unexpressed - and propose a robust method to estimate gene expression states using an order relationship between PM and MM measures. RESULTS: An indicator 'probability of a gene being expressed' was obtained using the number of probe pairs within a probe set where the PM measure exceeds the MM measure. We examined the validity of the proposed indicator using Human Genome U95 data sets provided by Affymetrix. The usefulness of 'probability of a gene being expressed' is illustrated through an exploration of candidate genes involved in neuroblastoma prognosis. We identified the candidate genes for which expression states differed (un-expressed or expressed) when compared between two outcomes. The validity of this result was subsequently confirmed by quantitative RT-PCR. CONCLUSION: The proposed qualitative evaluation, 'probability of a gene being expressed', is a useful indicator for improving microarray data analysis. It is useful to reduce the number of false discoveries. Expression states - expressed or unexpressed - correspond to the most fundamental gene function 'On' and 'Off', which can lead to biologically meaningful results.

Optimal fast T2-weighted magnetic resonance microscopy imaging of the eye and its clinical application.

PURPOSE: To compare a half-Fourier single-shot rapid acquisition with relaxation enhancement (RARE) sequence with a balanced steady-state free precession (b-SSFP) sequence in the evaluation of the eye using magnetic resonance (MR) microscopy imaging and to clarify the usefulness of RARE microscopy imaging in evaluating nonoperative glaucoma patients and patients who have undergone surgery for glaucoma or cataract. MATERIALS AND METHODS: One-mm and 2-mm slice thickness images of RARE sequence and b-SSFP sequence using a 1.5 T MR unit and a 23-mm microscopy coil were obtained in eight healthy volunteers. The signal-to-noise (S/N) ratio of aqueous humor in the anterior chamber was measured quantitatively and visualization of the anterior chamber anatomy was assessed qualitatively. Furthermore, we evaluated 21 glaucoma patients (including six postoperative patients) and four patients after cataract surgery with 2-mm slice thickness RARE MRI. RESULTS: The 2-mm slice thickness RARE imaging had a significantly greater S/N ratio than the 1-mm slice thickness RARE imaging (P < 0.05) and acquired the best image quality among the four types of images (P < 0.01). Additionally, 2-mm slice thickness RARE microscopy imaging could depict anterior chamber anatomy of glaucoma eyes and eyes after cataract surgery. CONCLUSION: We believe that optimal fast T2-weighted MR microimaging might become a useful ophthalmologic examination technique.

Quantitative Analysis of Effects of a Single 60Co Gamma Ray Point Exposure on Time-Dependent Change in Locomotor Activity in Rats.

Investigating initial behavioral changes caused by irradiation of animals might provide important information to aid understanding of early health effects of radiation exposure and clinical features of radiation injury. Although previous studies in rodents suggested that radiation exposure leads to reduced activity, detailed properties of the effects were unrevealed due to a lack of proper statistical analysis, which is needed to better elucidate details of changes in locomotor activity. Ten-week-old male Wistar rats were subjected to single point external whole-body irradiation with 60Co gamma rays at 0, 2.0, 3.5, and 5.0 Gy (four rats per group). Infrared sensors were used to continuously record the locomotor activity of each rat. The cumulative number of movements during the night was defined as "activity" for each day. A non-linear mixed effects model accounting for individual differences and daily fluctuation of activity was applied to analyze the rats' longitudinal locomotor data. Our statistical method revealed characteristics of the changes in locomotor activity after radiation exposure, showing that (1) reduction in activity occurred immediately-and in a dose-dependent manner-after irradiation and (2) recovery to pre-irradiation levels required almost one week, with the same recovery rate in each dose group.

Selection of a novel drug-response predictor in esophageal cancer: a novel screening method using microarray and identification of IFITM1 as a potent marker gene of CDDP response.

