Acceptance of physical activity monitoring in cancer patients during radiotherapy, the GIROfit phase 2 pilot trial.

Background: In radiotherapy the timely identification of patients needing intervention and supportive care due to side effects is an important task especially in the outpatient setting. Activity trackers as an increasingly used lifestyle device may enable physicians to monitor patient's physical activity (PA) and to intervene early during the course of radiotherapy. Objective: The primary aim of this trial was to assess patient acceptance of PA monitoring in an outpatient setting and to correlate changes in PA with toxicity and changes in quality of life. Methods: Patients undergoing radio(chemo-)therapy with a curative intent were eligible to participate in this prospective pilot phase II trial. Patients were instructed to wear a commercially available activity tracker during the course of radiotherapy and four weeks afterwards. Quality of life (QoL) and fatigue was scored using the Functional assessment of Chronic Illness Therapy questionnaire. A linear regression was performed to determine baseline activity and changes in step counts during radiotherapy. Results: We included 23 patients in this trial. Two withdrew consent before the start of treatment, two patients were excluded after prophylactic feeding tube placement and prolonged recovery. Compliance in the remaining 19 patients was high, with availability of step-counts on 92% of the days. Baseline step counts were 6274 for breast cancer patients and 3621 for patients with other entities. Decreasing activity during radiotherapy coincided with the development of side effects and declines in quality of life. Conclusions: Activity trackers as tool to monitor PA during and after radiotherapy were accepted by a majority of the patients included in the current trial. Observed changes in PA correlated with patient reported side effects and QoL in some of the patients.

Auger-spectroscopy in quantum Hall edge channels and the missing energy problem.

Quantum Hall edge channels offer an efficient and controllable platform to study quantum transport in one dimension. Such channels are a prospective tool for the efficient transfer of quantum information at the nanoscale, and play a vital role in exposing intriguing physics. Electric current along the edge carries energy and heat leading to inelastic scattering, which may impede coherent transport. Several experiments attempting to probe the concomitant energy redistribution along the edge reported energy loss via unknown mechanisms of inelastic scattering. Here we employ quantum dots to inject and extract electrons at specific energies, to spectrally analyse inelastic scattering inside quantum Hall edge channels. We show that the missing energy puzzle could be untangled by incorporating non-local Auger-like processes, in which energy is redistributed between spatially separate parts of the sample. Our theoretical analysis, accounting for the experimental results, challenges common-wisdom analyses which ignore such non-local decay channels.

Radiation Pneumonitis after Intensity-Modulated Radiotherapy for Esophageal Cancer: Institutional Data and a Systematic Review.

Radiation pneumonitis (RP) is a serious complication that can occur after thoracic radiotherapy. The goal of this study is to investigate the incidence of RP after radiochemotherapy with intensity modulated radiotherapy (IMRT) in patients with esophageal cancer and correlate this with dose volume histogram (DVH) related parameters. For this purpose, the clinical course of 73 patients was evaluated and irradiation doses to the lungs were extracted from radiotherapy treatment plans. Furthermore, a systematic review on this topic was conducted across PubMed. In our institutional cohort, Common Terminology Criteria for Adverse Events (CTCAE) grade II or higher RP occurred in four patients (5.5%). The systematic review identified 493 titles of which 19 studies reporting 874 patients qualified for the final analysis. No grade IV or V RP after radiochemotherapy with IMRT for esophageal cancer was reported in the screened literature. Grade II or higher RP is reported in 6.6% of the patients. A higher incidence can be seen with increasing values for lung V20. In conclusion, our institutional data and the literature consistently show a low incidence of symptomatic RP after radiochemotherapy in patients with esophageal cancer treated with IMRT. However, efforts should be made to keep the lung V20 below 23% and specific caution is warranted in patients with pre-existing lung conditions.

Cloning of the breakpoints of a de novo inversion of chromosome 8, inv (8)(p11.2q23.1) in a patient with Ambras syndrome.

Ambras syndrome (AMS) is a unique form of universal congenital hypertrichosis. In patients with this syndrome, the whole body is covered with fine long hair, except for areas where normally no hair grows. There is accompanying facial dysmorphism and teeth abnormalities, including retarded first and second dentition and absence of teeth. In 1993, Baumeister et al. reported an isolated case of Ambras syndrome in association with a pericentric inversion of chromosome 8. Subsequently, another patient with congenital hypertrichosis and rearrangement of chromosome 8 was reported by Balducci et al. (1998). Both of these patients have a breakpoint in 8q22 in common suggesting that this region of chromosome 8 contains a gene involved in regulation of hair growth. In order to precisely determine the nature of the rearrangement in the case of Ambras syndrome, we have used fluorescent in situ hybridization (FISH) analysis. We have cloned the inversion breakpoints in this patient and generated a detailed physical map of the inversion breakpoint interval. Analysis of the transcripts that map in the vicinity of the breakpoints revealed that the inversion does not disrupt a gene, and suggests that the phenotype is caused by a position effect.

