A platform trial of neoadjuvant and adjuvant antitumor vaccination alone or in combination with PD-1 antagonist and CD137 agonist antibodies in patients with resectable pancreatic adenocarcinoma.

A neoadjuvant immunotherapy platform clinical trial allows for rapid evaluation of treatment-related changes in tumors and identifying targets to optimize treatment responses. We enrolled patients with resectable pancreatic adenocarcinoma into such a platform trial (NCT02451982) to receive pancreatic cancer GVAX vaccine with low-dose cyclophosphamide alone (Arm A; n = 16), with anti-PD-1 antibody nivolumab (Arm B; n = 14), and with both nivolumab and anti-CD137 agonist antibody urelumab (Arm C; n = 10), respectively. The primary endpoint for Arms A/B - treatment-related change in IL17A expression in vaccine-induced lymphoid aggregates - was previously published. Here, we report the primary endpoint for Arms B/C: treatment-related change in intratumoral CD8+ CD137+ cells and the secondary outcomes including safety, disease-free and overall survivals for all Arms. Treatment with GVAX+nivolumab+urelumab meets the primary endpoint by significantly increasing intratumoral CD8+ CD137+ cells (p = 0.003) compared to GVAX+Nivolumab. All treatments are well-tolerated. Median disease-free and overall survivals, respectively, are 13.90/14.98/33.51 and 23.59/27.01/35.55 months for Arms A/B/C. GVAX+nivolumab+urelumab demonstrates numerically-improved disease-free survival (HR = 0.55, p = 0.242; HR = 0.51, p = 0.173) and overall survival (HR = 0.59, p = 0.377; HR = 0.53, p = 0.279) compared to GVAX and GVAX+nivolumab, respectively, although not statistically significant due to small sample size. Therefore, neoadjuvant and adjuvant GVAX with PD-1 blockade and CD137 agonist antibody therapy is safe, increases intratumoral activated, cytotoxic T cells, and demonstrates a potentially promising efficacy signal in resectable pancreatic adenocarcinoma that warrants further study.

Determination of ADH in textiles using the HPLC-MS/MS method and the study of its adsorption behaviour towards formaldehyde.

In the textile industry, formaldehyde-based resins are used as finishers to make the fabrics crease-resistant, which are the main source of formaldehyde in textiles. In our practical study, there are cases that prove that textile products containing adipic dihydrazide (ADH) will continuously adsorb formaldehyde from the surrounding environment during storage. In this study, a high performance liquid chromatography-tandem mass spectrometry method was established for the precise determination of ADH in textiles. The method was optimized in terms of instrument conditions, extraction temperature, extraction time, and extraction mode. Under optimum test conditions, ADH was determined precisely with the linearity range of 0.05-2 mg L-1 and correlation coefficient (R 2) of 0.9993. Recovery rate and repeatability were tested; the data showed that the recovery rate of ADH in textiles was in the range of 85-100%, and the RSD (relative standard deviation) was less than 10%. The ADH-positive textile samples were placed in designed environments for some time to adsorb formaldehyde. The adsorbed amounts of formaldehyde in the textile samples first increase and then decrease with time. The maximum amount of formaldehyde a sample can adsorb increases with an increase in its ADH content and will stop increasing once its ADH content exceeds 1700 mg kg-1. The placement environment has a little effect on the maximum adsorption capacity of the samples towards formaldehyde, but can significantly affect the adsorption rate and equilibrium adsorption capacity.

Bone mineral imaged in vivo by 31P solid state MRI of human wrists.

PURPOSE: To implement solid state (31)P MRI ((31)P SMRI) in a clinical scanner to visualize bone mineral. MATERIALS AND METHODS: Wrists of seven healthy volunteers were scanned. A quadrature wrist (31)P transmit/receive coil provided strong B(1) and good signal-to-noise ratio (SNR). A (1)H-(31)P frequency converter was constructed to enable detection of the (31)P signal by means of the (1)H channel. Data points lost in the receiver dead time were recovered by a second acquisition with longer dwell time and lower gradient strength. RESULTS: Three-dimensional (31)P images, showing only bone mineral of the wrist, were obtained with a clinical 3 Tesla (T) scanner. In the best overall case an image with isotropic resolution of ∼5.1 mm and SNR of 30 was obtained in 37 min. (31)P NMR properties (resonance line width 2 kHz and T(1) 17-19 s) of in vivo human bone mineral were measured. CONCLUSION: In vivo (31)P SMRI visualization of human wrist bone mineral with a clinical MR scanner is feasible with suitable modifications to circumvent the scanners' limitations in reception of short-T(2) signals. Frequency conversion methodology is useful for implementing (31)P SMRI measurements on scanners which do not have multinuclear capability or for which the multinuclear receiver dead time is excessive.

