URINE-BASED BIOMARKERS FOR EARLY CANCER DETECTION AND RELATED METHODS

Abstract

ABSTRACT A system and method for early cancer detection using a urine cancer detector strip are described. The system includes a detection module that utilizes colorimetry and fluorescence dual mode detection, assisted by duplex specific nuclease (DSN) and gold nanoparticle-based amplification, to detect miRNA-21 in urine samples. The detection process involves immobilizing FAM-labelled hairpin probes on gold nanoparticles, quenching fluorescence, capturing target miRNA-21, converting it into a DNA/RNA heteroduplex, and recognizing and digesting the heteroduplex with DSN. The presence of miRNA-21 is indicated by a color change from red to bluish gray. This non-invasive and cost-effective device aims to facilitate earlier diagnosis and treatment of cancer. Reference Fig 1

Specification

Description:TECHNICAL FIELD
[0001] The present invention is related to non-invasive cancer detection.
BACKGROUND
[0002] Cancer remains one of the leading causes of mortality worldwide, with early detection being crucial for improving patient outcomes. Traditional methods of cancer detection, such as imaging tests, biopsies, and blood tests, often have limitations in terms of invasiveness, cost, and the ability to detect cancer at an early stage. These methods may not always provide reliable results, especially in the early stages of cancer when treatment can be most effective. As a result, there is a significant need for non-invasive, cost-effective, and reliable diagnostic tools that can detect cancer early, thereby improving the chances of successful treatment and survival.

[0003] Urine-based biomarkers have emerged as a promising solution in the field of cancer diagnostics. Urine, being a non-invasive sample, offers a convenient and accessible medium for detecting biomarkers associated with cancer. Among these biomarkers, microRNAs (miRNAs) have gained attention due to their role in regulating gene expression and their association with cancer development and progression. Aberrant expression of specific miRNAs, such as miRNA-21, has been linked to various types of cancer, making them potential candidates for early detection. The development of sensitive and specific detection methods for these miRNAs in urine samples could revolutionize cancer diagnostics by providing a simple and effective means of early detection.
SUMMARY
[0004] In accordance with embodiments, a system for early cancer detection is provided, comprising a urine cancer detector strip that includes a sample pad, a nitrocellulose membrane, an amplification segment, and a plastic cassette. The system further includes a detection module configured to detect the presence of miRNA-21 in a urine sample using colorimetry and fluorescence dual mode detection. This detection is assisted with duplex specific nuclease (DSN) and gold nanoparticle-based amplification. The detection module comprises FAM-labelled hairpin probes (FAM-HPs) immobilized on gold nanoparticles (AuNPs), a mechanism for quenching the fluorescence of the FAM-HPs by the AuNPs, a capture mechanism for capturing the target miRNA-21 by the HP-AuNPs probe, a conversion mechanism for converting the captured miRNA-21 into a DNA/RNA heteroduplex, a recognition and digestion mechanism for specifically recognizing and digesting the DNA/RNA heteroduplex by the DSN, and a binding mechanism for further binding the released target miRNA-21 to other HP-AuNPs under the assistance of DSN. The system also includes a result section configured to display a color change indicating the presence of miRNA-21 in the urine sample, with the color change being from red to bluish gray.

[0005] In accordance with other embodiments, the system is designed such that miRNA-21 serves as a potential biomarker for early cancer detection, being upregulated in many types of cancer. The DSN is highlighted as a promising tool for improving the amplification of miRNA due to its unique selective cutting ability.

[0006] In yet other embodiments, the urine cancer detector strip is described as a non-invasive and cost-effective device for early cancer detection, facilitating earlier diagnosis and treatment of cancer. The amplification segment is specifically noted as the location where the actual amplification of miRNA-21 occurs if it is present in the urine sample.

