Field of the Invention
The present invention relates to a method of detecting cancer and, in particular, to a method of detecting cancer based on spermidine/spermine N1-acetyltransferase (SSAT) gene expression.
Description of the Related Art
U.S. Pat. No. 6,811,967 which issued to Sitar et al. on Nov. 4, 2004, and the full disclosure of which is incorporated herein by reference, discloses a method for assaying activity of the enzyme spermidine/spermine N1-acetyltransferase (SSAT) using SSAT substrates by detecting acetylated forms of the SSAT substrates. The SSAT substrates may include amantadine wherein metabolism of amantadine occurs in part by the action of the inducible enzyme SSAT to produce the acetylated metabolite N-acetylamantadine. Disclosed also is the correlation of SSAT activity to pathological conditions.
It is an object of the present invention to provide a method of detecting cancer based on spermidine/spermine N1-acetyltransferase (SSAT) gene expression.
Elevated levels of SSAT gene expression expression were observed in breast, prostate and lung cancer cell lines. Elevated levels of SSAT gene expression were verified in tissues from patients with breast, prostate and lung cancer. Elevated SSAT gene expression is elevated in different human cancers and elevated SSAT gene expression may accordingly serve as a companion diagnostic biomarker for detection and monitoring of cancer progression.
There is accordingly provided a method of detecting cancer comprising correlating spermidine/spermine N1-acetyltransferase gene expression to cancer. The method may comprise correlating elevated levels of spermidine/spermine N1-acetyltransferase gene expression to cancer. The method may comprise correlating spermidine/spermine N1-acetyltransferase gene expression to breast cancer. The method may comprise correlating spermidine/spermine N1-acetyltransferase gene expression to lung cancer. The method may comprise correlating spermidine/spermine N1-acetyltransferase gene expression to prostate cancer.
The invention will be more readily understood from the following description of the embodiments thereof given, by way of example only, with reference to the accompanying drawings, in which:
Spermidine/spermine N1-acetyltransferase (SSAT) is the rate-limiting enzyme in the polyamine metabolic pathway. SSAT plays a regulatory role in spermidine and spermine homeostasis and normally is present in very small amounts in mammalian cells. However, the increased production of polyamines in cancer results in increased levels of polyamines and N1-acetylspermidine, potentially reflecting increased SSAT activity. The elevation of polyamines triggers the increase in SSAT activity to remove polyamines as part of a cell defense system as shown in
Total RNA was extracted from the human cancer cell lines and the patient-derived breast cancer tissue, prostate cancer tissue and lung cancer tissue using a Qiagen QIA™ Shredder Kit and RNeasy™ Mini Kit obtained from Life Technologies Inc. of Ontario, Canada. RNA concentration in each extracted sample was confirmed by nanodrop spectrophotometric measurement. RNA integrity was evaluated by measurement of RNA Integrity Number (RIN). SSAT gene expression was determined by qRT-PCR using cDNA probe specific for SSAT and performed using Qiagen QuaniTect™ SYBR Green RT-PCR Kit obtained from Life Technologies Inc. of Ontario, Canada. The mRNA expression levels of the housekeeping genes, glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and hypoxanthine phosphoribosyltransferase 1 (HRPT1), were measured in parallel using the corresponding PCR primers for these genes. The SSAT gene expression levels were normalized with GAPDH or HPRT1 as the internal reference. Normalized SSAT gene expression was further analyzed by the ΔΔct method.
The statistical analysis of the data was performed using SigmaStat (SPSS In.) software. Values were expressed as mean±standard error of the mean. Differences between two groups were evaluated by Student's t-test. A probability of 95% or more (P<0.05) was considered significant.
Referring to
Western blots were also obtained for the patient-derived breast cancer tissue, lung cancer tissue, and prostate cancer tissue and are shown in
The quantified data of the Western blots expressed as a ratio of the internal control, tubulin, as the housekeeping protein is shown below.
The increased levels of SSAT mRNA expression in patient-derived tumors provide further evidence that elevated SSAT gene expression levels in cancer tissue may serve as a diagnostic marker for the detection of cancer and that SSAT may be a target for anti-cancer drug development.
It will be understood by a person skilled in the art that many of the details provided above are by way of example only, and are not intended to limit the scope of the invention which is to be determined with reference to the following claims.
Filing Document | Filing Date | Country | Kind |
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PCT/CA15/50534 | 6/9/2015 | WO | 00 |
Number | Date | Country | |
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62009864 | Jun 2014 | US |