Novel approaches using alkaline or acid/guanidine treatment to eliminate therapeutic antibody interference in the measurement of total target ligand

Measurement of the total target ligand can help to provide pharmacokinetic (PK) and pharmacodynamic (PD) informations. However, the presence of monocloncal antibody therapeutics (ThAs) interferes with ELISA determinations of the total target proteins. The interferences can cause over- or under-estimation of the target protein analysis. The nature of interferences was dependent upon the ThA, target protein, antibody reagents and assay conditions of the ELISA. We have developed novel alkaline and acid/guanidine treatment approaches to dissociate the protein binding and preferentially denature the ThA. The neutralized target proteins can be determined by ELISA. These methods provide reproducible measurements of total target protein without ThA interference. Serum samples, standards and QCs containing target protein and ThA were treated with alkaline buffer (pH > 13) containing casein or acid/guanidine buffer (pH < 1). Total target proteins for two different ThA systems were successfully measured and interferences were completely eliminated by the treatments. These methods were successfully applied to analysis in pre-clinical serum samples.     

Mesothelin, a novel immunotherapy target for triple negative breast cancer

Mesothelin is a cell-surface glycoprotein present on mesothelial cells and elicits T cell responses in a variety of cancers including pancreatic, biliary and ovarian cancer. Breast cancer is not known to express mesothelin. We postulated that mesothelin may be a unique tumor-associated antigen in triple negative breast cancer (TNBC), a less common breast cancer subtype which may have been under-represented in prior studies that characterized mesothelin expression. Therefore, we screened 99 primary breast cancer samples by immunohistochemistry analysis using formalin-fixed paraffin-embedded archival tumor tissues and confirmed that mesothelin was overexpressed in the majority of TNBC (67 %) but only rarely in <5 % ER(+) or Her2-neu(+) breast cancer, respectively. To determine whether mesothelin may be exploited as a novel immunotherapy target in breast cancer, an in vitro cell killing assay was performed to compare the ability of genetically modified T cells expressing a chimeric antibody receptor (CAR) specific for mesothelin (mesoCAR T cells) or non-transduced T cells to kill mesothelin-expressing primary breast cancer cells. A significantly higher anti-tumor cytotoxicity by mesoCAR T cells was observed (31.7 vs. 8.7 %, p < 0.001). Our results suggest that mesothelin has promise as a novel immunotherapy target for TNBC for which effective targeted therapy is lacking to date.     

Diagnostic values of GHSR DNA methylation pattern in breast cancer

DNA methylation patterns have been recognised as cancer-specific markers with high potential for clinical applications. We aimed at identifying methylation variations that differentiate between breast cancers and other breast tissue entities to establish a signature for diagnosis. Candidate genomic loci were analysed in 117 fresh-frozen breast specimens, which included cancer, benign and normal breast tissues from patients as well as material from healthy individuals. A cancer-specific DNA methylation signature was identified by microarray analysis in a test set of samples (n?=?52, p?0.99). To date, this represents the DNA methylation marker of the highest sensitivity and specificity for breast cancer diagnosis. Functionally, ectopic expression of GHSR in a cell line model reduced breast cancer cell invasion without affecting cell viability upon stimulation of cells with ghrelin. Our data suggest a link between epigenetic down-regulation of GHSR and breast cancer cell invasion.     

Visual spatial integrity in the absence of splenium

Several lines of evidence have confirmed that visual integrity among the two hemifields is severely impaired following transection of posterior part of the corpus callosum (splenium). But the extent of this impairment differs for various visual functions. Here, in a posterior callosectomized patient (MD), interhemispheric visual spatial integrity is investigated in three experiments. In the first experiment, MD was asked to make decision about horizontal alignment of two balls presented simultaneously in one of three conditions: both in right or left hemifield, or each in one hemifield. We have previously shown that she is not able to perform a shape-matching task for stimuli presented to different hemifields. But in this task, MD was able to compare location of the two bilaterally presented stimuli significantly above chance level.Then we investigated whether attentive visual object tracking across vertical meridian of the visual field is possible in the absence of splenium. MD had to attentively track one bouncing ball among three identical balls, while it crossed the vertical midline in half of trials. Her performance in crossed conditions was significantly above chance level, but it was lower than uncrossed conditions. Finally, we investigated the contribution of simple interhemispheric temporal signals in performing the attentive tracking task. Results suggest that the patient was not using such temporal signals. Our results suggest that interhemispheric connections other than splenium can contribute in making an integral visual map across hemifields. Such an integrated map can be used for bilateral visual spatial comparisons and visual spatial attention.     