[0007] In accordance with further embodiments, a method for early cancer detection is provided. This method involves providing a urine cancer detector strip comprising a sample pad, a nitrocellulose membrane, an amplification segment, and a plastic cassette. A urine sample is received on the sample pad and moved to the amplification segment for miRNA amplification, where the actual amplification of miRNA-21 occurs if present. The presence of miRNA-21 is detected using colorimetry and fluorescence dual mode detection assisted with DSN and gold nanoparticle-based amplification. This detection process involves immobilizing FAM-labelled hairpin probes (FAM-HPs) on gold nanoparticles (AuNPs), quenching the fluorescence of the FAM-HPs by the AuNPs, capturing the target miRNA-21 by the HP-AuNPs probe, converting the captured miRNA-21 into a DNA/RNA heteroduplex, specifically recognizing and digesting the DNA/RNA heteroduplex by the DSN, and further binding the released target miRNA-21 to other HP-AuNPs under the assistance of DSN. A color change indicating the presence of miRNA-21 in the urine sample is displayed, with the color change being from red to bluish gray.

[0008] In accordance with additional embodiments, the method emphasizes that miRNA-21 is a potential biomarker for early cancer detection, being upregulated in many types of cancer. The DSN is again noted as a promising tool for improving the amplification of miRNA due to its unique selective cutting ability. The urine cancer detector strip is reiterated as a non-invasive and cost-effective device for early cancer detection, allowing for earlier diagnosis and treatment of cancer. The method also includes a result section configured to display the color change indicating the presence of miRNA-21 in the urine sample.
BRIEF DESCRIPTION OF THE DRAWINGS

[0009] FIG. 1 illustrates a Proto-type of Cancer detection strip
[0010] FIG. 2 illustrates components of a urine cancer detection system, including detection, result, and amplification modules.
[0011] FIG. 3 illustrates a flowchart for detecting miRNA-21 in urine using a colorimetric strip.
DETAILED DESCRIPTION

[0012] Urine-based biomarkers are emerging as a promising non-invasive and cost-effective approach for early cancer detection. This technology leverages the detection of specific microRNAs (miRNAs) in urine samples, which are small, non-coding RNA molecules that regulate gene expression. miRNAs have been found to play a significant role in cancer development and progression, with aberrant expression detected in various cancers, including bladder, prostate, and breast cancer. Among these, miRNA-21 is particularly noteworthy as it is upregulated in many types of cancer, making it a potential biomarker for early detection. The detection of miRNA-21 is achieved through a dual-mode system utilizing colorimetry and fluorescence, assisted by duplex specific nuclease (DSN) and gold nanoparticle-based amplification. DSN is a tool that enhances miRNA amplification due to its selective cutting ability, cleaving only DNA sequences in double-stranded DNA or DNA/RNA heteroduplexes. This specificity allows the miRNA to remain intact for repeated cyclic reactions, facilitating signal amplification. The urine cancer detector strip, a key component of this system, includes a sample pad, nitrocellulose membrane, amplification segment, and plastic cassette. The sample pad receives the urine sample, which is then transferred to the amplification segment where miRNA-21, if present, undergoes amplification. The detection module employs FAM-labelled hairpin probes immobilized on gold nanoparticles, with fluorescence quenched by the nanoparticles. Upon capturing the target miRNA-21, the probe converts it into a DNA/RNA heteroduplex, which is digested by DSN, allowing further binding and amplification. The result section of the strip displays a color change from red to bluish gray, indicating the presence of miRNA-21. This system offers a non-invasive, cost-effective method for early cancer detection, with potential applications in clinical diagnostics, companion diagnostics, and research tools.

[0013] FIG. 1 describes the proto-type of urine-based cancer detection strip which comprises several key components that work together to detect miRNA-21, a cancer biomarker. It begins with a sample pad, where the urine sample is placed. From there, the sample passes through an absorption pad that ensures smooth flow and filtration of the sample. The central component of the strip is the amplification section, which contains hairpin probes (FAM-HP) immobilized on gold nanoparticles (AuNPs). In the presence of miRNA-21, these probes capture the biomarker, forming a DNA/RNA hybrid that is cleaved by duplex-specific nuclease (DSN) to amplify the signal. The detection process is facilitated by magnesium chloride, which ensures the proper chemical environment for the reaction. A color change from red to bluish-gray signals a positive detection of miRNA-21. All of these elements are housed on a nitrocellulose membrane, encased in a protective plastic cassette for structural integrity.

