Dynamic Comparison of Kaplan–Meier Proportions: Monitoring a Randomized Clinical Trial with a Long-Term Binary Endpoint

Summary .   The approach to early termination for efficacy in a trial where events occur over time but the primary question of interest relates to a long-term binary endpoint is not straightforward. This article considers comparison of treatment groups with Kaplan–Meier (KM) proportions evaluated at increasing times from randomization, at increasing calendar testing times. This strategy is employed to improve the ability to detect important treatment effects and provide critical treatments to patients in a timely manner. This dynamic Kaplan–Meier (DKM) approach is shown to be robust; that is, it produces high power and early termination time across a wide range of circumstances. In contrast, a fixed time KM comparison and the log-rank test are both shown to be more variable in performance. Practical considerations of implementing the DKM method are discussed.     

A regression-based method for estimating mean treatment cost in the presence of right-censoring

In many clinical trials and evaluations using medical care administrative databases it is of interest to estimate not only the survival time of a given treatment modality but also the total associated cost. The most widely used estimator for data subject to censoring is the Kaplan-Meier (KM) or product-limit (PL) estimator. The optimality properties of this estimator applied to time-to-event data (consistency, etc.) under the assumptions of random censorship have been established. However, whenever the relationship between cost and survival time includes an error term to account for random differences among patients' costs, the dependency between cumulative treatment cost at the time of censoring and at the survival time results in KM giving biased estimates. A similar phenomenon has previously been noted in the context of estimating quality-adjusted survival time. We propose an estimator for mean cost which exploits the underlying relationship between total treatment cost and survival time. The proposed method utilizes either parametric or nonparametric regression to estimate this relationship and is consistent when this relationship is consistently estimated. We then present simulation results which illustrate the gain in finite-sample efficiency when compared with another recently proposed estimator. The methods are then applied to the estimation of mean cost for two studies where right-censoring was present. The first is the heart failure clinical trial Studies of Left Ventricular Dysfunction (SOLVD). The second is a Health Maintenance Organization (HMO) database study of the cost of ulcer treatment.     

Thermosensitive Replication of a Kanamycin Resistance Factor

A strain of Proteus vulgaris isolated from the urinary tract of a patient with postoperative pyelonephritis and resistant to sulfonamide, streptomycin, tetracycline, and kanamycin (KM) was found to transfer only KM resistance by cell-to-cell conjugation. The genetic determinant controlling the transferable KM resistance was considered to be an R factor and was designated R (KM). Successive transfer of KM resistance was demonstrated also from Escherichia coli 20S0, which received the R (KM) factor, to other substrains of E. coli K-12 or Salmonella typhimurium LT-2. The transfer of the R (KM) factor was strongly affected by the temperature at which the mating culture was kept. The transfer frequency of R (KM) at 25 C was about 10(5) times higher than at 37 C. The R (KM) factor was spontaneously eliminated from the host bacterial cells when P. vulgaris was cultured at 42 C, but no elimination occurred at 25 C. This elimination of the R (KM) factor at elevated temperature was also observed when the R (KM) factor infected E. coli and S. typhimurium. On the other hand, a normal R factor could not be eliminated from the same E. coli host strain by cultivation at the higher temperature. We consider the thermosensitive transfer and the spontaneous elimination of the R (KM) factor at higher temperature to depend upon thermosensitive replication of the R (KM) factor.     

The dual klepsydra model of internal time representation and time reproduction

We present a model of the internal representation and reproduction of temporal durations, the 'dual klepsydra' model (DKM). Unlike most contemporary models operating on a 'pacemaker-counter' scheme, the DKM does not assume an oscillatory process as the internal time-base. It is based on irreversible, dissipative processes in inflow/outflow systems (leaky klepsydrae), whose states are continuously compared; if their states are equal, durations are subjectively perceived as equal. Model-based predictions fit experimental time reproduction data with good accuracy, and show qualitative features not accounted for by other models. The deterministic model is characterized by two parameters, kappa (outflow rate coefficient) and eta (ratio of inflow rates). A stochastic version of the model (SDKM) assumes randomly fluctuating inflows, involves two more parameters, and accounts for intra-individual variance of reproduced durations. Analysis of the SDKM leads to non-trivial problems in the stochastic theory, briefly sketched here. Methods of parameter estimation for both deterministic and stochastic versions are given. Applying the DKM to the subjective experience of time passage, we show how subjective measure of elapsed time is constituted. Finally, essential features of the model and its possible neurophysiological interpretation are discussed.     

