Monitoring of somatic mutations in circulating cell-free DNA by digital PCR and next-generation sequencing during afatinib treatment in patients with lung adenocarcinoma positive for EGFR activating mutations.

Background: Analysis of circulating cell-free DNA (cfDNA) is under intensive investigation for its potential to identify tumor somatic mutations. We have now explored the usefulness of such liquid biopsy testing with both the digital polymerase chain reaction (dPCR) and next-generation sequencing (NGS) during treatment of patients with the epidermal growth factor receptor (EGFR) inhibitor afatinib. Patients and methods: Eligible patients had advanced lung adenocarcinoma with EGFR activating mutations and were treated with afatinib. Plasma samples were collected before and during (4 and 24 weeks) afatinib treatment as well as at disease progression. Tumor and plasma DNA were analyzed by dPCR and NGS. Results: Thirty-five patients were enrolled. The objective response rate and median progression-free survival (PFS) were 77.1% and 13.8 months, respectively. Tumor and plasma DNA were available for 32 patients. dPCR and NGS detected EGFR activating mutations in 81.3% and 71.9% of baseline cfDNA samples, respectively. In 19 patients treated with afatinib for ≥24 weeks, the number of EGFR mutant alleles detected in cfDNA by dPCR declined rapidly and markedly after treatment onset, becoming undetectable or detectable at only a low copy number (<10 copies per milliliter) at 4 weeks. Median PFS was slightly longer for patients with undetectable EGFR mutant alleles in cfDNA at 4 weeks than for those in whom such alleles were detectable (14.3 versus 10.0 months). A total of 45 somatic mutations was identified in baseline tumor DNA, and 30 (66.7%) of these mutations were identified in cfDNA by NGS. Allele frequency for somatic mutations in cfDNA determined by NGS changed concordantly during afatinib treatment with the number of EGFR mutant alleles determined by dPCR. Conclusions: Monitoring of cfDNA by dPCR is informative for prediction of afatinib efficacy, whereas that by NGS is reliable and has the potential to identify mechanisms of treatment resistance.

Development of an oncolytic HSV vector fully retargeted specifically to cellular EpCAM for virus entry and cell-to-cell spread.

Oncolytic herpes simplex virus (HSV) vectors have attracted increasing attention as novel anti-cancer agents. HSV entry is triggered by the binding of glycoprotein D (gD) to its receptors, such as herpesvirus entry mediator or nectin-1. We have recently reported the construction of a fully retargeted HSV platform that incorporates single-chain antibodies (scFv) into gD to mediate entry exclusively via tumor-associated antigens. In this study, we created an scFv directed against epithelial cell adhesion molecule (EpCAM), a recognized carcinoma-associated antigen, and inserted it into the retargeted HSV platform that is ablated for gD recognition of its canonical receptors and contains the entry-enhancing mutations in gB we previously identified. We observed that both initial entry and subsequent cell-to-cell spread of the retargeted virus were stringently dependent on cellular EpCAM expression. Interestingly, the retargeted virus developed larger plaques on some of the human tumor lines tested than the control virus bearing wild-type gD. Intratumoral injection of the retargeted virus revealed antitumor activity in a mouse xenograft model. These observations illustrate the versatility of our retargeted HSV platform as it allows expansion of the oncolytic virus toolbox for the treatment of diverse cancers.

Interferon-lambda4 genetic polymorphism is associated with the therapy response for hepatitis C virus recurrence after a living donor liver transplant.

The standard therapy against hepatitis C virus (HCV) recurrence postliver transplantation includes interferon (IFN)α and ribavirin. IFNL4 ss469415590 polymorphism has been reported as a novel predictor of the response to IFN therapy for chronic HCV infection. We examined the impact of IFNL4 polymorphism on the responsiveness to IFN therapy after liver transplantation. Tissue specimens were collected from 80 HCV-infected recipients and 78 liver donors, and their IFNL4 ss469415590 genotype, hepatic IFNL4 and interferon-stimulated genes' mRNA expression levels were examined. The association of the polymorphism and expression levels in terms of the IFN therapy response to HCV recurrence was analysed. Most individuals who had rs8099917 risk alleles also had ss469415590 risk alleles (R(2) = 0.9). Sustained virological response (SVR) rates were higher in both liver graft recipients and transplants with ss469415590 TT/TT alleles than in those with the risk ΔG allele (P = 0.003 and P = 0.005, respectively). In recipients with ss469415590 TT/TT, IFNL4 TT mRNA levels showed no significant differences between livers of patients who responded to therapy and those who did not (P = 0.4). In recipients with the risk ΔG allele, IFNL4 ΔG mRNA expression levels were significantly lower in SVR patients than in non-SVR patients (P = 0.02). Hepatic interferon stimulable genes and IFNL4 mRNA expression were correlated. Our findings suggest that analysing the ss469415590 genotype and IFNL4 ΔG expression provides a novel prediction strategy for the possible response to IFN therapy after liver transplantation.

