Engineering Cancer/Testis Antigens With Reversible S-Cationization to Evaluate Antigen Spreading

Serum autoantibody to cancer/testis antigens (CTAs) is a critical biomarker that reflects the antitumor immune response. Quantitative and multiplexed anti-CTA detection arrays can assess the immune status in tumors and monitor therapy-induced antitumor immune reactions. Most full-length recombinant CTA proteins tend to aggregate. Cysteine residue-specific S-cationization techniques facilitate the preparation of water-soluble and full-length CTAs. Combined with Luminex technology, we designed a multiple S-cationized antigen-immobilized bead array (MUSCAT) assay system to evaluate multiple serum antibodies to CTAs. Reducible S-alkyl-disulfide-cationized antigens in cytosolic conditions were employed to develop rabbit polyclonal antibodies as positive controls. These control antibodies sensitively detected immobilized antigens on beads and endogenous antigens in human lung cancer-derived cell lines. Rabbit polyclonal antibodies successfully confirmed the dynamic ranges and quantitative MUSCAT assay results. An immune monitoring study was conducted using the serum samples on an adenovirus-mediated REIC/Dkk-3 gene therapy clinical trial that showed a successful clinical response in metastatic castration-resistant prostate cancer. Autoantibody responses were closely related to clinical outcomes. Notably, upregulation of anti-CTA responses was monitored before tumor regression. Thus, quantitative monitoring of anti-CTA antibody biomarkers can be used to evaluate the cancer-immunity cycle. A quality-certified serum autoantibody monitoring system is a powerful tool for developing and evaluating cancer immunotherapy

Characterization Experiments of Secondary Arcs on Solar Arrays: Threshold and Duration

An electrostatic discharge test performed on a solar array panel is one of the important tests carried out before spacecraft launch to ensure spacecraft reliability in orbit. In this study, the effects of secondary arcs on the solar array, which can cause catastrophic accidents, are considered. The sustained-arc threshold of multijunction and silicon solar arrays was investigated to establish the design guidelines for ensuring the safety of satellite solar arrays. In this experimental study, the string voltage, string current, gap length, and solar-cell type were selected as test parameters. Gap lengths were 0.5, 0.8, 1.0, and 2.0 mm. For 0.5 and 0.8mmof gap length, a permanent sustained arc occurred under the condition of 1.5 A of string current for 50, 70, 90, and 110 V of string voltage. Furthermore, temporary sustained-arc duration exponentially increased with increasing string current. Temporary sustained-arc duration longer than 1 ms can serve as a practical alarm for the imminent permanent sustained-arc inception

Statistical Number of Primary Discharges Required for Solar Array Secondary Arc Tests

In the present paper, we propose a method to assess the risk of a secondary arc on a solar array in orbit based on statistical theory. Ground experiments have been carried out to investigate the statistics of temporary-sustained-arc duration. We have found that secondary-arc duration can be modeled as a combination of multiple Poisson distributions. Once we know the formula of the Poisson distribution, we can easily calculate the probability of a secondary arc lasting longer than an acceptable limit. The quantity of secondary-arc-duration data needed to define the Poisson distribution formula has been discussed. Assuming that thousands or tens of thousands of primary arcs occur in orbit, if the average duration of secondary arcs in the test is less than 5% of the acceptable limit for secondary-arc duration, we can greatly reduce the number of experimental trials to 10 or less in most cases

Effect of Atomic Oxygen Exposure on Surface Resistivity Change of Spacecraft Insulator Material

Spacecraft surface charging can lead to arcing and a loss of electricity generation capability in solar panels or even loss of a satellite. The charging problem may be further aggravated by atomic oxygen (AO) exposure in Low Earth orbits, which modifies the surface of materials like polyimide, Teflon, anti-reflective coatings, cover glass etc, used on satellite surfaces, affecting materials properties, such as resistivity, secondary electron emissivity and photo emission, which govern the charging behavior. These properties are crucial input parameters for spacecraft charging analysis. To study the AO exposure effect on charging governing properties, an atomic oxygen exposure facility based on laser detonation of oxygen was built. The facility produces AO with a peak velocity value around 10-12km/s and a higher flux than that existing in orbit. After exposing the polyimide test material to the equivalent of 10 years of AO fluence at an altitude of 700-800 km, surface charging properties like surface resistivity and volume resistivity were measured. The measurement was performed in a vacuum using the charge storage decay method at room temperature, which is considered the most appropriate for measuring resistivity for space applications. The results show that the surface resistivity increases and the volume resistivity remains almost the same for the AO exposure fluence of 5.4×1018 atoms cm-2