[0014] FIG. 2 illustrates the detection system 200, which may be responsible for detecting miRNA-21 using colorimetry and fluorescence.

[0015] The detection module 202 may employ a dual-mode detection system that utilizes duplex specific nuclease (DSN) and gold nanoparticle-based amplification. The detection process may begin with the immobilization of FAM-labelled hairpin probes (FAM-HPs) on gold nanoparticles (AuNPs). The fluorescence of the FAM-HPs may be quenched by the AuNPs, which may prepare the system for the detection of miRNA-21. Upon introduction of the target miRNA-21 from the urine sample, the HP-AuNPs probe may capture the target miRNA-21 and convert it into a DNA/RNA heteroduplex. This heteroduplex may be specifically recognized and digested by the DSN, allowing the released target miRNA-21 to bind to other HP-AuNPs with the assistance of DSN. This cyclic reaction may enable signal amplification, which may be crucial for the detection of miRNA-21. The detection module may thus provide a non-invasive and cost-effective method for early cancer detection by utilizing the unique properties of DSN and gold nanoparticles. The detection module may be an integral part of the urine cancer detector strip, which may facilitate the early diagnosis and treatment of cancer by indicating the presence of miRNA-21 through a color change from red to bluish gray.

[0016] The result section, identified as 204, may serve a role in the system for early cancer detection by displaying a color change that indicates the presence of miRNA-21. This component may be integral to the detection module, which utilizes colorimetry and fluorescence dual mode detection. The result section may be designed to visually communicate the detection of miRNA-21 by exhibiting a color transition from red to bluish gray. This color change may be facilitated by the interaction of the detection module's components, including the FAM-labelled hairpin probes immobilized on gold nanoparticles and the duplex specific nuclease (DSN), which may assist in the amplification and detection process. The result section may thus provide a clear and immediate indication of the presence of miRNA-21, which is a potential biomarker for early cancer detection. The system may leverage the non-invasive and cost-effective nature of the urine cancer detector strip, allowing for earlier diagnosis and treatment of cancer. The result section's ability to display the color change may be crucial for the user to interpret the test results accurately, thereby enhancing the system's utility in clinical and non-clinical settings.

[0017] The amplification segment, identified as 206, may serve as the site where the actual amplification of miRNA-21 occurs. This segment may be integral to the system's function, as it may facilitate the amplification process necessary for detecting miRNA-21, a potential biomarker for early cancer detection. The amplification segment may work in conjunction with the detection module, which may utilize colorimetry and fluorescence dual mode detection, assisted by duplex specific nuclease (DSN) and gold nanoparticle-based amplification. The amplification segment may receive the urine sample, which may be moved from the sample pad, and may enable the amplification of miRNA-21 if present. This process may involve the immobilization of FAM-labelled hairpin probes on gold nanoparticles, which may be quenched by the AuNPs. Upon introduction of the target miRNA-21, the HP-AuNPs probe may capture the target miRNA and convert it into a DNA/RNA heteroduplex. This heteroduplex may then be specifically recognized and digested by DSN, allowing the released target RNA to further bind to other HP-AuNPs under DSN assistance. The amplification segment may thus play a pivotal role in the amplification of miRNA-21, enabling the subsequent detection and indication of its presence through a color change in the result section. This color change, from red to bluish gray, may indicate the presence of miRNA-21, thereby contributing to the system's capability for early cancer detection. The amplification segment may thus be essential for the system's operation, facilitating the amplification and detection processes that are central to the system's function.

[0018] FIG. 3 is a flowchart illustrating a method 300 for providing a urine cancer detector strip, according to an embodiment.