First-line nivolumab + ipilimumab in advanced NSCLC: CheckMate 227 subpopulation analyses in Asian patients.

BACKGROUND: Nivolumab plus ipilimumab demonstrated clinically meaningful improvement in efficacy versus chemotherapy with a manageable safety profile in patients with advanced non-small cell lung cancer (NSCLC) and tumor programmed death-ligand 1 (PD-L1) expression ≥1% or <1% in Part 1 of CheckMate 227. Here we report efficacy and safety results for the Asian subpopulation. METHODS: Patients with stage IV/recurrent NSCLC were randomized 1 : 1 : 1 to nivolumab plus ipilimumab, nivolumab monotherapy, or chemotherapy (PD-L1 ≥1%) or nivolumab plus ipilimumab, nivolumab plus chemotherapy, or chemotherapy (PD-L1 <1%). Overall survival (OS), progression-free survival, objective response rate, duration of response, and safety were evaluated among patients in Japan, South Korea, and Taiwan. RESULTS: In the Asian subpopulation with PD-L1 ≥1%, 81 patients received nivolumab plus ipilimumab and 81 received chemotherapy. Median OS was not reached with nivolumab plus ipilimumab versus 24.8 months with chemotherapy; 3-year OS rate was 53% versus 37% [hazard ratio (HR), 0.72; 95% confidence interval (CI) 0.47-1.11]. The 3-year progression-free survival rate was 26% versus 7% (HR, 0.65; 95% CI 0.45-0.96), objective response rate was 56% versus 37%, and median duration of response was 29.0 months (95% CI 15.0 months-not reached) versus 6.9 months (95% CI 3.9-11.1 months). Similar results were observed regardless of tumor PD-L1 expression and in Japanese patients. Grade 3-4 treatment-related adverse events occurred in 40% of patients receiving nivolumab plus ipilimumab and 36% receiving chemotherapy, in the overall Asian subpopulation (tumor PD-L1 expression ≥1% and <1%); no new safety signals were identified. CONCLUSIONS: At 3-year follow-up, nivolumab plus ipilimumab provided durable long-term efficacy benefits versus chemotherapy regardless of tumor PD-L1 expression in the Asian subpopulation, including Japanese patients. Consistent with findings for all randomized patients, these data support the use of nivolumab plus ipilimumab as first-line treatment of Asian patients with advanced NSCLC.

The impact of IL28B genetic variants on recurrent hepatitis C in liver transplantation: significant lessons from a dual graft case.

IL28B genetic polymorphism is related to interferon-sensitivity in chronic hepatitis C, but the significance of grafts carrying different genotypes from recipients is still unclear in liver transplantation. A 51-year-old Japanese male carrying a minor genotype underwent dual liver transplantation for liver cirrhosis due to hepatitis C virus (HCV). The left lobe graft carried a major genotype, and the right a minor genotype. He achieved virological response during the course of pegylated-interferon and ribavirin therapy against recurrent hepatitis C for 2 years, but HCV relapsed immediately at the end of the therapy. Two years after antiviral therapy, liver biopsy was performed from each graft. The specimens showed A1F0 in the left lobe graft and A2F2 in the right. Moreover, quantitative polymerase chain reaction was performed using RNA extracted from each specimen to see there was no HCV RNA in the left lobe whereas there was in the right. This case provides clear evidence that IL28B genetic variants determine interferon sensitivity in recurrent hepatitis C following liver transplantation, which could result in new strategies for donor selection or for posttransplant antiviral therapy to HCV positive recipients.

Evaluation of pharyngeal swallowing pressure using high-resolution manometry during transoral surgery for oropharyngeal cancer.

