The intelligent-Knife (i-Knife) and its intraoperative diagnostic advantage for the treatment of cervical disease

Clearance of surgical margins in cervical cancer prevents the need for adjuvant chemoradiation and allows fertility preservation. In this study, we determined the capacity of the rapid evaporative ionization mass spectrometry (REIMS), also known as intelligent knife (iKnife), to discriminate between healthy, preinvasive, and invasive cervical tissue. Cervical tissue samples were collected from women with healthy, human papilloma virus (HPV) ± cervical intraepithelial neoplasia (CIN), or cervical cancer. A handheld diathermy device generated surgical aerosol, which was transferred into a mass spectrometer for subsequent chemical analysis. Combination of principal component and linear discriminant analysis and least absolute shrinkage and selection operator was employed to study the spectral differences between groups. Significance of discriminatory m/z features was tested using univariate statistics and tandem MS performed to elucidate the structure of the significant peaks allowing separation of the two classes. We analyzed 87 samples (normal = 16, HPV ± CIN = 50, cancer = 21 patients). The iKnife discriminated with 100% accuracy normal (100%) vs. HPV ± CIN (100%) vs. cancer (100%) when compared to histology as the gold standard. When comparing normal vs. cancer samples, the accuracy was 100% with a sensitivity of 100% (95% CI 83.9 to 100) and specificity 100% (79.4 to 100). Univariate analysis revealed significant MS peaks in the cancer-to-normal separation belonging to various classes of complex lipids. The iKnife discriminates healthy from premalignant and invasive cervical lesions with high accuracy and can improve oncological outcomes and fertility preservation of women treated surgically for cervical cancer. Larger in vivo research cohorts are required to validate these findings

Point-of-care diagnosis of endometrial cancer using the surgical intelligent knife (iknife)-a prospective pilot study of diagnostic accuracy

Introduction: Delays in the diagnosis and treatment of endometrial cancer negatively impact patient survival. The aim of this study was to establish whether rapid evaporative ionisation mass spectrometry using the iKnife can accurately distinguish between normal and malignant endometrial biopsy tissue samples in real time, enabling point-of-care (POC) diagnoses. Methods: Pipelle biopsy samples were obtained from consecutive women needing biopsies for clinical reasons. A Waters G2-XS Xevo Q-Tof mass spectrometer was used in conjunction with a modified handheld diathermy (collectively called the ‘iKnife’). Each tissue sample was processed with diathermy, and the resultant surgical aerosol containing ionic lipid species was then analysed, producing spectra. Principal component analyses and linear discriminant analyses were performed to determine variance in spectral signatures. Leave-one-patient-out cross-validation was used to test the diagnostic accuracy. Results: One hundred and fifty patients provided Pipelle biopsy samples (85 normal, 59 malignant, 4 hyperplasia and 2 insufficient), yielding 453 spectra. The iKnife differentiated between normal and malignant endometrial tissues on the basis of differential phospholipid spectra. Cross-validation revealed a diagnostic accuracy of 89% with sensitivity, specificity, positive predictive value and negative predictive value of 85%, 93%, 94% and 85%, respectively. Conclusions: This study is the first to use the iKnife to identify cancer in endometrial Pipelle biopsy samples. These results are highly encouraging and suggest that the iKnife could be used in the clinic to provide a POC diagnosis

The loss of histone H3 lysine 9 acetylation due to dSAGA-specific dAda2b mutation influences the expression of only a small subset of genes

In Drosophila, the dADA2b-containing dSAGA complex is involved in histone H3 lysine 9 and 14 acetylation. Curiously, although the lysine 9- and 14-acetylated histone H3 levels are drastically reduced in dAda2b mutants, these animals survive until a late developmental stage. To study the molecular consequences of the loss of histone H3 lysine 9 and 14 acetylation, we compared the total messenger ribonucleic acid (mRNA) profiles of wild type and dAda2b mutant animals at two developmental stages. Global gene expression profiling indicates that the loss of dSAGA-specific H3 lysine 9 and 14 acetylation results in the expression change (up- or down-regulation) of a rather small subset of genes and does not cause a general transcription de-regulation. Among the genes up-regulated in dAda2b mutants, particularly high numbers are those which play roles in antimicrobial defense mechanisms. Results of chromatin immunoprecipitation experiments indicate that in dAda2b mutants, the lysine 9-acetylated histone H3 levels are decreased both at dSAGA up- and down-regulated genes. In contrast to that, in the promoters of dSAGA-independent ribosomal protein genes a high level of histone H3K9ac is maintained in dAda2b mutants. Our data suggest that by acetylating H3 at lysine 9, dSAGA modifies Pol II accessibility to specific promoters differently

Endogenous aldehyde accumulation generates genotoxicity and exhaled biomarkers in esophageal adenocarcinoma

Volatile aldehydes are enriched in esophageal adenocarcinoma (EAC) patients’ breath and could improve early diagnosis, however the mechanisms of their production are unknown. Here, we show that weak aldehyde detoxification characterizes EAC, which is sufficient to cause endogenous aldehyde accumulation in vitro. Two aldehyde groups are significantly enriched in EAC biopsies and adjacent tissue: (i) short-chain alkanals, and (ii) medium-chain alkanals, including decanal. The short-chain alkanals form DNA-adducts, which demonstrates genotoxicity and confirms inadequate detoxification. Metformin, a putative aldehyde scavenger, reduces this toxicity. Tissue and breath concentrations of the medium-chain alkanal decanal are correlated, and increased decanal is linked to reduced ALDH3A2 expression, TP53 deletion, and adverse clinical features. Thus, we present a model for increased exhaled aldehydes based on endogenous accumulation from reduced detoxification, which also causes therapeutically actionable genotoxicity. These results support EAC early diagnosis trials using exhaled aldehyde analysis