Prior laboratory prediction of individual drug response is of key importance in esophageal squamous cell carcinoma (ESCC), because of the extremely narrow therapeutic index of chemotherapy. However, very few critical markers have been validated to date for ESCC. We previously demonstrated that simultaneous performance of two different types of comprehensive gene expression analysis might provide a way to identify potent marker genes for drug sensitivity from the expression-sensitivity correlation analysis alone, but the screening method appeared not to be always effective. Therefore, we attempted to identify novel potent marker genes using a new statistical analysis of oligonucleotide microarray expression data, based on a two-dimensional mixed normal model, and selected 3 and 7 novel candidates for 5-fluorouracil (5-FU) and cis-platinum (CDDP), respectively. Interferon induced transmembrane protein 1 (IFITM1) gene alone, being suggested as a key gene of Wnt pathway, was commonly selected in both screening methods. The transfection analyses and siRNA-mediated knock-down experiments revealed that expression of IFITM1 closely related to cellular sensitivity to CDDP. Considering the fact that drug sensitivity is determined by multiple genes, we established the best linear model using quantified expression data of a set of all the selected marker genes including IFITM1, which converted the quantified expression data of ESCC cell lines into an IC50 value of each drug. In the same way, using the representative genes selected in vitro, we developed highly predictive formulae for disease-free survival (DFS) of the CDDP/5-FU combination after curative operation in esophageal cancer patients (R=0.917). A two-dimensional mixed normal model can be a powerful tool to identify novel drug-response determinants, and the IFITM1 gene selected by the statistical method a novel critical biomarker of CDDP response in ESCC.

Regression analysis of maternal smoking effect on birth weight.

Risk factors for low birth weight (LBW) have been indicated in many studies, but in Japan few studies have examined the amount of reduction in birth weight (BW). The goal of this study was to examine the association between maternal smoking among pregnant women and subsequent reduction in BW, corrected by the effect of potential confounders. To assess the effect of background factors other than maternal smoking, we established a model to estimate the contribution of each explanatory variable using logarithmic multiple regression. We then used the adjusted BW model to evaluate the direct effect of maternal smoking. To obtain information on maternal characteristics, including smoking status and characteristics of their infants, including BW, we conducted a questionnaire survey. After statistical adjustment for background factors, the mean of BW among infants of participants who smoked during pregnancy was roundtable significantly lower than that of non-smoking participants, but there was no verification of a dose-response relationship. However, mean BWs were not significantly different when comparing participants who quit smoking during pregnancy to non-smoking participants, suggesting that stopping smoking during pregnancy is beneficial.

Prediction of individual response to platinum/paclitaxel combination using novel marker genes in ovarian cancers.

We attempted to identify potent marker genes using a new statistical analysis and developed a prediction system for individual response to platinum/paclitaxel combination chemotherapy in ovarian cancer patients based on the hypothesis that expression analysis of a set of the key drug sensitivity genes for platinum and paclitaxel could allow us to predict therapeutic response to the combination. From 10 human ovarian cancer cell lines, genes correlative in the expression levels with cytotoxicities of cisplatin (CDDP) and paclitaxel were chosen. We first selected five reliable prediction markers for the two drugs from 22 genes already known as sensitivity determinants and then identified another 8 novel genes through a two-dimensional mixed normal model using oligomicroarray expression data. Using expression data of genes quantified by real-time reverse transcription-PCR, we fixed the best linear model, which converted the quantified expression data into an IC(50) of each drug. Multiple regression analysis of the selected genes yielded three prediction formulae for in vitro activity of CDDP and paclitaxel. In the same way, using the same genes selected in vitro, we then attempted to develop prediction formulae for progression-free survival to the platinum/paclitaxel combination. We therefore constructed possible formulae using different sets of 13 selected marker genes (5 known and 8 novel genes): Utility confirmation analyses using another nine test samples seemed to show that the formulae using a set of 8 novel marker genes alone could accurately predict progression-free survival (r = 0.683; P = 0.042).

Chemosensitivity prediction in esophageal squamous cell carcinoma: novel marker genes and efficacy-prediction formulae using their expression data.

Esophageal cancer is a highly lethal disease and the optimal therapy remains unclear. Since adjuvant chemotherapy gives a better chance of survival, we attempted to develop a chemosensitivity prediction model to improve individual responses to therapy. Comprehensive gene expression analyses (cDNA and oligonucleotide microarrays) and MTT assay of 8 drugs in 20 KYSE squamous cell carcinoma cell lines were performed to distinguish candidate marker genes whose expression levels reproducibly correlated with cellular drug sensitivities. After confirmation with real-time RT-PCR, we performed multiple regression analyses to develop drug-sensitivity prediction formulae using the quantified expression data of selected marker genes. Using the same sets of genes, we also constructed prediction models for individual clinical responses to 5-FU-based chemotherapy using 18 cases. We selected 5 better marker genes, known as drug sensitivity determinants, identified 9 novel predictive genes for 4 of 8 anticancer drugs [5-FU, CDDP, DOX, and CPT-11 (SN-38)], and developed highly predictive formulae of in vitro sensitivities to the 4 drugs and clinical responses to 5-FU-based adjuvant chemotherapies in terms of overall and disease-free survivals. Our selected genes are likely to be effective drug-sensitivity markers and formulae using the 9 novel genes would provide advantages in prediction.