Oncolytic viruses and cancer treatment.

BACKGROUND: The fundamental difference between tumor and normal tissue growth is the emergence of the microenvironment with diminished or extinguished immunogenicity. One of the main functions of oncolytic viruses is the formation of such a microenvironment, which leads to a revival of immunological processes and loss of viability of cancer cells. Oncolytic viruses are being continuously improved and should be considered as a possible adjuvant immunomodulatory cancer treatment. A key requirement for the success of this cancer therapy is the specificity of the oncolytic viruses, which replicate only in tumor cells but do not affect normal cells. In this review, optimization strategies to achieve cancer specificity with increased efficacy are discussed and the most interesting results from preclinical and clinical trials are presented. PURPOSE: This review provides information on the current status of the development and use of oncolytic viruses as part of the bio-logical treatment of cancer.

[The substance of genetic information--nucleic acids]. / Hmotná podstata genetických informací - nukleové kyseliny.

If we look at the history of our knowledge of nucleic acids, we would see in the distant past of 140 years Friedrich Miescher who had identified the acidic substance within the cell nucleus, which he called nuclein. About 70 years after his initial observation, this substance was connected with genetic information. This very substantial finding happened during the World War II. This was the impulse that research of nucleic acids received to speed up continuously growing mountain of information, which is more and more difficult to understand. Another eruption of new information about our genome was the result of ten years of intensive cooperation of many manufacturers divided into two competitive blocks which offered us knowledge of nucleotide sequence of all 46 DNA molecules. The year 2000 became the landmark marking the start of the postgenomic era. It did not mean that human genome was totally explored, but the cornerstone has been settled. Since then, we could concentrate our efforts on variability; use of the project of 1,000 genomes brought many important findings, eg. copy number variability (CNV) exceeds the single nucleotide polymophisms (SNP). Also intergenomic relationships, studies on function and pathways began to be much more understandable by elucidation of the genome primary structure. NGS as a tool also accelerated the epigenetic research. All this improved molecular diagnostics by discovering many new markers playing their role in disease and treatment and allowed us to enter the field of multifactorial illnesses including cancer. The progress in diagnostic technologies which has happened during the last decade forced our research teams to include other professions - eg. bioinformatics.

Transcriptome alterations in maternal and fetal cells induced by tobacco smoke.

OBJECTIVES: Maternal smoking has a negative effect on all stages of pregnancy. Tobacco smoke-related defects are well established at the clinical level; however, less is known about molecular mechanisms underlying these pathologic conditions. We thus performed a comprehensive analysis of transcriptome alterations induced by smoking in maternal and fetal cells. STUDY DESIGN: Samples of peripheral blood (PB), placenta (PL), and cord blood (UCB) were obtained from pregnant smokers (n = 20) and gravidas without significant exposure to tobacco smoke (n = 52). Gene expression profiles were assayed by Illumina Expression Beadchip v3 for analysis of 24,526 transcripts. The quantile method was used for normalization. Differentially expressed genes were analyzed in the Limma package and the P-values were corrected for multiple testing. Unsupervised hierarchical clustering was performed using average linkage and Euclidean distance. The enrichment of deregulated genes in biological processes was analyzed in DAVID database. RESULTS: Comparative analyses defined significant deregulation of 193 genes in PB, 329 genes in PL, and 49 genes in UCB of smokers. The deregulated genes were mainly related to xenobiotic metabolism, oxidative stress, inflammation, immunity, hematopoiesis, and vascularization. Notably, functional annotation of the affected genes identified several deregulated pathways associated with autoimmune diseases in the newborns of smokers. CONCLUSIONS: The study demonstrated maternal smoking causes significant changes in transcriptome of placental and fetal cells that deregulate numerous biological processes important for growth and development of the fetus. An activation of fetal CYP genes showed a limited ability of the placenta to modulate toxic effects of maternal tobacco use.

HVRII of mtDNA in cord blood cells of newborn children and in their saliva 10 years later.

Comparison of hypervariable region II nucleotide sequences of mitochondrial DNA obtained from cord blood cells and saliva cells of the same individual at birth and after ten years revealed a few differences at the so-called mutation hot spots (three transitions and three indels within the C-tract). The personal identity of samples was proved by short tandem repeat profiling. Comparison of individuals living in two regions that differ by air pollution, however, did not reveal statistically significantly increased number of mutations in the population from the region of poorer environmental conditions, although indicating such tendency.

Effect of maternal tobacco smoke exposure on the placental transcriptome.