Suppression of premature termination codons as a therapeutic approach

In this review, we describe our current understanding of translation termination and pharmacological agents that influence the accuracy of this process. A number of drugs have been identified that induce suppression of translation termination at in-frame premature termination codons (PTCs; also known as nonsense mutations) in mammalian cells. We discuss efforts to utilize these drugs to suppress disease-causing PTCs that result in the loss of protein expression and function. In-frame PTCs represent a genotypic subset of mutations that make up ~11% of all known mutations that cause genetic diseases, and millions of patients have diseases attributable to PTCs. Current approaches aimed at reducing the efficiency of translation termination at PTCs (referred to as PTC suppression therapy) have the goal of alleviating the phenotypic consequences of a wide range of genetic diseases. Suppression therapy is currently in clinical trials for treatment of several genetic diseases caused by PTCs, and preliminary results suggest that some patients have shown clinical improvements. While current progress is promising, we discuss various approaches that may further enhance the efficiency of this novel therapeutic approach.     

Suppression of premature termination codons as a therapeutic approach

In this review, we describe our current understanding of translation termination and pharmacological agents that influence the accuracy of this process. A number of drugs have been identified that induce suppression of translation termination at in-frame premature termination codons (PTCs; also known as nonsense mutations) in mammalian cells. We discuss efforts to utilize these drugs to suppress disease-causing PTCs that result in the loss of protein expression and function. In-frame PTCs represent a genotypic subset of mutations that make up ~11% of all known mutations that cause genetic diseases, and millions of patients have diseases attributable to PTCs. Current approaches aimed at reducing the efficiency of translation termination at PTCs (referred to as PTC suppression therapy) have the goal of alleviating the phenotypic consequences of a wide range of genetic diseases. Suppression therapy is currently in clinical trials for treatment of several genetic diseases caused by PTCs, and preliminary results suggest that some patients have shown clinical improvements. While current progress is promising, we discuss various approaches that may further enhance the efficiency of this novel therapeutic approach.     

中国昆明小鼠亚群蛋白质多态性的研究

本文报道采用电泳技术对北京、上海、长春3市的4个中国昆明小鼠(简称KM)实验群体中24个蛋白质标志研究的结果,与我国1981年从美国引进的NIH小鼠进行比较,显示出:(1)KM小鼠亚群间等位基因组成无明显差异,它们间遗传距离为0.008—0.027,与群体封闭时间成正相关;(2)4个KM小鼠群体间聚类分析发现S:KM群体为特殊一类,该结果与KM亚群间下颌骨分析结果相符;(3)KM与Swiss来源的NIH小鼠群体间在E(?)-3、Es-10、Got-2、Glo-1、Gpt-1、和Mpi-1座位上的等位基因组成存在显著差异,它们之间的遗传距离平均值为0.131±0.011,证实中国KM小鼠为非Swiss来源的一个亚种。     