Bioinformatic analysis of autism positional candidate genes using biological databases and computational gene network prediction.

Common genetic disorders are believed to arise from the combined effects of multiple inherited genetic variants acting in concert with environmental factors, such that any given DNA sequence variant may have only a marginal effect on disease outcome. As a consequence, the correlation between disease status and any given DNA marker allele in a genomewide linkage study tends to be relatively weak and the implicated regions typically encompass hundreds of positional candidate genes. Therefore, new strategies are needed to parse relatively large sets of 'positional' candidate genes in search of actual disease-related gene variants. Here we use biological databases to identify 383 positional candidate genes predicted by genomewide genetic linkage analysis of a large set of families, each with two or more members diagnosed with autism, or autism spectrum disorder (ASD). Next, we seek to identify a subset of biologically meaningful, high priority candidates. The strategy is to select autism candidate genes based on prior genetic evidence from the allelic association literature to query the known transcripts within the 1-LOD (logarithm of the odds) support interval for each region. We use recently developed bioinformatic programs that automatically search the biological literature to predict pathways of interacting genes (PATHWAYASSIST and GENEWAYS). To identify gene regulatory networks, we search for coexpression between candidate genes and positional candidates. The studies are intended both to inform studies of autism, and to illustrate and explore the increasing potential of bioinformatic approaches as a compliment to linkage analysis.

Familial primary pulmonary hypertension (gene PPH1) is caused by mutations in the bone morphogenetic protein receptor-II gene.

Familial primary pulmonary hypertension is a rare autosomal dominant disorder that has reduced penetrance and that has been mapped to a 3-cM region on chromosome 2q33 (locus PPH1). The phenotype is characterized by monoclonal plexiform lesions of proliferating endothelial cells in pulmonary arterioles. These lesions lead to elevated pulmonary-artery pressures, right-ventricular failure, and death. Although primary pulmonary hypertension is rare, cases secondary to known etiologies are more common and include those associated with the appetite-suppressant drugs, including phentermine-fenfluramine. We genotyped 35 multiplex families with the disorder, using 27 microsatellite markers; we constructed disease haplotypes; and we looked for evidence of haplotype sharing across families, using the program TRANSMIT. Suggestive evidence of sharing was observed with markers GGAA19e07 and D2S307, and three nearby candidate genes were examined by denaturing high-performance liquid chromatography on individuals from 19 families. One of these genes (BMPR2), which encodes bone morphogenetic protein receptor type II, was found to contain five mutations that predict premature termination of the protein product and two missense mutations. These mutations were not observed in 196 control chromosomes. These findings indicate that the bone morphogenetic protein-signaling pathway is defective in patients with primary pulmonary hypertension and may implicate the pathway in the nonfamilial forms of the disease.

Integrated mapping package--a physical mapping software tool kit.

We have developed an integrated physical mapping computer software package (IMP), originally designed to support the physical mapping of human chromosome 13 and expanded to support several gene-identification projects based on the positional candidate approach. IMP displays map data in a form that provides useful guidelines to the end users. An integrated map with high resolution and confidence is constructed from different types of mapping data, including hybridization experiments, STS-based PCR assays, genetic linkage mapping, cDNA localization, and FISH data. The map is also designed to provide suggestions for specific experiments that are required to obtain maps with even higher resolution and confidence. To this end, the optimization employs multiple constraints that take into account already established STS "scaffold" maps. This software thus serves as an important general tool kit for physical mapping, sequencing, and gene-hunting projects.

Load-controlled compression of articular cartilage induces a transient stimulation of aggrecan gene expression.

