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The CLN3 gene is a novel molecular target for cancer drug discovery
Svetlana N Rylova1, Andrea Amalfitano, Dixie-Ann Persaud-Sawin
1Department of Pediatrics, Duke University Medical Center, Durham, North Carolina 27710, USA.
Abstract:
Juvenile Batten disease is a neurodegenerative disease caused by accelerated apoptotic death of photoreceptors and neurons attributable to defects in the CLN3 gene. CLN3 is antiapoptotic when overexpressed in NT2 neuronal precursor cells. CLN3 negatively modulates endogenous ceramide levels in NT2 cells and acts upstream of ceramide generation. Because defects in regulation of apoptosis are involved in the development of cancer, we evaluated the expression of CLN3 on both mRNA and protein levels in a variety of cancer cell lines and solid colon cancer tissue. We also observed the effect of the blocking of CLN3 protein expression on cancer cell growth, survival, ceramide production, and apoptosis by using an adenovirus-bearing antisense CLN3 construct. We show that CLN3 mRNA and protein are overexpressed in glioblastoma (U-373G and T98g), neuroblastoma (IMR-32 and SK-N-MC), prostate (Du145, PC-3, and LNCaP), ovarian (SK-OV-3, SW626, and PA-1), breast (BT-20, BT-549, and BT-474), and colon (SW1116, SW480, and HCT 116) cancer cell lines but not in pancreatic (CAPAN and As-PC-1) or lung (A-549 and NCI-H520) cancer cell lines. CLN3 is also up-regulated in mouse melanoma and breast carcinoma cancer cell lines. We found CLN3 expression is 22-330% higher than in corresponding normal colon control tissue in 8 of 10 solid colon tumors. An adenovirus-expressing antisense CLN3 (Ad-AS-CLN3) blocks CLN3 protein expression in DU-145, BT-20, SW1116, and T98g cancer cell lines as seen by Western blot. Blocking of CLN3 expression using Ad-AS-CLN3 inhibits growth and viability of cancer cells. It also causes elevation in endogenous ceramide production through de novo ceramide synthesis and results in increased apoptosis as shown by propidium iodide and JC-1 staining. This suggests that Ad-AS-CLN3 may be an option for therapy in some cancers. More importantly these results suggest that CLN3 is a novel molecular target for cancer drug discovery.
Insights
Juvenile Batten disease gene CLN3 is overexpressed in many cancers and promotes cancer cell survival. Blocking CLN3 expression inhibits cancer growth and induces apoptosis, suggesting CLN3 as a potential cancer drug target.
Area of Science:
- Molecular Biology
- Cancer Research
- Neurodegenerative Diseases
Background:
- Juvenile Batten disease involves accelerated neuronal and photoreceptor apoptosis due to CLN3 gene defects.
- CLN3 exhibits antiapoptotic properties and negatively modulates ceramide levels.
- Dysregulation of apoptosis is implicated in cancer development.
Purpose of the Study:
- To investigate CLN3 gene and protein expression in various cancer cell lines and solid tumor tissues.
- To evaluate the impact of blocking CLN3 expression on cancer cell growth, survival, ceramide production, and apoptosis.
Main Methods:
- Analysis of CLN3 mRNA and protein levels in cancer cell lines and colon tumor tissues.
- Utilized an adenovirus-bearing antisense CLN3 construct (Ad-AS-CLN3) to block CLN3 protein expression.
- Assessed cancer cell growth, viability, ceramide production, and apoptosis using techniques like Western blot, propidium iodide, and JC-1 staining.
Main Results:
- CLN3 mRNA and protein were overexpressed in glioblastoma, neuroblastoma, prostate, ovarian, breast, and colon cancer cell lines, as well as in mouse melanoma and breast carcinoma cell lines.
- CLN3 expression was significantly higher in 80% of tested solid colon tumors compared to normal tissue.
- Blocking CLN3 expression with Ad-AS-CLN3 inhibited cancer cell growth and viability, increased endogenous ceramide production, and induced apoptosis.
Conclusions:
- CLN3 is overexpressed in a wide range of cancers and plays a role in cancer cell survival.
- Blocking CLN3 expression demonstrates therapeutic potential for certain cancers.
- CLN3 represents a novel molecular target for the development of new cancer drugs.