低温和光周期对绿盲蝽越冬卵滞育解除和发育历期的影响

为了探讨温度和光周期对绿盲蝽Apolygus lucorum Meyer-Dür越冬卵滞育解除和发育历期的影响, 系统调查了绿盲蝽越冬卵在不同温度和不同光照组合下的孵化率和孵化时间, 结果显示：绿盲蝽的越冬卵均为滞育卵,低温和光周期对绿盲蝽越冬卵的滞育解除均有影响。2℃的低温处理能够显著促进其滞育解除,在0~65 d范围内,随着低温处理时间增长,其滞育解除时间缩短,未经低温处理的越冬卵T₅₀为68.5 d,低温处理65 d的T₅₀为12.25 d,绿盲蝽越冬卵在2℃低温处理65 d后完全解除滞育;在0~40 d范围内,低温处理时间越长,绿盲蝽越冬卵的孵化率越高,在25℃、全光照的条件下不经低温处理的孵化率为68.65%,低温处理40 d后在25℃的条件下的孵化率达到99.46%。在20~26℃范围内,绿盲蝽越冬卵的滞育后发育历期随着温度的上升而缩短, 随着光周期的延长而缩短。结果说明低温处理能够提高绿盲蝽越冬卵滞育解除率,但不是其滞育解除的必要条件,低温处理与自然变温对绿盲蝽滞育解除的作用相似;高温和长光照能够促进绿盲蝽的滞育解除,缩短发育历期。     

Pregnancy termination

During pregnancy, the antiprogestin mifepristone will induce uterine contractions, increase the sensitivity of the myometrium to prostaglandin, and ripen the cervix. These effects indicate that mifepristone can be used for termination of pregnancy. The clinical experience has shown that mifepristone is sufficiently effective for this purpose only if combined with a suitable prostaglandin, e.g. gemeprost or misoprostol. The combined treatment has been used for termination of early pregnancy (up to 63 days of amenorrhea) and for termination of second trimester pregnancy. During early pregnancy, the recommended dose of mifepristone is 600 mg (although 200 mg seems sufficient), followed 36-48 h later by 0.4-0.8 mg misoprostol administered either orally or vaginally, or vaginal administration of 1.0 mg gemeprost. For termination of second trimester pregnancy, the treatment with mifepristone is most commonly combined with 1.0 mg gemeprost repeated at 3-6-h intervals. The combined treatment is as effective and safe during early pregnancy as is the alternative vacuum aspiration and is also equally acceptable if the woman is allowed to choose the method she prefers. During the second trimester, the pretreatment will significantly reduce the duration of labor, dose of prostaglandin, and the frequency of side effects.     

An intravascular chemoattractant lectin inhibits neutrophil migration

KM+, a lectin purified from Artocarpus integrifolia seeds, is an attractant for neutrophils, and has properties similar to fMLP, IL-8 and MNCF. The endogenous lectin MNCF, inhibits carrageenan-induced neutrophil migration when intravenously administered in rats. In an attempt to mimic the activity of MNCF with KM+, we determined the effect of intravenous (iv) injection of KM+ (5 g) on neutrophil migration to the peritoneal cavity of Wistar rats induced by KM+ (50 g, intraperitoneal, ip), fMLP (5 ng, ip) and carrageenan (300 g, ip). Initially we evaluated the effect of the time interval between intravenous and intraperitoneal administration of KM+. The intervals ranged from 20 to 120 min and progressively stronger inhibition was observed with increasing time intervals up to a maximum of 60 min, with effect decreasing thereafter. With injections at the optimum interval of 60 min, we observed that KM+ inhibited KM+- and carrageenan-induced neutrophil migration by 72%, and fMLP-induced migration by 56%. White cell counts for Wistar rats that only received KM+iv, performed at 0 to 120 min intervals after injection, revealed early neutropenia lasting 60 min, followed by a marked increase in circulating neutrophils that reached a maximum of twice the initial levels within 90 min and after 120 min returned to levels near to that observed before intravenous administration of KM+. These results indicate that when KM+ is present in the intravascular space, it produces an inhibitory effect on neutrophil migration similar to that caused by the intravenous administration of other chemoattractants, regardless of whether they act through a mechanism independent of carbohydrate recognition, as does IL-8, or are dependent on carbohydrate recognition, like MNCF.     

Estimating number of events from the Kaplan-Meier curve for incorporation in a literature-based meta-analysis: what you don't see you can't get!