Pragmatic and rapid analysis of carbonyl, oxidation and chlorination nucleoside-adducts in murine tissue by UPLC-ESI-MS/MS

Nucleoside-adduct analysis by liquid chromatography mass spectrometry is a powerful tool in genotoxicity studies. Efforts to date have quantified an impressive array of DNA damage products, although methodological diversity suggests quantification is still a challenging task. For example, inadequate co-examination of normal nucleosides, cumbersome sample preparation and large DNA requirements were identified to be recurring issues. A six-minute ultra-performance liquid chromatography method is presented which adequately separates seven candidate nucleoside-adducts from the four unmodified nucleosides. The method was sensitive to 1 adduct per 108 normal bases with 20 µg DNA input for most targets. The method was shown to be accurate (81–119% across quintuplets of six tissue types) and precise (relative standard deviation 4–13%). The fast method time facilitated a second quantitation for normal nucleosides at an appropriate dilution, allowing DNA damage concentrations to be contextualised accurately sample-to-sample. From DNA samples, the analytical processing time was < 8 h, and 96 samples can easily be prepared in a day. The method was used to quantify carbonyl, chloro- and oxo- adducts in murine tissue samples

Rapid Evaporative Ionisation Mass Spectrometry (REIMS) provides accurate direct from culture species identification within the genus Candida

Members of the genus Candida, such as C. albicans and C. parapsilosis, are important human pathogens. Other members of this genus, previously believed to carry minimal disease risk, are increasingly recognised as important human pathogens, particularly because of variations in susceptibilities to widely used anti-fungal agents. Thus, rapid and accurate identification of clinical Candida isolates is fundamental in ensuring timely and effective treatments are delivered. Rapid Evaporative Ionisation Mass Spectrometry (REIMS) has previously been shown to provide a high-throughput platform for the rapid and accurate identification of bacterial and fungal isolates. In comparison to commercially available matrix assisted laser desorption ionisation time of flight mass spectrometry (MALDI-ToF), REIMS based methods require no preparative steps nor time-consuming cell extractions. Here, we report on the ability of REIMS-based analysis to rapidly and accurately identify 153 clinical Candida isolates to species level. Both handheld bipolar REIMS and high-throughput REIMS platforms showed high levels of species classification accuracy, with 96% and 100% of isolates classified correctly to species level respectively. In addition, significantly different (FDR corrected P value < 0.05) lipids within the 600 to 1000 m/z mass range were identified, which could act as species-specific biomarkers in complex microbial communities

Validation of ultrasonic harmonic scalpel for real-time tissue identification using rapid evaporative ionization mass spectrometry

In this study, we integrate rapid evaporative ionization mass spectrometry (REIMS) with the Harmonic scalpel, an advanced laparoscopic surgical instrument that utilizes ultrasound energy to dissect and coagulate tissues. It provides unparalleled manipulation capability to surgeons and has superseded traditional electrosurgical tools particularly in abdominal surgery, but is yet to be validated with REIMS. The REIMS platform coupled with the Harmonic device was shown to produce tissue-specific lipid profiles through the analysis of porcine tissues in both negative and positive ionization modes. Comparison with other methods of electrosurgical dissection, such as monopolar electrosurgery and CO2 laser, showed spectral differences in the profile dependent on the energy device used. The Harmonic device demonstrated major spectral differences in the phospholipid region of m/z 600–1000 compared with the monopolar electrosurgical and CO2 laser-generated spectra. Within the Harmonic REIMS spectra, high intensities of diglycerides and triglycerides were observed. In contrast, monopolar electrosurgical and laser spectra demonstrated high abundances of glycerophospholipids. The Harmonic scalpel was able to differentiate between the liver, muscle, colon, and small intestine, demonstrating 100% diagnostic accuracy. The validation of the Harmonic device–mass spectrometry combination will allow the platform to be used safely and robustly for real-time in vivo surgical tissue identification in a variety of clinical applications

Shotgun Lipidomic Profiling of the NCI60 Cell Line Panel Using Rapid Evaporative Ionization Mass Spectrometry.

Rapid evaporative ionization mass spectrometry (REIMS) was used for the rapid mass spectrometric profiling of cancer cell lines. Spectral reproducibility was assessed for three different cell lines, and the extent of interclass differences and intraclass variance was found to allow the identification of these cell lines based on the REIMS data. Subsequently, the NCI60 cell line panel was subjected to REIMS analysis, and the resulting data set was investigated for its distinction of individual cell lines and different tissue types of origin. Information content of REIMS spectral profiles of cell lines were found to be similar to those obtained from mammalian tissues although pronounced differences in relative lipid intensity were observed. Ultimately, REIMS was shown to detect changes in lipid content of cell lines due to mycoplasma infection. The data show that REIMS is an attractive means to study cell lines involving minimal sample preparation and analysis times in the range of seconds