Bone matrix imaged in vivo by water- and fat-suppressed proton projection MRI (WASPI) of animal and human subjects.

PURPOSE: To demonstrate water- and fat-suppressed proton projection MRI (WASPI) in a clinical scanner to visualize the solid bone matrix in animal and human subjects. MATERIALS AND METHODS: Pig bone specimens and polymer pellets were used to optimize the WASPI method in terms of soft-tissue suppression, image resolution, signal-to-noise ratio, and scan time on a 3T MRI scanner. The ankles of healthy 2-3-month-old live Yorkshire pigs were scanned with the optimized method. The method was also applied to the wrists of six healthy adult human volunteers to demonstrate the feasibility of the WASPI method in human subjects. A transmit/receive coil built with proton-free materials was utilized to produce a strong B(1) field. A fast transmit/receive switch was developed to reduce the long receiver dead time that would otherwise obscure the signals. RESULTS: Clear 3D WASPI images of pig ankles and human wrists, showing only the solid bone matrix and other tissues with high solid content (eg, tendons), with a spatial resolution of 2.0 mm in all three dimensions were obtained in as briefly as 12 minutes. CONCLUSION: WASPI of the solid matrix of bone in humans and animals in vivo is feasible.

Quantitative (31)P NMR spectroscopy and (1)H MRI measurements of bone mineral and matrix density differentiate metabolic bone diseases in rat models.

In this study, bone mineral density (BMD) of normal (CON), ovariectomized (OVX), and partially nephrectomized (NFR) rats was measured by (31)P NMR spectroscopy; bone matrix density was measured by (1)H water- and fat-suppressed projection imaging (WASPI); and the extent of bone mineralization (EBM) was obtained by the ratio of BMD/bone matrix density. The capability of these MR methods to distinguish the bone composition of the CON, OVX, and NFR groups was evaluated against chemical analysis (gravimetry). For cortical bone specimens, BMD of the CON and OVX groups was not significantly different; BMD of the NFR group was 22.1% (by (31)P NMR) and 17.5% (by gravimetry) lower than CON. For trabecular bone specimens, BMD of the OVX group was 40.5% (by (31)P NMR) and 24.6% (by gravimetry) lower than CON; BMD of the NFR group was 26.8% (by (31)P NMR) and 21.5% (by gravimetry) lower than CON. No significant change of cortical bone matrix density between CON and OVX was observed by WASPI or gravimetry; NFR cortical bone matrix density was 10.3% (by WASPI) and 13.9% (by gravimetry) lower than CON. OVX trabecular bone matrix density was 38.0% (by WASPI) and 30.8% (by gravimetry) lower than CON, while no significant change in NFR trabecular bone matrix density was observed by either method. The EBMs of OVX cortical and trabecular specimens were slightly higher than CON but not significantly different from CON. Importantly, EBMs of NFR cortical and trabecular specimens were 12.4% and 26.3% lower than CON by (31)P NMR/WASPI, respectively, and 4.0% and 11.9% lower by gravimetry. Histopathology showed evidence of osteoporosis in the OVX group and severe secondary hyperparathyroidism (renal osteodystrophy) in the NFR group. These results demonstrate that the combined (31)P NMR/WASPI method is capable of discerning the difference in EBM between animals with osteoporosis and those with impaired bone mineralization.

Quantitative bone matrix density measurement by water- and fat-suppressed proton projection MRI (WASPI) with polymer calibration phantoms.