BACKGROUND: Transoral robotic surgery is frequently described, driven by the desire to offer a less morbid alternative to chemoradiation. However, the objective evaluation of post-operative function has rarely been reported. Therefore, high-resolution manometry was used in this study to evaluate the impact of changes in peri-operative swallowing function on pharyngeal pressure events. METHODS: Ten patients with various stages of oropharyngeal cancer underwent transoral surgery. High-resolution manometry and videofluoroscopic swallow studies were performed before surgery and two months afterwards. The following parameters were obtained: velopharyngeal and mesopharyngeal post-deglutitive upper oesophageal sphincter pressures, velo-meso-hypopharyngeal contractile integral, upper oesophageal sphincter relaxation pressure, and pharyngeal velocity. RESULTS: There was no significant difference in pharyngeal pressure or contractile integral pre- versus post-operatively. However, pharyngeal velocity was significantly higher post-operatively than pre-operatively. CONCLUSION: High-resolution manometry showed that transoral surgery in patients without pre-operative dysphagia preserved pharyngeal constriction. However, transoral surgery might produce scar formation in the pharynx, which could lead to narrowing of the pharynx.

A phase II study of amrubicin combined with carboplatin for elderly patients with small-cell lung cancer: North Japan Lung Cancer Study Group Trial 0405.

BACKGROUND: Amrubicin, a new anthracycline agent, has shown high activity for small-cell lung cancer (SCLC) in previous studies. However, a combination regimen with amrubicin and platinum has been investigated little. On the basis of previous phase I study, we conducted this study to evaluate the efficacy and the safety of amrubicin and carboplatin for elderly patients with SCLC. METHODS: Chemotherapy-naive elderly patients with SCLC received amrubicin (35 mg/m(2), days 1-3) and carboplatin [area under the curve (AUC) 4.0, day1] every 3 weeks. The primary end point was overall response rate (ORR), and secondary end points were progression-free survival (PFS), overall survival and toxicity profile. RESULTS: From January 2005 to November 2007, 36 patients were enrolled [median age 76 (range 70-83); ECOG performance status of zero and one in 17 and 19 patients, respectively]. One complete response and 31 partial responses were observed (ORR 89%). Median PFS was 5.8 months and median survival time was 18.6 months. Grade 3-4 neutropenia was observed in 97% of the patients and six patients (17%) suffered from grade 3-4 febrile neutropenia. Other toxic effects were moderate and treatment-related death was not observed. CONCLUSIONS: Amrubicin combined with carboplatin is quite effective for SCLC with acceptable toxic effects even for the elderly population. Further evaluation of this regimen is warranted.

EVALUATIONS OF INVENTORY AND ACTIVITY CONCENTRATION OF RADIOCESIUM IN SOIL AT A RESIDENTIAL HOUSE 3 YEARS AFTER THE FUKUSHIMA NUCLEAR ACCIDENT.

Soil samples from the surface to a 5 cm depth were collected at a residential house in Koriyama City, Fukushima Prefecture using a scraper plate every three months from March 2014 to September 2014 to evaluate the vertical distribution profiles and inventories of 134Cs and 137Cs in soil. The vertical distribution profiles of radiocesium (134Cs and 137Cs) in soil showed that greater than 86% of the total radiocesium was absorbed in the upper 2 cm 3 years after the accident. Radiocesium in the surface layer seems to move to the lower layer over time. The migration of radiocesium in surface layer might be influenced by the ground surface runoff by rainfall. Radiocesium inventories in June increased significantly over the short period between March and June. In contrast, the radiocesium inventories in September did not increase significantly compared to the values in June. Radiocesium resuspension and deposition caused by decontamination work and meteorological events might be one possible reason for the increased radiocesium inventories observed in June.

FEASIBILITY STUDY ON THE FUSION OF PHITS SIMULATIONS AND THE DLNN ALGORITHM FOR A NEW QUANTITATIVE METHOD OF IN-SITU MULTIPLE-CHANNEL DEPTH DISTRIBUTION SPECTROMETRY.

We have recently have developed an in-situ multiple-channel depth distribution spectrometer (DDS) that can easily acquire on-site measurements of the depth distribution of specific radioactivities of Cs-134 and Cs-137 underground. Despite considerable improvements in the hardware developed for this device, the quantitative method for determining of radioactivities with this DDS device cannot yet achieve satisfactory performance for practical use. For example, this method cannot discriminate each γ-ray spectra of Cs-134 and Cs-137 acquired by the 20 thallium-doped caesium iodine CsI(Tl) scintillation crystal detectors of the DDS device from corresponding depth levels of underground soil. Therefore, we have applied deep learning neural network (DLNN) as a novel radiation measurement technique to discriminate the spectra and to determine the specific radioactivities of Cs-134 and Cs-137. We have developed model soil layers on a virtual space in Monte-Carlo based PHITS simulations and transported γ-ray radiation generated from a particular single soil layer or multiple layers as radiation sources; next, we performed PHITS calculations of those specific radioactivity measurements for each soil layer using DDS device based on machine learning via the DLNN algorithm. In this study, we obtained informative results regarding the feasibility of the proposal innovative radiation measurement method for further practical use in on-site applications.