In literature-based meta-analyses of time-to-event data, the number of events in the treated and control groups together with the total number of patients randomized to the two treatment arms are often used as summary statistics. If interest is in mortality at a specified moment in time, the number of events can, in most cases, only be obtained from the Kaplan-Meier curve. The estimated number of events, however, is typically larger than the true number of events. The effect of this overestimation on the Mantel-Haenszel test and the odds ratio is studied in this paper. From these results, it can be concluded that the number of events should not be estimated from the Kaplan-Meier curves for meta-analytic purposes unless virtually no patients are lost to follow-up or censored and there are still many patients at risk in the two groups at the time at which the number of events is to be determined.     

Addressing an Idiosyncrasy in Estimating Survival Curves Using Double Sampling in the Presence of Self-Selected Right Censoring

We investigate the use of follow-up samples of individuals to estimate survival curves from studies that are subject to right censoring from two sources: (i) early termination of the study, namely, administrative censoring, or (ii) censoring due to lost data prior to administrative censoring, so-called dropout. We assume that, for the full cohort of individuals, administrative censoring times are independent of the subjects' inherent characteristics, including survival time. To address the loss to censoring due to dropout, which we allow to be possibly selective, we consider an intensive second phase of the study where a representative sample of the originally lost subjects is subsequently followed and their data recorded. As with double-sampling designs in survey methodology, the objective is to provide data on a representative subset of the dropouts. Despite assumed full response from the follow-up sample, we show that, in general in our setting, administrative censoring times are not independent of survival times within the two subgroups, nondropouts and sampled dropouts. As a result, the stratified Kaplan-Meier estimator is not appropriate for the cohort survival curve. Moreover, using the concept of potential outcomes, as opposed to observed outcomes, and thereby explicitly formulating the problem as a missing data problem, reveals and addresses these complications. We present an estimation method based on the likelihood of an easily observed subset of the data and study its properties analytically for large samples. We evaluate our method in a realistic situation by simulating data that match published margins on survival and dropout from an actual hip-replacement study. Limitations and extensions of our design and analytic method are discussed.     

Differential changes in nucleolar size and ribosomal RNA synthesis during diapause break by prolonged chilling in Bombyx eggs

The activity of ribosomal RNA (rRNA) genes as judged by nucleolar size and rRNA synthesis has been shown to depend upon the phase of diapause in the eggs of Bombyx mori. In the present study, we found that nucleolar size in diapausing eggs was enlarged at a very early stage during cold treatment, a procedure necessary for the termination of diapause. In contrast, the intrinsic capacity of ribosomal RNA synthesis in the chilled eggs, as examined at 25°C by radioactive precursor incorporation into rRNA, increased much later, in parallel with the break of diapause. The early phase of cold treatment is the period when the eggs undergo some important changes (the so-called diapause development), preparing for diapause termination. Thus we infer that the above mentioned increase in nucleolar size may be one of the features of diapause development.     

An enhanced mass spectrometry approach reveals human embryonic stem cell growth factors in culture

The derivation and long-term maintenance of human embryonic stem cells (hESCs) has been established in culture formats that are both dependent and independent of support (feeder) cells. However, the factors responsible for preserving the viability of hESCs in a nascent state remain unknown. We describe a mass spectrometry-based method for probing the secretome of the hESC culture microenvironment to identify potential regulating protein factors that are in low abundance. Individual samples were analyzed several times, using successive mass (m/z) and retention time-directed exclusion, without sampling the same peptide ion twice. This iterative exclusion -mass spectrometry (IE-MS) approach more than doubled protein and peptide metrics in comparison to a simple repeat analysis method on the same instrument, even after extensive sample pre-fractionation. Furthermore, implementation of the IE-MS approach was shown to enhance the performance of an older quadrupole time of flight (Q-ToF) MS. The resulting number of identified peptides approached that of a parallel repeat analysis on a newer LTQ-Orbitrap MS. The combination of the results of both instruments proved to be superior to that achieved by a single instrument in the identification of additional proteins. Using the IE-MS strategy, combined with complementary gel- and solution-based fractionation methods, the hESC culture microenvironment was extensively probed. Over 10 to 12 times more extracellular proteins were observed compared with previously published surveys. The detection of previously undetectable growth factors, present at concentrations ranging from 10(-9) to 10(-11) g/ml, highlights the depth of our profiling. The IE-MS approach provides a simple and reliable technique that greatly enhances instrument performance by increasing the effective depth of MS-based proteomic profiling. This approach should be widely applicable to any LC-MS/MS instrument platform or biological system.     