The density of the organic matrix of bone substance is a critical parameter necessary to clinically evaluate and distinguish structural and metabolic pathological conditions such as osteomalacia in adults and rickets in growing children. Water- and fat-suppressed proton projection MRI (WASPI) was developed as a noninvasive means to obtain this information. In this study, a density calibration phantom was developed to convert WASPI intensity to true bone matrix density. The phantom contained a specifically designed poly(ethylene oxide)/poly(methyl methacrylate) (PEO/PMMA) blend, whose MRI properties (T(1), T(2), and resonance linewidth) were similar to those of solid bone matrix (collagen, tightly bound water, and other immobile molecules), minimizing the need to correct for differences in T(1) and/or T(2) relaxation between the phantom and the subject. Cortical and trabecular porcine bone specimens were imaged using WASPI with the calibration phantom in the field of view (FOV) as a stable intensity reference. Gravimetric and amino acid analyses were carried out on the same specimens after WASPI, and the chemical results were found to be highly correlated (r(2) = 0.98 and 0.95, respectively) to the WASPI intensity. By this procedure the WASPI intensity can be used to obtain the true bone matrix mass density in g cm(-3).

Murine intramyocellular lipids quantified by NMR act as metabolic biomarkers in burn trauma.

It has been suggested that intramyocellular lipids (IMCLs) may serve as biomarkers of insulin resistance and mitochondrial dysfunction. Using a hind-limb mouse model of burn trauma, we tested the hypothesis that severe localized burn trauma involving 5% of the total body surface area causes a local increase in IMCLs in the leg skeletal muscle. We quantified IMCLs from ex vivo intact tissue specimens using High-Resolution Magic Angle Spinning (HRMAS) 1H NMR and characterized the accompanying gene expression patterns in burned versus control skeletal muscle specimens. We also quantified plasma-free fatty acids (FFAs) in burn versus control mice. Our results from HRMAS 1H NMR measurements indicated that IMCL levels were significantly increased in mice exposed to burn trauma. Furthermore, plasma FFA levels were also significantly increased, and gene expression of Glut4, insulin receptor substrate 1 (IRS1), glycolytic genes, and PGC-1beta was downregulated in these mice. Backward stepwise multiple linear regression analysis demonstrated that IMCL levels correlated significantly with FFA levels, which were a significant predictor of IRS1 and PGC-1beta gene expression. We conclude from these findings that IMCLs can serve as metabolic biomarkers in burn trauma and that FFAs and IMCLs may signal altered metabolic gene expression. This signaling may result in the observed burn-induced insulin resistance and skeletal muscle mitochondrial dysfunction. We believe that IMCLs may therefore be useful biomarkers in predicting the therapeutic effectiveness of hypolipidemic agents for patients with severe burns.

Reduced rate of adenosine triphosphate synthesis by in vivo 31P nuclear magnetic resonance spectroscopy and downregulation of PGC-1beta in distal skeletal muscle following burn.

Using a mouse model of burn trauma, we tested the hypothesis that severe burn trauma corresponding to 30% of total body surface area (TBSA) causes reduction in adenosine triphosphate (ATP) synthesis in distal skeletal muscle. We employed in vivo 31P nuclear magnetic resonance (NMR) in intact mice to assess the rate of ATP synthesis, and characterized the concomitant gene expression patterns in skeletal muscle in burned (30% TBSA) versus control mice. Our NMR results showed a significantly reduced rate of ATP synthesis and were complemented by genomic results showing downregulation of the ATP synthase mitochondrial F1 F0 complex and PGC-1beta gene expression. Our findings suggest that inflammation and muscle atrophy in burns are due to a reduced ATP synthesis rate that may be regulated upstream by PGC-1beta. These findings implicate mitochondrial dysfunction in distal skeletal muscle following burn injury. That PGC-1beta is a highly inducible factor in most tissues and responds to common calcium and cyclic adenosine monophosphate (cAMP) signaling pathways strongly suggests that it may be possible to develop drugs that can induce PGC-1beta.

Combination of high-resolution magic angle spinning proton magnetic resonance spectroscopy and microscale genomics to type brain tumor biopsies.

Advancements in the diagnosis and prognosis of brain tumor patients, and thus in their survival and quality of life, can be achieved using biomarkers that facilitate improved tumor typing. We introduce and implement a combinatorial metabolic and molecular approach that applies state-of-the-art, high-resolution magic angle spinning (HRMAS) proton (1H) MRS and gene transcriptome profiling to intact brain tumor biopsies, to identify unique biomarker profiles of brain tumors. Our results show that samples as small as 2 mg can be successfully processed, the HRMAS 1H MRS procedure does not result in mRNA degradation, and minute mRNA amounts yield high-quality genomic data. The MRS and genomic analyses demonstrate that CNS tumors have altered levels of specific 1H MRS metabolites that directly correspond to altered expression of Kennedy pathway genes; and exhibit rapid phospholipid turnover, which coincides with upregulation of cell proliferation genes. The data also suggest Sonic Hedgehog pathway (SHH) dysregulation may play a role in anaplastic ganglioglioma pathogenesis. That a strong correlation is seen between the HRMAS 1H MRS and genomic data cross-validates and further demonstrates the biological relevance of the MRS results. Our combined metabolic/molecular MRS/genomic approach provides insights into the biology of anaplastic ganglioglioma and a new potential tumor typing methodology that could aid neurologists and neurosurgeons to improve the diagnosis, treatment, and ongoing evaluation of brain tumor patients.