[0019] The urine sample may be received on the sample pad, which is a component of the urine cancer detector strip. At step 302, the sample pad may serve as the initial point of contact for the urine sample, facilitating its collection and subsequent processing. The sample pad may be designed to efficiently absorb and hold the urine sample, ensuring that it is adequately prepared for further analysis. This step may involve the integration of the sample pad with other components, such as the nitrocellulose membrane and plastic cassette, to form a cohesive unit that supports the overall functionality of the system. The sample pad may be configured to work in conjunction with the nitrocellulose membrane, allowing the urine sample to be effectively transferred to the amplification segment. At step 304, this transfer may be essential for the amplification of miRNA-21, a potential biomarker for early cancer detection. The sample pad's role in receiving and processing the urine sample may be crucial for the accurate detection of miRNA-21, as it sets the stage for the subsequent steps in the detection process. The design and material properties of the sample pad may be optimized to ensure compatibility with the detection module, which utilizes colorimetry and fluorescence dual mode detection. This compatibility may enhance the system's ability to detect miRNA-21 with high sensitivity and specificity. The sample pad's integration into the urine cancer detector strip may also contribute to the system's non-invasive and cost-effective nature, making it a viable option for early cancer detection.

[0020] The urine sample may be moved to the amplification segment for miRNA amplification. At step 304, this step may involve the transition of the urine sample from the initial collection point to the designated amplification area within the system. The amplification segment may serve as the zone where the miRNA-21, if present, can undergo amplification. This process may be facilitated by the amplification segment, which is designed to enhance the detection capabilities of the system. The movement of the urine sample to this segment may be crucial for ensuring that the miRNA-21 is adequately amplified, thereby increasing the likelihood of its detection. The amplification segment may be equipped with the necessary components to support this amplification process, potentially involving the interaction with other system elements such as the detection module. This step may be integral to the overall functionality of the system, as it may set the stage for subsequent detection and analysis of the miRNA-21, contributing to the system's ability to provide early cancer detection.

[0021] In the context of detecting the presence of miRNA-21 in a urine sample, the detection module may utilize colorimetry and fluorescence dual mode detection, which is assisted by duplex specific nuclease (DSN) and gold nanoparticle-based amplification. In step 306, the process may begin with the immobilization of FAM-labelled hairpin probes (FAM-HPs) on gold nanoparticles (AuNPs). This immobilization may serve as a preparatory step for detection, where the fluorescence of the FAM-HPs may be quenched by the AuNPs. The quenching mechanism may facilitate the subsequent capture of the target miRNA-21 by the HP-AuNPs probe. Upon capturing the target miRNA-21, the probe may convert it into a DNA/RNA heteroduplex. This conversion may be crucial for the specific recognition and digestion of the heteroduplex by the DSN, which may enhance the amplification process due to DSN's unique selective cutting ability. Following digestion, the released target miRNA-21 may further bind to other HP-AuNPs under the assistance of DSN, potentially allowing for repeated cycles of amplification. This cyclic reaction may contribute to the overall sensitivity and specificity of the detection process. The detection module, therefore, may play a pivotal role in the early detection of cancer by identifying miRNA-21, a potential biomarker, through a non-invasive and cost-effective method. The integration of these components and actions may provide a comprehensive approach to detecting miRNA-21, leveraging the advantages of both colorimetry and fluorescence in a dual mode detection system.

[0022] The result section may display a color change to indicate the presence of miRNA-21 in the urine sample. At step 308, this color change may transition from red to bluish gray, suggesting the detection of miRNA-21. The miRNA-21 may serve as a potential biomarker for early cancer detection, as it is often upregulated in various types of cancer. The DSN may be utilized as a promising tool to enhance the amplification of miRNA due to its unique selective cutting ability. The urine cancer detector strip may be characterized as a non-invasive and cost-effective device, potentially allowing for earlier diagnosis and treatment of cancer. The result section of the strip may be configured to display the color change, thereby indicating the presence of miRNA-21 in the urine sample. This process may involve the use of colorimetry and fluorescence dual mode detection, assisted by duplex specific nuclease and gold nanoparticle-based amplification, to achieve the desired result. The integration of these components and mechanisms may facilitate the effective detection of miRNA-21, contributing to the overall functionality of the system for early cancer detection.