Environmental Effects on Solar Array Electrostatic Discharge Current Waveforms and Test Results

A solar array electrostatic discharge ground test is necessary to assure spacecraft reliability in orbit. Laboratory experiments were carried out to characterize an electrostatic discharge current waveform with different background pressures and charging environments to identify the importance of the test setup. The waveform strongly depended on the background pressure. This difference can affect the result of the solar cell degradation test. However, in the case of the secondary arc test, the difference of the primary arc current waveform did not affect the duration of the secondary arc. The current available from a power supply mostly determined the duration of the secondary, irrespective of the test environment. Methods to control the primary arc current supplied by an external capacitance are proposed

Identification of Pou5f1, Sox2, and Nanog downstream target genes with statistical confidence by applying a novel algorithm to time course microarray and genome-wide chromatin immunoprecipitation data

<p>Abstract</p> <p>Background</p> <p>Target genes of a transcription factor (TF) <it>Pou5f1 </it>(<it>Oct3/4 </it>or <it>Oct4</it>), which is essential for pluripotency maintenance and self-renewal of embryonic stem (ES) cells, have previously been identified based on their response to <it>Pou5f1 </it>manipulation and occurrence of Chromatin-immunoprecipitation (ChIP)-binding sites in promoters. However, many responding genes with binding sites may not be direct targets because response may be mediated by other genes and ChIP-binding site may not be functional in terms of transcription regulation.</p> <p>Results</p> <p>To reduce the number of false positives, we propose to separate responding genes into groups according to direction, magnitude, and time of response, and to apply the false discovery rate (FDR) criterion to each group individually. Using this novel algorithm with stringent statistical criteria (FDR < 0.2) to a compendium of published and new microarray data (3, 6, 12, and 24 hr after <it>Pou5f1 </it>suppression) and published ChIP data, we identified 420 tentative target genes (TTGs) for <it>Pou5f1</it>. The majority of TTGs (372) were down-regulated after <it>Pou5f1 </it>suppression, indicating that the <it>Pou5f1 </it>functions as an activator of gene expression when it binds to promoters. Interestingly, many activated genes are potent suppressors of transcription, which include polycomb genes, zinc finger TFs, chromatin remodeling factors, and suppressors of signaling. Similar analysis showed that <it>Sox2 </it>and <it>Nanog </it>also function mostly as transcription activators in cooperation with <it>Pou5f1</it>.</p> <p>Conclusion</p> <p>We have identified the most reliable sets of direct target genes for key pluripotency genes – <it>Pou5f1</it>, <it>Sox2</it>, and <it>Nanog</it>, and found that they predominantly function as activators of downstream gene expression. Thus, most genes related to cell differentiation are suppressed indirectly.</p

Midkine promoter-based conditionally replicative adenovirus therapy for midkine-expressing human pancreatic cancer

