Page 70 - JCTR-11-3

Journal of Clinical and
Translational Research N-NOSE: Early cervical cancer screening

effects of adaptation – was spotted at four locations on the 3. Results
plate (for the urine side and non-urine side). Subsequently,
1 μL of urine sample, diluted 10-fold and 100-fold with 3.1. Sensitivity of N-NOSE in detecting cervical
ultrapure water, was added to two spots on the urine side. cancer
We confirmed that nematodes showed no chemotaxis Early identification of cervical carcinogenesis is crucial
behavior to 1 μL of water (data not shown). for reducing the mortality risk associated with cervical
2,29
Approximately 100 adult nematodes were collected and cancer. Therefore, our investigation focused on
washed three times using chemotaxis buffer – prepared evaluating the capability of our N-NOSE technology to
using 0.05% gelatin, 5 mM potassium phosphate (made by detect CIN, an early-stage pre-cursor to cervical cancer,
combining appropriate amounts of monobasic potassium with high sensitivity. While the absence of a concurrent
phosphate and dipotassium phosphate in ultrapure water), control group is a limitation, the specificity of the
1 mM calcium chloride, and 1 mM magnesium sulfate. The N-NOSE test has been previously established in large-
nematodes were then placed at the center of the plate. After scale studies involving healthy individuals, demonstrating
removing the excess buffer, the nematodes were allowed to high specificity rates of 90% and 95% using the same
19,22
roam freely for 30 min. cutoff values and methodologies applied in this study.
This methodological consistency allows us to infer the
The chemotaxis index was calculated 19,24,28 using the specificity of the test within our study population. Our
following equation: primary objective was to assess the test’s ability to identify
true positive cases within a diseased cohort correctly. The
(A B)− (1)
Chemotaxis index = high sensitivity observed, particularly the 100% sensitivity
+
(A B) for detecting CIN3 lesions, underscores the test’s potential
Where (A) is the number of nematodes near the urine clinical utility in the early detection and intervention of
samples, and (B) is the number of nematodes in the region cervical carcinogenesis. Early identification of CIN is
without the urine samples. critical to prevent the progression of invasive cervical
cancer, and our findings support the role of N-NOSE as a
For each sample, chemotaxis assays were performed non-invasive and sensitive screening tool.
using both 10-fold and 100-fold urine dilutions (n = 10
for each dilution). A negative chemotaxis index (ranging In this study, we employed the same cutoff values as those
from −1 to 0) indicates repulsion to the sample, while a used in our previous clinical investigations, as illustrated
positive chemotaxis index (ranging from 0 to 1) indicates in Figure 2, to ensure consistency and comparability. This
attraction to the sample. 19,24 A chemotaxis index was methodological alignment allows us to infer specificity
considered “positive” if it was positive in at least one of the based on established data while focusing on evaluating
two dilutions. 22,24 sensitivity within the present patient cohort. A “negative”
result is defined as nematode repulsion to both 10- and
2.4. Statistical analyses 100-fold diluted urine samples, while a “positive” result
One-way analysis of variance was used to calculate indicates nematode attraction to at least one of the diluted
the correlation coefficients and/or p-values between urine samples. In our previous studies, specificity rates of
the N-NOSE index at both 10-fold and 100-fold urine 90% and 95% were confirmed using the same cutoff values
19,22
dilutions and variables such as age and each biochemical in two independent populations of healthy individuals.
blood test (hepatic alanine transaminase, hepatic Conducting the N-NOSE chemotaxis assays under the
aspartate aminotransferase, renal blood urea nitrogen, same conditions as we did in our previous studies allows us
or renal creatinine). Statistically significant differences to maintain consistency and comparability with our earlier
in the N-NOSE index for both urine dilutions (10- findings while acknowledging the inherent constraints
fold and 100-fold) between cervical cancer patients posed by the study design.
and CIN cases were evaluated using Welch’s t-test. The The test was conducted on urine samples from 74 female
Wilcoxon signed-rank test was employed to calculate patients with cervical cancer and 245 female patients with
the p-values between the N-NOSE index at both urine CIN. Statistical analysis revealed no significant correlation
dilutions (10-fold and 100-fold) and each urine general between the results of N-NOSE and age, with correlation
qualitative test (urinary glucose, urinary ketone bodies, coefficients of -0.04166 for 10-fold dilution (p=0.4585)
urinary proteins, or urinary occult blood), respectively. and -0.00517 for 100-fold dilution (p=0.9267). Among the
All statistical analyses were performed using JMP 14 cervical cancer patients, 53 out of 74 patients (71.6%) tested
®
software (SAS Institute, USA). A p<0.05 was considered positive using the N-NOSE test. Stage-specific sensitivities
statistically significant. of cervical cancer using the N-NOSE test, based on the

Volume 11 Issue 3 (2025) 64 doi: 10.36922/jctr.24.00080