Non-invasive brain mapping of motor-related areas of four limbs in patients with clinically isolated syndrome compared to healthy normal controls

Functional MRI studies on patients with multiple sclerosis (MS) have demonstrated widespread cortical reorganization of the motor network. However, few functional studies have addressed cortical plasticity in patients with clinically isolated syndrome (CIS). The activity of the lower limb motor system, despite its highlighted involvement in patients with CIS and MS, has been little studied. Thus, brain activation was compared in CIS patients with clinically intact motor systems with that in healthy control participants while they were performing motor tasks with four limbs. A total of 26 right-handed patients with CIS with clinically intact motor systems and 28 right-handed age and sex-matched controls participated in the functional MRI (fMRI) motor task. Patients with CIS showed greater activation in the ipsilateral secondary somatosensory cortex, cingulate gyrus and precuneus cortex while performing the ankle movement task compared to healthy controls. In the finger-tapping task, patients with CIS showed greater activity in the contralateral thalamus, ipsilateral premotor and superior temporal gyrus. In addition, the left inferior frontal gyrus was activated more in patients with CIS, regardless of the hand used. Therefore, despite having clinically intact motor systems, patients with CIS had different motor networks. All novel recruited regions were adjacent to the somatotopy of the primary motor areas of the limbs. Our finding confirm that brain reorganization precedes clinical manifestation, as no patient had any clinical manifestation that suggested involvement of the motor system.     

Effects of silicone gel on burn scars

Aim

To study the efficacy of silicone gel applied to hypertrophic burn scars, in reducing scar interference with normal function and improving cosmesis.

Methods

A randomised, double-blind, placebo-controlled trial involving 38 people with hypertrophic burn scars. Each scar was divided into two segments; silicone gel sheet was applied randomly to one of the two and placebo to the other. Participants were seen again after 1 and 4 months. Their data and wound characteristics were collected using the Vancouver scar scale.

Results

The median age of participants was 22 years (1.5–60 years) and 16 were male; 4 did not attend follow-up and were excluded from the study. There were no significant differences in baseline characteristics. Although after 1 month all scar scale measures were lower in treated areas, only the vascularity scale was significantly different between the two areas. After 4 months, all scale measures were significantly lower in the silicone gel group than in the control group, except for the pain score.

Conclusion

Silicone gel is an effective treatment for hypertrophic burn scars.     

Robotic Surgical Education: a Collaborative Approach to Training Postgraduate Urologists and Endourology Fellows

Objective:

Currently, robotic training for inexperienced, practicing surgeons is primarily done vis-à-vis industry and/or society-sponsored day or weekend courses, with limited proctorship opportunities. The objective of this study was to assess the impact of an extended-proctorship program at up to 32 months of follow-up.

Methods:

An extended-proctorship program for robotic-assisted laparoscopic radical prostatectomy was established at our institution. The curriculum consisted of 3 phases: (1) completing an Intuitive Surgical 2-day robotic training course with company representatives; (2) serving as assistant to a trained proctor on 5 to 6 cases; and (3) performing proctored cases up to 1 year until confidence was achieved. Participants were surveyed and asked to evaluate on a 5-point Likert scale their operative experience in robotics and satisfaction regarding their training

Results:

Nine of 9 participants are currently performing robotic-assisted laparoscopic radical prostatectomy (RALP) independently. Graduates of our program have performed 477 RALP cases. The mean number of cases performed within phase 3 was 20.1 (range, 5 to 40) prior to independent practice. The program received a rating of 4.2/5 for effectiveness in teaching robotic surgery skills.

Conclusion:

Our robotic program, with extended proctoring, has led to an outstanding take-rate for disseminating robotic skills in a metropolitan community.