The geomorphology, color, and thermal properties of Ryugu: Implications for parent-body processes.

The near-Earth carbonaceous asteroid 162173 Ryugu is thought to have been produced from a parent body that contained water ice and organic molecules. The Hayabusa2 spacecraft has obtained global multicolor images of Ryugu. Geomorphological features present include a circum-equatorial ridge, east-west dichotomy, high boulder abundances across the entire surface, and impact craters. Age estimates from the craters indicate a resurfacing age of [Formula: see text] years for the top 1-meter layer. Ryugu is among the darkest known bodies in the Solar System. The high abundance and spectral properties of boulders are consistent with moderately dehydrated materials, analogous to thermally metamorphosed meteorites found on Earth. The general uniformity in color across Ryugu's surface supports partial dehydration due to internal heating of the asteroid's parent body.

Efficacy of fluoroscopy-guided endoscopic cricopharyngeal myotomy.

BACKGROUND: In endoscopic cricopharyngeal myotomy, surgeons sometimes have concerns about performing an adequate incision with only a narrow intra-cavital view from one direction. In order to overcome these issues, fluoroscopic radiography was used during endoscopic cricopharyngeal myotomy. METHODS: Peri-operative fluoroscopic radiography was utilised to check the position of the diverticuloscope, and to confirm the extent of the incision during surgery. A balloon catheter was used to determine whether the cricopharyngeal muscle was sufficiently resected. Blood loss, peri-operative complications, and functional oral swallowing scale and penetration aspiration scale scores were evaluated. RESULTS: In 12 out of 15 patients, intra-operative fluoroscopic radiography showed the diverticuloscope positioned in the post-cricoid area, and the cricopharyngeal muscle was raised and the surgery completed without adverse effect. Swallowing functions improved following surgery. CONCLUSION: Intra-operative fluoroscopy might improve endoscopic cricopharyngeal myotomy by allowing surgeons to confirm the extent of resection, and by reducing peri-operative morbidity and complication rates.

Analysis of donor-type chimerism in lineage-specific cell populations after allogeneic myeloablative and non-myeloablative stem cell transplantation.

We analyzed donor-type chimerism in CD3+, CD14.15+ and CD56+ cells from 36 patients who had undergone conventional-intensity allogeneic stem cell transplantation (CST) and 34 patients who had undergone non-myeloablative allogeneic stem cell transplantation (NST) for hematological malignancies. On day 28 after transplantation, all fractions in NST patients and CD3+ cells in CST patients who received a non-total body irradiation (TBI) regimen showed more frequent mixed chimerism (<90% donor cells) than those in patients who had received TBI. NST patients with acute graft-versus-host disease (grade II-IV) frequently showed more than 50% donor-type chimerism in CD3+ cells on day 14 (P=0.029). NST patients with <50% donor-type chimerism on day 14 and with <90% donor-type chimerism on day 28 in CD56+ cells had significantly poor 1-year overall survival (0 vs 91%, P<0.001 and 20 vs 74%, P=0.002, respectively). Both NST and CST patients with <90% donor-type chimerism in CD14.15+ cells on day 28 had significantly poor 1-year overall survival (14 vs 70%, P=0.005 and 0 vs 66%, P=0.002, respectively). Our data show that the extent of donor-type chimerism in lineage-specific cells appears to have an impact on outcome after allogeneic stem cell transplantation.

Effect of pharmacological doses of estrogen on ovary-independent rat mammary carcinoma containing estrogen and progesterone receptors.