Smoking in pregnancy increases a woman's risk of preterm delivery resulting in serious neonatal health problems and chronic lifelong disabilities for the children (e.g., mental retardation, learning problems). To study the effects of tobacco smoke on the placental transcriptome, we performed gene expression profiling on placentas from women exposed to tobacco smoke in pregnancy (N = 12) and from those without significant exposure (N = 64). Gene expression profiles were determined by Illumina HumanRef-8 v2 Expression BeadChips with 18,216 gene probes. Microarray data were normalized by quantile method and filtered for a detection P-value <0.01. Differential gene expression was determined by moderated t-statistic. A linear model was fitted for each gene given a series of arrays using lmFit function. Multiple testing correction was performed using the Benjamini and Hochberg method. Abundant levels of transcripts were found for genes encoding placental hormones (CSH1, CSHL1), pregnancy-specific proteins (PSG3, PSG4, PAPPA), and hemoglobins (HBB, HBG, HBA). Comparative analysis of smokers vs nonsmokers revealed the differential expression of 241 genes (P < 0.05). In smoker cohort, we detected high up-regulation of xenobiotic genes (CYP1A1, CYP1B1, CYB5A, COX412), collagen genes (e.g., COL6A3, COL1A1, COL1A2), coagulation genes (F5, F13A1) as well as thrombosis-related genes (CD36, ADAMTS9, GAS6). In smokers, we identified deregulated genes that show tissue non-specific induction and may be considered as general biomarkers of tobacco smoke exposure. Further, we also found genes specifically deregulated in the exposed placentas. Functional annotation analysis suggested processes and pathways affected by tobacco smoke exposure that may represent molecular mechanisms of smoke-induced placental abnormalities.

[On human ageing and longevity--2. Internal determinants]. / O lidském stárnutí a dlouhovekosti--2. Vnitrní podmínky.

Longevity as a result of prolonged or postponed ageing is substantially influenced by genetic determinants. Most of them were analyzed by studying different progeria syndromes and their genetic control. From this point of view the use of experimental animals, because it enables to identify genes involved in shortening or prolongation of life-span by changing experimental conditions, has been also very effective. Expression of ageing processes at cellular as well on organism level as a biological unit revealed sets of genetic pathways involved in longevity. Several theories which elicit different aspects have been constructed to explain the ageing process.

[On human longevity--1. external influences]. / O lidské dlouhovekosti--1. vnejsí vlivy.

Human life-span is controlled by many factors, both internal (genomic) and external. In this paper only impact of external conditions is to be discussed. External factors are very variable, sometimes difficult to ascertain, but their control is relatively easier to address than the internal ones. The effect of human activities on human lifespan can be positive or negative. Experimental results have shown that one of the ways leading to prolongation of life-span is caloric restriction (CR). There are, of course, many other factors, not only of nurture character, inflicting on quality and length of human life. Some of them are human products including stress with different levels of inherited ability to resist it, but also possibility to improve individual capacity to control its effect due to hormesis. Prolongation of life expectancy--ageing of the population has become an economic burden in many countries, especially for social and health services.

[Clinical validity and clinical utility of molecular-genetic laboratory testing]. / Klinická validita a uzitecnost molekulárne genetických laboratorních zkousek.

Intensity of genetic determination of our phenotypes differs both quantitatively and qualitatively and concerns not only physiological, but also pathological features. Clinical diagnoses are more and more dependent on laboratory findings, whose quality and utility have to be objectively estimated and assesed. For medical practice the knowledge of genetic determination is substantial and for this reason several approaches have beeen adopted. Genetic determination can be measured and expressed by several parameters, of which heritability and genetic risk are discussed in this paper to elucidate their properties, importace, significance, and limitations.

[Direct-to-consumer genetic testing also in our country]. / Genetické testování zpusobem DTC i v nasich krajích.

The main features of direct-to-consumer (DTC) genetic testing suffers from the loss of personal contact between a client and a genetic consultant, who is apt to assess the usefulness of genetic testing and to interpret results of genetic tests and who is also sufficiently trained to provide professional way of consultation. Laboratories offering these services often inform clients by phone, and neglect their identification - the samples examined may be anonymous. Also samples from children are accepted for testing. Quality control is not mandatory and laboratory genetic testing is not a licenced business in the Czech Republic. Because of these problems, which are not specific to our situation, but rather global, the possibility of the ban concerning these activities is being discussed in several European countries.

Genetic variability of HVRII mtDNA in cord blood and respiratory morbidity in children.