The effects of short- and long-term load-controlled compression on the levels of aggrecan mRNA have been determined. Results show that a compressive stress of 0.1 MPa on bovine articular cartilage explants for 1, 4, 12, and 24 h produces a transient up-regulation of aggrecan mRNA synthesis. At 1 h, aggrecan mRNA levels in loaded explants were increased 3.2-fold compared to control explants. At longer times (>/=4 h), the levels of aggrecan mRNA returned to baseline values or stayed slightly higher. There is a dose dependence in the response of the explant to increasing levels of compressive stress (0-0.5 MPa) for 1 h. Aggrecan mRNA levels increased 2- to 3-fold at 0-0.25 MPa. At 0.5 MPa, the level of aggrecan mRNA was lower than those at 0.1 and 0.25 MPa. This dose-dependent effect suggests a reversal of the stimulatory effects of compression on aggrecan gene expression at higher loads. After 24 h of compression, the levels of aggrecan mRNA in explants subjected to any of the stress levels were not significantly different from those in control explants. The stimulatory effect of 0.1 MPa compressive stress on aggrecan mRNA levels was blocked by Rp-cAMP and U-73122, indicating the involvement of the classical signal transduction pathways in the mechanical modulation of aggrecan gene expression. The responses of link protein mRNA to compression paralleled those of aggrecan, while there was no significant change in expression of the gene for the housekeeping protein elongation factor-1 alpha. The results indicate that articular cartilage chondrocytes can respond to short-term compressive loads by transiently up-regulating expression of the aggrecan gene. The fact that long-term compression did not significantly alter aggrecan mRNA levels suggests that previously observed inhibitory effects of prolonged static compression on proteoglycan synthesis in articular cartilage may be, for the most part, mediated through mechanisms other than suppression of aggrecan mRNA levels.

Regulatory activities of the 5'- and 3'-untranslated regions and promoter of the human aggrecan gene.

Identification and characterization of the regulatory elements of the human aggrecan gene are necessary first steps in addressing the molecular mechanisms through which the gene is regulated. Using luciferase reporter constructs driven by the human aggrecan promoter or the cytomegalovirus promoter, the 5'- and 3'-untranslated regions of the human aggrecan gene were found to regulate gene expression transcriptionally in a promoter- and/or cell type-specific manner. Independent of cell type, the 5'-untranslated region was inhibitory with respect to the cytomegalovirus promoter, but it was stimulatory to the human aggrecan promoter. The 5'-untranslated region inhibited the cytomegalovirus promoter by approximately 60% in both chondrocytes and NIH 3T3 cells, but it stimulated the activity of the human aggrecan promoter about 8-fold in chondrocytes and 40-fold in NIH 3T3 cells. In contrast, the 3'-untranslated region inhibited the activities of the human aggrecan promoter by 40-70% in both cell types, but it stimulated the cytomegalovirus promoter activities by 50-60% in NIH 3T3 cells and inhibited its activity by 70% in chondrocytes. The differential effects of the untranslated regions on the two types of promoters may be a reflection of differences in regulation of TATA-less promoters, such as the human aggrecan promoter, and TATA-containing promoters, such as the cytomegalovirus promoter.

High-resolution YAC-cosmid-STS map of human chromosome 13.

We have assembled a high-resolution physical map of human chromosome 13 DNA (approximately 114 Mb) from hybridization, PCR, and FISH mapping data using a specifically designed set of computer programs. Although the mapping of 13p is limited, 13q (approximately 98 Mb) is covered by an almost continuous contig of 736 YACs aligned to 597 contigs of cosmids. Of a total of 10,789 cosmids initially selected from a chromosome 13-specific cosmid library (16,896 colonies) using inter-Alu PCR probes from the YACs and probes for markers mapped to chromosome 13, 511 were assembled in contigs that were established from cross-hybridization relationships between the cosmids. The 13q YAC-cosmid map was annotated with 655 sequence tagged sites (STSs) with an average spacing of 1 STS per 150 kb. This set of STSs, each identified by a D number and cytogenetic location, includes database markers (198), expressed sequence tags (93), and STSs generated by sequencing of the ends of cosmid inserts (364). Additional annotation has been provided by positioning 197 cosmids mapped by FISH on 13q. The final (comprehensive) map, a list of STS primers, and raw data used in map assembly are available at our Web site (genome1.ccc.columbia.edu/ approximately genome/) and can serve as a resource to facilitate accurate localization of additional markers, provide substrates for sequencing, and assist in the discovery of chromosome 13 genes associated with hereditary diseases.

A single gene for human TRAF-3 at chromosome 14q32.3 encodes a variety of mRNA species by alternative polyadenylation, mRNA splicing and transcription initiation.