Mammary carcinoma induced in male Sprague-Dawley rats by multiple intragastric intubations of 7,12-dimethylbenz-(a)anthracene showed ovary-independent growth but contained estrogen receptors (ER) and progesterone receptors (Yoshida, Yoshida. Fukunishi, Sato, Okamoto, and Matsumoto, Cancer Res., 42: 2434-2439, 1982). Transplantable carcinoma (MT6) was obtained from dimethylbenz(a)anthracene-induced mammary carcinoma and then maintained in male rats. MT6 tumors with ER grew equally well in males, females, gonadectomized males, males given injections of bromocryptine (1 mg/day) or lisuride hydrogen maleate (50 micrograms/day), and gonadectomized males receiving smaller doses of 17 beta-estradiol (1 to 100 micrograms/2 days). However, the growth of MT6 was inhibited markedly by injection of a very large amount of 17 beta-estradiol (1 mg/2 days). Although transplanted MT6 tumors were ductal carcinoma with cribriform pattern with ER, tumors recurring after injection with 1 mg 17 beta-estradiol were found to be spindle cell carcinoma without ER, which could grow equally well in recipients treated with or without 1 mg 17 beta-estradiol. These observations suggest that the growth of ovary-independent MT6 tumors with ER and progesterone receptors is inhibited only by pharmacological doses of estrogens and that the loss of growth-inhibiting effect of pharmacological doses of estrogen of MT6 tumors occurs during high-dose estrogen treatment.

Different mechanisms of inhibition by alkyl-lysophospholipid and phorbol ester of granulocyte-macrophage colony-stimulating factor binding to human leukemic cell lines.

We investigated the effects of rac-1-O-octadecyl-2-O-methyl-glycero-3-phosphocholine (ET-18-OCH3) and 12-O-tetradecanoylphorbol-13-acetate (TPA) on granulocyte-macrophage colony-stimulating factor (GM-CSF) binding to human leukemic cell lines HL60, U937, KG-1, KG-1a, and K562. HL60, U937, and KG-1 exhibited the high-affinity receptors, but KG-1a and K562 revealed no demonstrable receptors. ET-18-OCH3 inhibited GM-CSF binding to HL60, U937, and KG-1 cells with a half-maximal inhibitory concentration of 16, 10, and 78 microM, respectively. ET-18-OCH3 at 10 microM reduced GM-CSF binding sites on HL60, U937, and KG-1, but had little effect on the dissociation constant (Kd). ET-18-OCH3 at 10 and 30 microM significantly (p < 0.01) decreased, in a dose-dependent manner, the total uptake, surface binding, and internalization of GM-CSF. The internalization of GM-CSF was more profoundly inhibited than its surface binding. TPA at 1 and 10 nM inhibited GM-CSF binding. Inhibition of GM-CSF binding by a combination of ET-18-OCH3 (10 microM) and TPA (1 or 10 nM) was less than additive, and ET-18-OCH3 partially inhibited TPA-induced protein kinase C (PKC) depletion in the cytosol and translocation to the particulate fractions. It is suggested that the inhibition of GM-CSF binding by ET-18-OCH3 is due in part to disruption of the plasma membrane and that the inhibition of GM-CSF binding by TPA is due to activation of PKC.

Collaboration of local government and experts responding to increase in environmental radiation level due to the nuclear disaster: focusing on their activities and latest radiological discussion.

Activities were introduced in Kashiwa city in the Tokyo metropolitan area to correspond to the elevated environmental radiation level after the disaster of the Fukushima Daiichi nuclear power plant. These were based on a strong cooperation between local governments and experts. Ambient dose rate and radioactivity of foodstuff produced inside of the city have been monitored. Representative ambient dose rates around living environments have almost already become their original levels of the pre-accident because of the decontamination activity, natural washout and effective half-lives of radioactivity. The internal annual dose due to radioactive cesium under the policy of 'Local Production for Local Consumption' is estimated as extremely low comparing the variation range due to natural radioactivity. Systematic survey around a retention basin has been started. All of these latest monitoring data would be one of the core information for the policy making as well as a cost-benefit discussion and risk communication.

Crystallization in Ising quantum magnets.

Dominating finite-range interactions in many-body systems can lead to intriguing self-ordered phases of matter. For quantum magnets, Ising models with power-law interactions are among the most elementary systems that support such phases. These models can be implemented by laser coupling ensembles of ultracold atoms to Rydberg states. Here, we report on the experimental preparation of crystalline ground states of such spin systems. We observe a magnetization staircase as a function of the system size and show directly the emergence of crystalline states with vanishing susceptibility. Our results demonstrate the precise control of Rydberg many-body systems and may enable future studies of phase transitions and quantum correlations in interacting quantum magnets.

Two patients treated with simeprevir plus pegylated-interferon and ribavirin triple therapy for recurrent hepatitis C after living donor liver transplantation: case report.