Uncoupling protein 3 expression and intramyocellular lipid accumulation by NMR following local burn trauma.

Burn trauma is a clinical condition accompanied by muscle wasting that severely impedes rehabilitation in burn survivors. Mitochondrial uncoupling protein 3 (UCP3) is uniformly expressed in myoskeletal mitochondria and its expression has been found to increase in other clinical syndromes that, like burn trauma, are associated with muscle wasting (e.g., starvation, fasting, cancer, sepsis). The aim of this study was to explore the effects of burn trauma on UCP3 expression, intramyocellular lipids, and plasma-free fatty acids. Mice were studied at 6 h, 1 d and 3 d after nonlethal hindlimb burn trauma. Intramyocellular lipids in hindlimb skeletal muscle samples collected from burned and normal mice were measured using 1H NMR spectroscopy on a Bruker 14.1 Tesla spectrometer at 4 degrees C. UCP3 mRNA and protein levels were also measured in these samples. Plasma-free fatty acids were measured in burned and normal mice. Local burn trauma was found to result in: 1) upregulation of UCP3 mRNA and protein expression in hindlimb myoskeletal mitochondria by 6 h postburn; 2) increased intramyocellular lipids; and 3) increased plasma-free fatty acids. Our findings show that the increase in UCP3 after burn trauma may be linked to burn-induced alterations in lipid metabolism. Such a link could reveal novel insights into how processes related to energy metabolism are controlled in burn and suggest that induction of UCP3 by burn in skeletal muscle is protective by either activating cellular redox signaling and/or mitochondrial uncoupling.

Synthesis and characterization of poly(DL-lactide)-grafted gelatins as bioabsorbable amphiphilic polymers.

A series of poly(DL-lactide) grafted gelatins, as new bioabsorbable amphiphilic polymers useful in parenteral drug delivery systems and in tissue engineering, were synthesized by the ring opening polymerization of DL-lactide onto a fractionated gelatin with the molecular weight of 1.02 x 10(5). Using tin(II) bis(2-ethylhexanoate) as catalyst, the bulk copolymerization at 140 degrees C and solution copolymerization in dimethylsulfoxide (DMSO) at 80 degrees C were firstly performed in the presence of gelatin. The results showed that the solution copolymerization in DMSO could afford the expected copolymers but the bulk copolymerization would result in an insoluble crosslinked product. The number of grafting sites on gelatin chain could be adjusted by the partial trimethylsilylation of side hydroxy, amino and carboxylic groups in gelatin. The solution copolymerization of DL-lactide on the partially protected gelatin in DMSO was also successful in providing copolymers with different molecular weights. The synthesized copolymers were characterized on the basis of elemental analysis, IR, 1H-NMR and thermal analysis. The IR and 1H-NMR data of these produced copolymers suggested that polylactide branches could be grafted onto gelatin via the side groups such as hydroxyl and amino groups in the solution copolymerization as well as carboxylic groups in bulk copolymerization. The molecular weights of the copolymers could be calculated from the difference of nitrogen contents between a copolymer and free gelatin. The results indicated that molecular weight of the copolymers and those of polylactide branches were increased with the feeding ratio of DL-lactide to gelatin in the copolymerization. However, because of the steric hindrance of some grafting sites on gelatin and the transesterifications of the propagating polylactide branches on gelatin with possibly formed homo-polymeric polylactide chains, the finally formed polylactide branches on gelatin were not very large and the highest average molecular weight of a polylactide branch was not over 4500 in any solution copolymerizations. The results from the thermal analysis of some copolymers, including thermogravimetry and differential scanning calorimetry, showed that the absorbed water in the samples could be lost at a temperature range below 150 degrees C and melting point decreased with increase of polylactide branches in the poly(DL-lactide)-grafted gelatins.