Inducible HSP70 (HSP70i) is implicated in the development of autoimmune vitiligo and is overexpressed in melanoma. We generated an expression construct encoding a single amino acid modification to the dendritic cell (DC) activating moiety (HSP70i Q435A ). A topical application of the DNA construct has now demonstrated efficacy in reversing autoimmunity in Sinclair swine without obstructing anti-melanoma responses. We postulate that treatment with the HSP70i Q435A construct elicits a humoral response to stress protein selectively expressed on the surface of melanoma cells, providing measurable anti-tumor protection. To test this, we evaluated humoral responses to the protein in mice to measure their specificity and anti-tumor activity. Biolistic delivery of HSP70i Q435A or empty vector DNA to C57BL/6 mice was provided 5 times, every 6 days before a subcutaneous B16F10 tumor challenge. Melanoma growth was monitored and serum was collected at end point to determine antibody isotype/subclass titers by ELISA, measure anti-HSP70-mediated ADCC via an ELISAbased natural kill cell (NK) degranulation assay and perform epitope scanning analysis to detect binding epitopes to anti-HSP70 serum antibodies. Tumor growth was delayed in HSP70i Q435A -treated mice with measurable anti-HSP70i IgG and IgM titers found exclusively in HSP70i Q435A -treated mice. A consensus C-terminal peptide sequence was identified via epitope mapping, and significant degranulation of NK cells was observed in response to anti-HSP70 serum antibodies. Thus, topical application of HSP70i Q435A DNA might offer a safe treatment for the autoimmune side effects of immunotherapy for melanoma.
Background: Current guidelines suggest the use of three-dimensional left ventricular ejection fraction (3DLVEF) to assess left ventricular function, with its use favoured over Simpson's biplane method. It is also recommended that this value be cross checked with a visual estimation of left ventricular ejection fraction (LVEF). Aims: We sought to examine the accuracy of visual LVEF estimation by two experienced sonographers (SH, KS), compared to the gold standard obtained by 3DLVEF. Methods: A total of 363 consecutive patients had echocardiograms performed for a clinically indicated reason. All echocardiograms included a full volume threedimensional image acquisition from the apical windows. Both sonographers sequentially assessed visual LVEF first, then 3DLVEF on all cases in a blinded fashion. All measurements were performed using the AutoLVQ package on an EchoPac workstation. Results: Patient characteristics included age 57 ± 17 years, BMI 27.6 ± 5.9 kg/m 2 , 58% males and Simpsons biplane LVEF 58.3 ± 12.0% (range 15-87%). 3DLVEF was not considered feasible in 20% of patients and these cases were excluded from further analysis. There was an excellent correlation between visual LVEF between the two sonographers, with a Pearson's correlation coefficient r = 0.96 (95% CI 0.95-0.97), Bland Altman mean bias 0.2% (95% limits of agreement 6.9 to −7.4%). This was superior to the correlation between 3DLVEF between the two sonographers: r = 0.84 (95% CI 0.80-0.87), and between visual LVEF and 3DLVEF by each sonographer (KS: r = 0.83 [95%
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