, Claims:CLAIMS

We claim:
1. A system for early cancer detection, comprising:
a urine cancer detector strip comprising a sample pad, a nitrocellulose membrane, an amplification segment, and a plastic cassette;
a detection module configured to detect the presence of miRNA-21 in a urine sample using colorimetry and fluorescence dual mode detection assisted with duplex specific nuclease (DSN) and gold nanoparticle-based amplification, wherein the detection module
comprises:
FAM-labelled hairpin probes (FAM-HPs) immobilized on gold nanoparticles (AuNPs),
a mechanism for quenching the fluorescence of the FAM-HPs by the AuNPs,
a capture mechanism for capturing the target miRNA-21 by the HP-AuNPs probe,
a conversion mechanism for converting the captured miRNA-21 into a DNA/RNA heteroduplex,
a recognition and digestion mechanism for specifically recognizing and digesting the DNA/RNA heteroduplex by the DSN, and
a binding mechanism for further binding the released target miRNA-21 to other HP-AuNPs under the assistance of DSN; and
a result section configured to display a color change indicating the presence of miRNA-21 in the urine sample, wherein the color change is from red to bluish gray.

2. The system as claimed in claim 1, wherein the miRNA-21 is a potential biomarker for early cancer detection and is upregulated in many types of cancer.

3. The system as claimed in claim 1, wherein the DSN is a promising tool for the improvement of amplification of miRNA due to its unique selective cutting ability.

4. The system as claimed in claim 1, wherein the urine cancer detector strip is a non-invasive and cost-effective device for early cancer detection and allows for earlier diagnosis and treatment of cancer.
5. The system as claimed in claim 1, wherein the amplification segment is where the actual amplification of the miRNA-21 occurs if it is present in the urine sample.

6. A method for early cancer detection, comprising:
providing a urine cancer detector strip comprising a sample pad, a nitrocellulose membrane,
an amplification segment, and a plastic cassette;
receiving a urine sample on the sample pad;
moving the urine sample to the amplification segment for miRNA amplification, wherein the amplification segment is where the actual amplification of the miRNA-21 occurs if it is present in the urine sample;
detecting the presence of miRNA-21 in the urine sample using colorimetry and fluorescence dual mode detection assisted with duplex specific nuclease (DSN) and gold nanoparticle-based amplification, wherein detecting the presence of miRNA-21 comprises:
immobilizing FAM-labelled hairpin probes (FAM-HPs) on gold nanoparticles (AuNPs),
quenching the fluorescence of the FAM-HPs by the AuNPs,
capturing the target miRNA-21 by the HP-AuNPs probe,
converting the captured miRNA-21 into a DNA/RNA heteroduplex,
specifically recognizing and digesting the DNA/RNA heteroduplex by the DSN, and
further binding the released target miRNA-21 to other HP-AuNPs under the assistance of DSN; and
displaying a color change indicating the presence of miRNA-21 in the urine sample, wherein the color change is from red to bluish gray.

7. The method as claimed in claim 6, wherein the miRNA-21 is a potential biomarker for early cancer detection and is upregulated in many types of cancer.

8. The method as claimed in claim 6, wherein the DSN is a promising tool for the improvement of amplification of miRNA due to its unique selective cutting ability.

9. The method as claimed in claim 6, wherein the urine cancer detector strip is a non-invasive and cost-effective device for early cancer detection and allows for earlier diagnosis and treatment of cancer.

10. The method as claimed in claim 6, wherein the urine cancer detector strip further comprises a result section configured to display the color change indicating the presence of miRNA-21 in the urine sample.

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