Genetic polymorphisms were examined using direct sequencing of the hypervariable region II (HVRII) in the D-loop of mtDNA in the cord blood of 355 children living in two areas of the Czech Republic - the industrial district of Teplice and the agricultural district of Prachatice. The incidence of the most frequent nucleotide variants of HVRII, C150T (10.1%), T152C (19.7%), T195C (19.7%) and 309.nC (41.4% for 309.2C and 13.8% for 309.3C), and the respiratory morbidity at the ages of 0-2 years and 2-6 years were investigated, considering many other factors such as locality, gender, ethnicity, heating by coal in household, maternal age, asthma bronchiale, allergic rhinitis, pollinosis, conjunctivitis and maternal tobacco exposure during and after pregnancy. We found that the T195C transversion in HVRII is connected with an increased risk of early childhood (0-2 years) bronchitis (RR 1.38, p=0.034, 95% CI 1.04-1.85) and with increased risk of otitis media in children aged 2-6 years (RR 1.62, p=0.032, 95% CI 1.04-2.53). Another polymorphism, 309.nC, is associated with an increased risk of bronchitis in children aged 2-6 years (RR 1.46, p=0.030, 95% CI 1.04-2.06). The results indicate that genetic polymorphisms in mtDNA may be an important factor not only for various types of cancers and neurodegenerative diseases, but also for respiratory morbidity in children.

[Personalized medicine--genomic phase]. / Personalizovaná medicína--genomická fáze. Proc a zac.

Personalized medicine has become declared to be a strategic goal of nearly all medical disciplines, a modern fashion of medicine, which corresponds to unexpected progress in knowledge of structure and sometimes also understanding the function of our genome. This was made possible by development of new technologies and methodological approaches, which led to improved and more detailed diagnostics and categorization of diseases. Also pharmaceutical industry is producing many new drugs, but not all are effective as supposed, and sometimes also cause adverse reactions (ADR). To prevent uneffectiveness and ADR, preliminary genetic testing of patients seems to be necessary. Utility of personalized medicine is sometimes discussed from the point of view of supposed inevitable increase of cost and problems with its adoption by existing system of healthcare.

JNK2 and p38 MAPK over-expressions do not represent key events in chronic myeloid leukemia transformation.

Over-expression of two members of MAP kinase family (JNK2 and p38) has been already observed in chronic myeloid leukemia (CML). In the present study, significance of this deregulation was investigated. Impacts of JNK2/p38 suppression on gene expression profile of CML cell lines (K562/KU-812) were studied using an experimental approach that combines siRNA-mediated specific inhibition of the genes and array-based expression analyses. After JNK2 depletion, 27 out of 588 tested genes showed significant expression changes, with 13 down-regulated genes and 14 up-regulated genes. Among others, expression of MSH2 and MSH6, mdm2, and caspase-2 was reduced and, on the other hand, MKK1 and MKK6, RFC2, cytokeratins K18 and K19, BAD, and DR5 expression was up-regulated. In the case of p38 silencing, 20 genes were considered as significantly deregulated (7 genes reduced, 13 over-expressed). These genes included caspase-10, SOD1, and Notch4 (down-regulation) and caspase-2 and caspase-3, CDC2, CDK4, and c-kit (up-regulation). In conclusion, comparison of expression profiles after JNK2 or p38 gene silencing revealed distinct sets of affected genes. The results implied an unequal impact of the MAPK deregulation on the CML cells. Further, we demonstrated that neither JNK2 nor p38 siRNAmediated inhibition led to significant change of CML cell proliferation. It suggests that there are other important, likely upstream regulators essential for CML malignant cell growth/transformation; therefore, separate inhibition of JNK2 or p38 MAPK gene is not sufficient for a proliferation arrest.

[Homage to McKusick's "Mendelian inheritance in man"]. / Hold McKusickove "Mendelian inheritance in man".

McKusick's database MIM has grown since its early beginning in sixties to 1985 when the online version (OMIM) appeared. The last edition of three volumes was printed in 1998. It has become a very valuable tool for all geneticists, and also clinicians of other disciplines started using it as a source of important information. The original limitation to disorders with mendelian inheritance has been step by step broken down, all components of human genome and also genes without known function and their epigenetic changes have been included. It was a pleasure for all of us to congratulate to McKusick's honorary degree obtained this year by the oldest European university in Bologna (with a short biography).

A bidirectional corridor in the Sahel-Sudan belt and the distinctive features of the Chad Basin populations: a history revealed by the mitochondrial DNA genome.