Human TRAF-3 is a signaling molecule that interacts with the cytoplasmic tails of CD40 and other TNF-receptor family members. TRAF-3 mRNA is expressed as two major classes of approximately 2 and 8 kb and a number of TRAF-3 encoding cDNA clones differ in discrete gene segments. Because this variety of mRNA species could result from mRNA processing events and/or multiple genes, the structure and localization of TRAF-3 encoding gene elements were determined. FISH and radiation hybrid mapping demonstrated that TRAF-3 is located at chromosome 14q32.3, approximately 1 Mb centromeric to the Ig heavy chain gene complex. Physical mapping of four overlapping genomic PAC clones established that TRAF-3 transcripts are encoded by a single gene, comprised of 13 exons and spanning 130 kb. Alternative polyadenylation in the mRNA segment encoded by exon 12 accounts for the difference between the 2 kb and the 8 kb classes of transcripts. Alternative mRNA splicing in the coding region (encoded by exons 3-12) generates transcripts which delete exons 8 (75 nt), 7+8 (156 nt) or 8+9 (168 nt) and that encode distinct protein isoforms (delta25, delta52 and delta56 aa, respectively). Alternative splicing of exon 2 (139 nt) and alternative transcriptional initiation result in mRNA species with distinct 5'UTRs. Together, these data indicate that a single TRAF-3 gene encodes a variety of mRNA species by a combination of alternative polyadenylation, alternative mRNA splicing and/or alternative initiation.

Cloning and gene mapping of the chromosome 13q14 region deleted in chronic lymphocytic leukemia.

Frequent deletions and loss of heterozygosity in a segment of chromosome 13 (13q14) in cases of B-cell chronic lymphocytic leukemia (CLL) have suggested that this malignancy is caused by inactivation of an unknown tumor suppressor gene located in this region. Toward the identification of the putative CLL tumor suppressor, we have constructed a high-resolution physical map of YAC, PAC, and cosmid contigs covering 600 kb of the 13q14 genomic region. In addition to densely positioned genetic markers and STSs, this map was further annotated by localization of 32 transcribed sequences (ESTs) using a combination of exon trapping, direct cDNA selection, sample sequencing of cosmids and PACs, and homology searches. On the basis of these mapping data, allelic loss analyses at 13q14 using CLL tumor samples allowed narrowing of the genomic segment encompassing the putative CLL gene to <300 kb. Twenty-three ESTs located within this minimally deleted region are candidate exons for the CLL-associated tumor suppressor gene.

Genomic cloning and chromosomal assignment of the E2F dimerization partner TFDP gene family.

The TFDP genes encode a family of transcription factors that can form heterodimers with E2F family proteins in vivo. The E2F-TFDP transcription factors are major regulators of genes that are required for the progression of S-phase, such as DHFR and DNA polymerase alpha, and they play a critical role in cell cycle regulation and differentiation. The retinoblastoma tumor suppressor protein has been shown to induce growth arrest by binding to E2F-TFDP and repressing its activity. Two human TFDP genes have been cloned, namely TFDP1 and TFDP2 (or DP1 and DP2). In the present study, we identified genomic clones of TFDP1, its pseudogene TFDP1P and TFDP2, and we mapped them to chromosome 13q34, 1q32.3, and 3q23, respectively. Chromosomal abnormalities involving regions 13q34 and 3q23 have been reported in certain lymphomas and other diseases associated with loss of cell cycle regulation, and the involvement of the TFDP transcription factors remains to be elucidated.

A high-resolution annotated physical map of the human chromosome 13q12-13 region containing the breast cancer susceptibility locus BRCA2.

Various types of physical mapping data were assembled by developing a set of computer programs (Integrated Mapping Package) to derive a detailed, annotated map of a 4-Mb region of human chromosome 13 that includes the BRCA2 locus. The final assembly consists of a yeast artificial chromosome (YAC) contig with 42 members spanning the 13q12-13 region and aligned contigs of 399 cosmids established by cross-hybridization between the cosmids, which were selected from a chromosome 13-specific cosmid library using inter-Alu PCR probes from the YACs. The end sequences of 60 cosmids spaced nearly evenly across the map were used to generate sequence-tagged sites (STSs), which were mapped to the YACs by PCR. A contig framework was generated by STS content mapping, and the map was assembled on this scaffold. Additional annotation was provided by 72 expressed sequences and 10 genetic markers that were positioned on the map by hybridization to cosmids.

Quantitative polymerase chain reaction assay for aggrecan and link protein gene expression in cartilage.

A procedure has been developed to quantify the levels of aggrecan and link protein mRNAs in small amounts of various tissues, including cartilage, using the power of PCR to amplify extremely low levels of specific templates. The PCR protocol which was selected allows for a simple assay procedure, with standards in different tubes from the samples. This straightforward procedure is quantitative, inexpensive, and allows for many samples to be analyzed at one time.

Tumor response, toxicity, and survival after neoadjuvant organ-preserving chemotherapy for advanced laryngeal carcinoma. The Department of Veterans Affairs Cooperative Laryngeal Cancer Study Group.

