Raman chemometric urinalysis (Rametrix) as a screen for bladder cancer.

Bladder cancer (BCA) is relatively common and potentially recurrent/progressive disease. It is also costly to detect, treat, and control. Definitive diagnosis is made by examination of urine sediment, imaging, direct visualization (cystoscopy), and invasive biopsy of suspect bladder lesions. There are currently no widely-used BCA-specific biomarker urine screening tests for early BCA or for following patients during/after therapy. Urine metabolomic screening for biomarkers is costly and generally unavailable for clinical use. In response, we developed Raman spectroscopy-based chemometric urinalysis (Rametrix™) as a direct liquid urine screening method for detecting complex molecular signatures in urine associated with BCA and other genitourinary tract pathologies. In particular, the RametrixTM screen used principal components (PCs) of urine Raman spectra to build discriminant analysis models that indicate the presence/absence of disease. The number of PCs included was varied, and all models were cross-validated by leave-one-out analysis. In Study 1 reported here, we tested the Rametrix™ screen using urine specimens from 56 consented patients from a urology clinic. This proof-of-concept study contained 17 urine specimens with active BCA (BCA-positive), 32 urine specimens from patients with other genitourinary tract pathologies, seven specimens from healthy patients, and the urinalysis control SurineTM. Using a model built with 22 PCs, BCA was detected with 80.4% accuracy, 82.4% sensitivity, 79.5% specificity, 63.6% positive predictive value (PPV), and 91.2% negative predictive value (NPV). Based on the number of PCs included, we found the RametrixTM screen could be fine-tuned for either high sensitivity or specificity. In other studies reported here, RametrixTM was also able to differentiate between urine specimens from patients with BCA and other genitourinary pathologies and those obtained from patients with end-stage kidney disease (ESKD). While larger studies are needed to improve RametrixTM models and demonstrate clinical relevance, this study demonstrates the ability of the RametrixTM screen to differentiate urine of BCA-positive patients. Molecular signature variances in the urine metabolome of BCA patients included changes in: phosphatidylinositol, nucleic acids, protein (particularly collagen), aromatic amino acids, and carotenoids.

Spectral characteristics of urine from patients with end-stage kidney disease analyzed using Raman Chemometric Urinalysis (Rametrix).

Raman Chemometric Urinalysis (RametrixTM) was used to discern differences in Raman spectra from (i) 362 urine specimens from patients receiving peritoneal dialysis (PD) therapy for end-stage kidney disease (ESKD), (ii) 395 spent dialysate specimens from those PD therapies, and (iii) 235 urine specimens from healthy human volunteers. RametrixTM analysis includes spectral processing (e.g., truncation, baselining, and vector normalization); principal component analysis (PCA); statistical analyses (ANOVA and pairwise comparisons); discriminant analysis of principal components (DAPC); and testing DAPC models using a leave-one-out build/test validation procedure. Results showed distinct and statistically significant differences between the three types of specimens mentioned above. Further, when introducing "unknown" specimens, RametrixTM was able to identify the type of specimen (as PD patient urine or spent dialysate) with better than 98% accuracy, sensitivity, and specificity. RametrixTM was able to identify "unknown" urine specimens as from PD patients or healthy human volunteers with better than 96% accuracy (with better than 97% sensitivity and 94% specificity). This demonstrates that an entire Raman spectrum of a urine or spent dialysate specimen can be used to determine its identity or the presence of ESKD by the donor.

Dipstick analysis of urine chemistry: benefits and limitations of dry chemistry-based assays.

Urinalysis is a commonly utilized laboratory test, and analysis of urine has been studied and used since ancient times. Urine contains a wide array of metabolites that can provide information regarding the current physiologic state of the body and clinical manifestations of disease. In this review, we discuss the mechanics of the dry chemistry component of the urine dipstick such as the reaction principles underlying various assays and potential effects of collection and storage on results. Additionally, we discuss the benefits and limitations of the urine dipstick as it pertains to its use as a low-cost tool in point-of-care settings and the reasoning for a lack of its use as a broad screening tool.

Spectral characteristics of urine specimens from healthy human volunteers analyzed using Raman chemometric urinalysis (Rametrix).

Raman chemometric urinalysis (Rametrix™) was used to analyze 235 urine specimens from healthy individuals. The purpose of this study was to establish the "range of normal" for Raman spectra of urine specimens from healthy individuals. Ultimately, spectra falling outside of this range will be correlated with kidney and urinary tract disease. Rametrix™ analysis includes direct comparisons of Raman spectra but also principal component analysis (PCA), discriminant analysis of principal components (DAPC) models, multivariate statistics, and it is available through GitHub as the Rametrix™ LITE Toolbox for MATLAB®. Results showed consistently overlapping Raman spectra of urine specimens with significantly larger variances in Raman shifts, found by PCA, corresponding to urea, creatinine, and glucose concentrations. A 2-way ANOVA test found that age of the urine specimen donor was statistically significant (p < 0.001) and donor sex (female or male identification) was less so (p = 0.0526). With DAPC models and blind leave-one-out build/test routines using the Rametrix™ PRO Toolbox (also available through GitHub), an accuracy of 71% (sensitivity = 72%; specificity = 70%) was obtained when predicting whether a urine specimen from a healthy unknown individual was from a female or male donor. Finally, from female and male donors (n = 4) who contributed first morning void urine specimens each day for 30 days, the co-occurrence of menstruation was found statistically insignificant to Rametrix™ results (p = 0.695). In addition, Rametrix™ PRO was able to link urine specimens with the individual donor with an average of 78% accuracy. Taken together, this study established the range of Raman spectra that could be expected when obtaining urine specimens from healthy individuals and analyzed by Rametrix™ and provides the methodology for linking results with donor characteristics.