We previously reported our data on telaprevir (TVR) used in combination with pegylated-interferon and ribavirin (PEG-IFN/RBV) for the treatment of recurrent hepatitis C virus (HCV) genotype 1 infection after liver transplantation (LT). TVR substantially increases the blood levels of immunosuppressive agents such as cyclosporine and tacrolimus for drug-drug interactions. On the other hand, the effect of simeprevir (SMV) on the blood levels of these immunosuppressive agents is unclear. We report 2 patients who achieved viral responses with little effect on the blood levels of cyclosporine and tacrolimus using SMV plus PEG-IFN/RBV treatment. The first was a 71-year-old woman with HCV-related liver cirrhosis and hepatocellular carcinoma who failed to respond to PEG-IFN/RBV after living donor LT. She was treated with 40 mg/d of cyclosporine, and received SMV plus PEG-IFN/RBV treatment. The second was a 65-year-old man with HCV-related liver cirrhosis who failed to respond to PEG-IFN/RBV after living donor LT. He was treated with 3 mg/d of tacrolimus, and received SMV plus PEG-IFN/RBV treatment. Serum HCV RNA became undetectable using TaqMan polymerase chain reaction (PCR) test after 4 weeks of treatment in both patients, and no remarkable fluctuation in blood concentration was observed either in cyclosporine or tacrolimus during the 12 weeks of SMV treatment. Completion of 12-week SMV triple therapy was followed by PEG-IFNα2b plus RBV, and both patients achieved sustained virological response 12 weeks after the end of treatment. SMV plus PEG-IFNRBV treatment showed a remarkable viral response with little effect on blood levels of immunosuppressive agents for recurrent HCV genotype 1 infection after LT.

Cancer incidence in Hiroshima and Nagasaki, Japan, 1958-1987.

The Hiroshima and Nagasaki tumour registries, which have been in operation since 1958, are among the few population-based cancer registries in Japan. This analysis evaluated cancer incidence in Hiroshima and Nagasaki between 1958 and 1987. The overall age-adjusted (World Population Standard) cancer incidence has increased from 217 to 301 per 100,000 among males, and from 176 to 197 per 100,000 among females during the first 30 years of cancer registration. The most recent rates are intermediate to rates in other countries. Despite a gradual decrease, gastric cancer remained the most common malignancy among males and females throughout the surveillance period, accounting for 24% of all cancers by the late 1980s. The rate of liver cancer has increased dramatically among males during the past 20 years, with a 2-fold increase in incidence in the past 10 years alone. The populations of Hiroshima and Nagasaki now have among the highest rates of liver cancer in the world. Breast cancer incidence in Hiroshima and Nagasaki, in contrast, is among the lowest in the world, although incidence rates have doubled since the 1960s. Other common malignancies include cancers of the lung, colon and rectum among males and cancers of the colon, cervix and lung among females.

The activity of suppressor cells in the spleen of murine bone marrow chimeras.

An intrasplenic injection (i.s.) of BALB/c bone marrow cells induces a higher survival rate than an intravenous injection (i.v.) in irradiated C3H/He recipients. Coculture experiments revealed the presence of alloantigen-specific and nonspecific suppressor cells in the spleens of mice injected i.s. and i.v. Suppressor activity decreased 50-60 days after bone marrow transplantation in i.v. chimeras, while there was no decrease in i.s. chimeras. In vitro suppressor activity was correlated with in vivo activity. Histopathological changes in the liver were examined. A total of 28% of the i.v. chimeras showed severe changes compared with 6% of the i.s. chimeras. The spleen indices of i.v. and i.s. chimeras were compared. Although there was no statistically significant difference between the spleen indices of i.v. chimeras and those of i.s. chimeras, spleen indices of i.v. chimeras tended to be higher than those of i.s. chimeras. These results show that suppressor cells in i.s. chimeras appear to inhibit graft-versus-host reactions more efficiently. Furthermore, an adoptive transfer assay showed that suppressor cells detected in i.s. chimeras were effective in vivo. We therefore suggest that suppressor cells detected in vitro correlate with in vivo activity and may play some role in the induction and maintenance of transplantation tolerance.

Use of the polymerase chain reaction to detect hypermethylation in the calcitonin gene. A new, sensitive approach to monitor tumor cells in acute myelogenous leukemia.

Based on the recent observations that, in a majority of patients with acute leukemia, the 5' end of the calcitonin gene was hypermethylated and abnormal DNA fragments were observed following HpaII restriction digestion, we have developed a PCR-based method to sensitively detect this abnormal methylation of the calcitonin gene in AML. Applying the concept of competitive PCR, a semi-quantitative correlation was obtained between the amount of hypermethylation and the amount of leukemic cells present. These results suggest that this method will be useful to monitor the amount of tumor cells in bone marrow from patients with AML.