<p>Abstract</p> <p>Background</p> <p>To develop a novel therapeutic strategy for human pancreatic cancer using a midkine promoter-based conditionally replicating adenovirus.</p> <p>Methods</p> <p>We examined midkine mRNA expression and midkine protein expression by seven human pancreatic cancer cell lines (AsPC-1, BxPC-3, CFPAC-1, HPAC, MIAPaCa-2, PANC-1, and Suit-2), as well as by non-cancerous pancreatic tissue and pancreatic cancers. Midkine promoter activity was measured in cancer cell lines by the dual luciferase reporter assay. Adenoviral transduction efficiency was assessed by fluorescent staining of cancer cell lines using adenovirus type 5 containing the green fluorescent protein gene (Ad5GFP). Replication of adenovirus type 5 containing the 0.6 kb midkne promoter (Ad5MK) was assessed by the detection of E1 protein in cancer cell lines. The cytotoxicity of Ad5MK for cancer cells was evaluated from the extent of growth inhibition after viral infection. Infection and replication were also assessed in nude mice with subcutaneous Suit-2 tumors by intratumoral injection of Ad5MK, Ad5GFP, or vehicle. E1a mRNA expression in the treated tumors and expression of the replication-specific adenoviral hexon protein were evaluated. Finally, the anti-tumor activity of Ad5MK against intraperitoneal xenografts of Suit-2 pancreatic cancer cells was examined after intraperitoneal injection of the virus.</p> <p>Results</p> <p>Both midkine mRNA expression and midkine protein expression were strong in AsPC-1 and CFPAC-1 cell liens, moderate in BxPC-3, HPAC, and Suit-2 cell lines, and weak in PANC-1 and MIAPaCa-2 cell lines. Expression of midkine mRNA was significantly stronger in pancreatic cancers than in non-cancerous pancreatic tissues. The relative luciferase activity mediated by the 0.6 kb midkne fragment in AsPC-1, PANC-1, and Suit-2 cell lines was approximately 6 to 20 times greater than that in midkne-negative MIAPaCa-2 cell lines. Pancreatic cancer cell lines exhibited a heterogeneous adenoviral transduction profile. E1A expression was higher in cell lines with strong midkine expression than in cell lines with weak midkine expression. Ad5MK showed much greater cytotoxicity for midkine-expressing Suit-2 and PANC-1 cell lines than for midkine-negative MIAPaCa-2 cell lines. In the Suit-2 subcutaneous xenograft model, expression of E1A was detected in Ad5MK-treated tumors, but not in untreated and Ad5GFP-treated tumors. In the Suit-2 intraperitoneal xenograft model, the Ad5MK group survived for significantly longer than the Ad5GFP, PBS, and untreated groups.</p> <p>Conclusion</p> <p>Ad5MK has an anti-tumor effect against human pancreatic cancer cell lines that express midkine mRNA. Midkine promoter-based conditionally replicative adenovirus might be a promising new gene therapy for pancreatic cancer.</p

Spectral evolution of GRB 060904A observed with Swift and Suzaku -- Possibility of Inefficient Electron Acceleration

We observed an X-ray afterglow of GRB 060904A with the Swift and Suzaku satellites. We found rapid spectral softening during both the prompt tail phase and the decline phase of an X-ray flare in the BAT and XRT data. The observed spectra were fit by power-law photon indices which rapidly changed from $\Gamma = 1.51^{+0.04}_{-0.03}$ to $\Gamma = 5.30^{+0.69}_{-0.59}$ within a few hundred seconds in the prompt tail. This is one of the steepest X-ray spectra ever observed, making it quite difficult to explain by simple electron acceleration and synchrotron radiation. Then, we applied an alternative spectral fitting using a broken power-law with exponential cutoff (BPEC) model. It is valid to consider the situation that the cutoff energy is equivalent to the synchrotron frequency of the maximum energy electrons in their energy distribution. Since the spectral cutoff appears in the soft X-ray band, we conclude the electron acceleration has been inefficient in the internal shocks of GRB 060904A. These cutoff spectra suddenly disappeared at the transition time from the prompt tail phase to the shallow decay one. After that, typical afterglow spectra with the photon indices of 2.0 are continuously and preciously monitored by both XRT and Suzaku/XIS up to 1 day since the burst trigger time. We could successfully trace the temporal history of two characteristic break energies (peak energy and cutoff energy) and they show the time dependence of $\propto t^{-3} \sim t^{-4}$ while the following afterglow spectra are quite stable. This fact indicates that the emitting material of prompt tail is due to completely different dynamics from the shallow decay component. Therefore we conclude the emission sites of two distinct phenomena obviously differ from each other.Comment: 19 pages, 9 figures, accepted for publication in PASJ (Suzaku 2nd Special Issue