The Chad Basin was sparsely inhabited during the Stone Age, and its continual settlement began with the Holocene. The role played by Lake Chad in the history and migration patterns of Africa is still unclear. We studied the mitochondrial DNA (mtDNA) variability in 448 individuals from 12 ethnically and/or economically (agricultural/pastoral) different populations from Cameroon, Chad, Niger and Nigeria. The data indicate the importance of this region as a corridor connecting East and West Africa; however, this bidirectional flow of people in the Sahel-Sudan Belt did not erase features peculiar to the original Chad Basin populations. A new sub-clade, L3f2, is described, which together with L3e5 is most probably autochthonous in the Chad Basin. The phylogeography of these two sub-haplogroups seems to indicate prehistoric expansion events in the Chad Basin around 28,950 and 11,400 Y.B.P., respectively. The distribution of L3f2 is virtually restricted to the Chad Basin alone, and in particular to Chadic speaking populations, while L3e5 shows evidence for diffusion into North Africa at about 7,100 Y.B.P. The absence of L3f2 and L3e5 in African-Americans, and the limited number of L-haplotypes shared between the Chad Basin populations and African-Americans, indicate the low contribution of the Chad region to the Atlantic slave trade.

Y-chromosomal variation in the Czech Republic.

To analyze the contribution of the Czech population to the Y-chromosome diversity landscape of Europe and to reconstruct past demographic events, we typed 257 males from five locations for 21 UEPs. Moreover, 141 carriers of the three most common haplogroups were typed for 10 microsatellites and coalescent analyses applied. Sixteen Hg's characterized by derived alleles were identified, the most common being R1a-SRY(10831) and P-DYS257*(xR1a). The pool of haplogroups within I-M170 represented the third most common clade. Overall, the degree of population structure was low. The ages for Hg I-M170, P-DYS257*(xR1a), and R1a-SRY(10831) ap peared to be comparable and compatible with their presence during or soon after the LGM. A signal of population growth beginning in the first millennium B.C. was detected. Its similarity among the three most common Hg's indicated that growth was characteristic for a gene pool that already contained all of them. The Czech population appears to be influenced, to a very moderate extent, by genetic inputs from outside Europe in the post-Neolithic and historical times. Population growth postdated the archaeologically documented introduction of Neolithic technology and the estimated central value coincides with a period of repeated changes driven by the development of metal technologies and the associated social and trade organization.

[What is the future of our genome?]. / Kam smeruje nase genetická výbava?

Where do we go and what will happen with our genomes? Viewed as a reflection of what we know about our origin as Homo sapiens, history and forecast. Archeogenetics tries to find our sourcing place and to explain how humans occupied the globe. One of the first theories based on mtDNA polymorphisms@ studies stressed on our African origin followed by stepwise dispersal all over the world which took approximately 80,000 years. But not all findings agreed with this assumption if based on nuclear loci and supported multifocal origin of humans. The process of establishing a new species--Homo sapiens, is still not fully understood and many questions remained unanswered. From the point of view of population genetics we can assume that: 1. Mutability (natural or from internal causes) does not change though we cannot neglect suspicion that environment could influence its increase. The content of harming mutations in our genome, due to the protective effect of health care, which blocks natural selection, is increasing and moreover changes of our life style opened the door for manifestation of week deleterious alleles accumulated during foregoing period of evolution. Also prolongation of our life span is accompanied with effects of genotypes positively selected because of their positive effect on our reproductive period but which could be harmful during postreproductive stage--antagonistic pleiotropy. 2. According traditional assumption on the quality of new mutations is that they are either neutral or harming. Changes which are drift-dependent are becoming reduced. 3. Effective population size (steady state could be supposed or some increase due to more intensive local migration). 4. Migration (In spite of absence of corresponding demographic data) seems to be nowadays more intensive than it was in the past.

MtDNA of Fulani nomads and their genetic relationships to neighboring sedentary populations.

Despite the large size of the contemporary nomadic Fulani population (roughly 13 million people), the genetic diversity and degree of differentiation of Fulanis compared to other sub-Saharan populations remain unknown. We sampled four Fulani nomad populations (n = 186) in three countries of sub-Saharan Africa (Chad, Cameroon, and Burkina Faso) and analyzed sequences of the first hypervariable segment of the mitochondrial DNA. Most of the haplotypes belong to haplogroups of West African origin, such as L1b, L3b, L3d, L2b, L2c, and L2d (79.6% in total), which are all well represented in each of the four geographically separated samples. The haplogroups of Western Eurasian origin, such as J1b, U5, H, and V, were also detected but in rather low frequencies (8.1% in total). As in African hunter-gatherers (Pygmies and Khoisan) and some populations from central Tunisia (Kesra and Zriba), three of the Fulani nomad samples do not reveal significant negative values of Fu's selective neutrality test. The multidimensional scaling of FST genetic distances of related sub-Saharan populations and the analysis of molecular variance (AMOVA) show clear and close relationships between all pairs of the four Fulani nomad samples, irrespective of their geographic origin. The only group of nomadic Fulani that manifests some similarities with geographically related agricultural populations (from Guinea-Bissau and Nigeria) comes from Tcheboua in northern Cameroon.