Growth characteristics of CHO cells in culture

The dependence of the growth characteristics and monolayer formation on the initial cell plating concentration were studied on a permanent CHO cell line. The cells were cultivated under standard conditions on plastic substrate. Initial plating concentrations were varied as 1000, 2000, 3000, 4000, 5000, and 6000 cells/cm². It was shown that the cell growth can be formally described by a standard S-shaped dependence. However, a more detailed analysis revealed inconsistency of the experimental and expected data. Specifically, the cell growth termination produced by monolayer formation does not coincide with the time when the theoretical curves approach a plateau. It is concluded that cell proliferation and monolayer formation are independent processes (at least in CHO cells). Both processes may be considered as analogs of proliferation and morphogenesis in metazoa. In addition, it is shown that the cessation of cell division is induced by reduction in the cell size to some limiting dimension and increasing of the cell polarization rather than contact inhibition of proliferation after the monolayer formation.     

Efficacy of a complex program for controlling influenza in a large industrial city

The epidemiological and economic effectiveness of the realization of the complex three-year (1980-1982) program for the control of influenza in Perm is shown. The coverage of 46-51% of the city population, including working people, pensioners and children, with immunization carried out with the use of live and inactivated vaccines made it possible to decrease morbidity rate 2.12 times in comparison with the average data for many years. The greatest effect was achieved at large industrial enterprises where, simultaneously with vaccination covering 90% of the employees, urgent prophylaxis with remantadin and the early treatment of influenza patients were carried out. Due to these measures the morbidity rate and the number of disability days per 100 employees decreased 3-6 times.     

Rheology of protein gels synthesized through a combined enzymatic and heat treatment method

Whey protein gels prepared under acidic conditions (pH<4.6) remain largely unutilized because of their weak and brittle nature in contrast to the favorable elastic gels produced at neutral or basic conditions. However, such usage is important, as low pH food products are desirable due to their shelf stability and less stringent sterilization processes. In this study, we use a two-step process involving enzyme followed by heat treatment to produce whey protein gels at low pH (4.0). Dynamic rheological measurements reveal that the gel elastic modulus and yield stress increase substantially when heat treatment is supplemented with enzyme treatment. Both the elastic modulus and yield stress increase with increasing enzyme concentration or treatment time. In contrast, the dynamic yield strain decreases with enzyme concentration but increases with time of enzyme treatment. These results are explained in terms of the enzyme treatment time affecting the diffusion of the enzyme within the gel. This in turn leads to two types of gel microstructure at short and long enzyme treatment times, with the extent of enzyme diffusion modulating the structure at intermediate times.     

Schistosoma mansoni: a diagnostic approach to detect acute schistosomiasis infection in a murine model by PCR

Schistosomiasis represents an increasing problem in non-endemic areas, due to the growing number of immigrants and to tourists contracting this disease in "off-the-beaten-track" tourism. Acute schistosomiasis is not diagnosed early due to the lack of diagnostic tools that are sufficiently sensitive enough to detect the parasite during the first weeks of infection. We have developed a diagnostic approach based on the detection of parasite DNA by polymerase chain reaction (PCR) in urine, comparing the performance of this new approach with the two currently used schistosomiasis diagnostic tools (Kato-Katz and ELISA) and the PCR in stool samples. This comparison was done in a Schistosoma mansoni murine experimental model, which permits follow up of the parasite from the acute to the chronic stage of infection. Our results suggest that this new PCR-based approach could be useful for the detection of acute schistosomiasis in easy-to-handle clinical samples such the urine.