PURPOSE: In 1984, the Department of Veterans Affairs Cooperative Studies Program began a trial in which patients with resectable squamous cell carcinoma of the larynx were randomized to receive standard surgery followed by radiation therapy or to receive neoadjuvant therapy with cisplatin and fluorouracil (5-FU) followed by radiation therapy for those achieving a greater than 50% tumor response to chemotherapy. This analysis reviews the tumor responses, toxicity, compliance, and long-term survival for those patients randomized to the chemotherapy arm. PATIENTS AND METHODS: One hundred sixty-six patients were randomized to the chemotherapy arm. Standard tumor response data, chemotherapy toxicity, and survival have been examined using standard statistical methods. RESULTS: The high response rates and acceptable toxicity to cisplatin and 5-FU of previously untreated patients were confirmed. Long-term disease-free survival was more likely to occur in patients who achieved a complete response to chemotherapy, particularly in those who had a confirmed histologic response to chemotherapy. Pretreatment histologic growth patterns were highly predictive of responses to chemotherapy. CONCLUSION: Neoadjuvant chemotherapy was well tolerated and did not negatively affect the definitive treatment that followed. The survival of nonresponding patients who underwent prompt salvage surgery was also not impaired. The role of organ preservation should be explored in other head and neck sites.

Assembly of ordered contigs of cosmids selected with YACs of human chromosome 13.

We have developed an efficient method for assembling ordered cosmid contigs aligned to mega-YACs and midi-YACs (average insert sizes of 1.0 and 0.35 Mb, respectively) and used this general method to initiate high-resolution physical mapping of human chromosome 13 (Chr 13). Chr 13-enriched midi-YAC (mYAC) and mega-YAC (MYAC) sublibraries were obtained from corresponding CEPH total human YAC libraries by selecting colonies with inter-Alu PCR probes derived from Chr 13 monochromosomal cell hybrid DNA. These sublibraries were arrayed on filters at high density. In our approach, the MYAC 13 sublibrary is screened by hybridization with cytogenetically assigned Chr 13 DNA probes to select one or a small subset of MYACs. Inter-Alu PCR products from each MYAC are then hybridized to the MYAC and mYAC sublibraries to identify overlapping YACs and to an arrayed Chr 13-specific cosmid library to select corresponding cosmids. The set of selected cosmids, gridded on filters at high density, is hybridized with inter-Alu PCR products from each of the overlapping YACs to identify subsets of cosmids and also with riboprobes from each cosmid of the arrayed set ("cosmid matrix cross-hybridization"). From these data, cosmid contigs are assembled by a specifically designed computer program. Application of this method generates cosmid contigs spanning the length of a MYAC with few gaps. To provide a high-resolution map, ends of cosmids are sequenced at preselected sites to position densely spaced sequence-tagged sites.

An algorithm based on graph theory for the assembly of contigs in physical mapping of DNA.

An algorithm is described for mapping DNA contigs based on an interval graph (IG) representation. In general terms, the input to the algorithm is a set of binary overlapping relations among finite intervals spread along a real line, from which the algorithm generates sets of ordered overlapping fragments spanning that line. The implications of a more general case of the IG, called a probe interval graph (PIG), in which only a subset of cosmids are used as probes, are also discussed. In the specific case of cosmids hybridizing to regions of a YAC, the algorithm takes cross-hybridization information using the cosmids as probes, and orders them along the YAC; if gaps exist due to insufficient coverage of cosmid contigs along the length of the YAC, repetitive use of the algorithm generates sets of ordered overlapping fragments. Both the IG and the PIG can expose problems caused by false overlaps, such as hybridizations due to repetitive elements. The algorithm, has been coded in C; CPU time is essentially linear with respect to the number of cosmids analyzed. Results are presented for the application of a PIG to cosmid contig assembly along a human chromosome 13-specific YAC. An alignment of 67 cosmids spanning a YAC took 0.28 seconds of CPU time on a Convex 220 computer.

Mapping, cloning and genetic characterization of the region containing the Wilson disease gene.

Wilson disease (WD) is an autosomal recessive disorder of copper transport which map to chromosome 13q14.3. In pursuit of the WD gene, we developed yeast artificial chromosome and cosmid contigs, and microsatellite markers which span the WD gene region. Linkage disequilibrium and haplotype analysis of 115 WD families confined the disease locus to a single marker interval. A candidate cDNA clone was mapped to this interval which, as shown in the accompanying paper, is very likely the WD gene. Our haplotype and mutation analyses predict that approximately half of all WD mutations will be rare in the American and Russian populations.