Bladder Cancer Biomarker May Lend New Diagnostic and Therapeutic Strategies

bladder cancer research

Diane M. Simeone, MD

A paper published October 2015 inCancer Researchspotlights a potential biomarker and therapeutic target for bladder cancer (BC). The authors write that although BC is recognized as a common and deadly malignancy, a lack of understanding of the molecular pathways involved has hindered development of effective treatments compared with progress made with other malignancies.¹

Using a transgenic mouse model, human BC tissue and human BC cell lines, the team led by Diane M. Simeone, MD, Lazar J Greenfield Professor of Surgery and chief of molecular and integrative physiology, Division of Gastrointestinal Surgery at the University of Michigan, Ann Arbor, examined the role of the Ataxia-telangiectasia group D complementing gene (ATDC), also known asTRIM29in BC.¹Simeone et al showed thatATDCdrives invasive BC development and identified the microRNA-mediated and epigenetic pathways involved.

The authors cite previous studies indicating a precedent for suspecting a role forATDCin BC and that it is highly expressed in several types of cancer including pancreatic, lung, head and neck, and cervical cancer.¹

The Transgenic Mouse Model

The group generated a transgenic mouse model by injecting theCAG-ATDCtransgene into the pronuclei of fertilized eggs of FVB/NJ mice. The resulting mice overexpressedATDC. The mice were divided into three groups on the basis of how many copies of theCAG-ATDCtransgene they had, namely 2, 4 or 6. Analysis revealed that the overexpression ofATDCwas transgene dose dependent, and this was reflected in bladder tumor formation (Table).¹

Table: Tumor Histology and Number of Transgene Copies

Number of Copies of Transgene

Hyperplasia

N (%)

Noninvasive

Tumors N (%)

Invasive Tumors N (%)

2

2/19 (10.5)

4

2/22 (9)

1/22 (5)

6

3/21 (14)

6/21 (29) including CIS

CIS, carcinoma in situ

On examination of the histology, the tumors were found to be indistinguishable from human urothelial cancers.¹

To further establish the validity of the model, the gene expression signature of the mice withATDC-induced tumors was built by identifying all genes over expressed >2-fold versus littermate controls. The authors state that, “The genes over-expressed inCAG-ATDCbladder tumors significantly overlapped with multiple expression profiling signatures of genes over-expressed in human BCs and human infiltrating vs superficial human urothelial BCs.”¹

Proliferation characteristics were decreased in human BC cell lines when ATDC knockdown was achieved by transfection of two ATDC-shRNAs, while the over expression ofATDCin a BC cell line with low ATDC levels resulted in a 37% increase in invasiveness when measured with a modified Boyden-chamber invasion assay. Again using a cell line, the growth of orthotopic bladder xenograft tumors was significantly reduced when ATDC was silenced using ATDC shRNA vectors versus control with no silencing.¹

Downstream of ATDC

Knowing from previous studies, that loss of phosphatase and tensin homolog (PTEN) expression contributes to the appearance of noninvasive and invasive BCs in transgenic mice, the authors conducted experiments to test involvement ofATDCin loss ofPTEN.^1-4They found using theirCAG-ATDCdriven BCs mouse model, thatPTENexpression was lost in noninvasive and invasive BC tumors but present in control mice. Using the human cell lines they were able to demonstrate thatATDCachieves silencing ofPTENby upregulating DNMT3A, a DNA methyltransferase. This epigenetic mechanism leads to the methylation of the PTEN promoter preventing expression ofPTEN, and the team further showed that this step drives tumor invasion and proliferation, in effect, driving a malignant phenotype.¹

But the mechanism does not end there. More details emerged when the experiments evaluated the activity of miR-29 (micro RNA). The miR-29 family regulate mRNA (messenger RNA) levels of DNMT3A, and experiments revealed that blocking miR-29 family members in human BC cell lines led to an increase in DNMT3A. Establishing a link toATDC, they showed that a knockdown of ATDC in human BC cell lines led to an increase in miR-29 A and B and a subsequent reduction of DNMT3A mRNA. Furthermore, they showed thatATDCexpression upregulated MYC (a regulator of gene transcription^5,6) leading to a reduced transcription of miR-29 and so an upregulation of DNMT3A.¹

Human Tumors

Analyzing ATDC mRNA expression in two large multicancer studies in the Oncomine database,^7,8the team showed that human BCs exhibited the highest expression ofATDCamong other cancer types and that compared with normal bladder tissue, both noninvasive and invasive BCs had significantly higher levels ofATDC. An analysis of tissue microarrays of 283 human primary BC showed thatATDCwas present in 53% of muscle-invasive BCs and 50% of CIS BCs. Elevated ATDC staining was detected in 22% of noninvasive BCs. Importantly, elevatedATDCexpression correlated with invasive disease.¹

Future Directions

Referring to the 20% of human noninvasive BCs that highly expressed ATDC the authors stated, “This raises the intriguing hypothesis thatATDCexpression in noninvasive BC may predict progression from noninvasive tumors to muscle invasive disease,” and concluded with, “Given its relevance to human BC, further studies of ATDC-induced tumorigenesis are likely to provide insight into the biology of invasive BC, and potentially provide novel diagnostic and therapeutic strategies for patients.”¹

References

1. Palmbos PL, Wang L, Yang H, et al. ATDC (TRIM29) Drives invasive BC formation.Cancer Res. 2015 Oct 15. pii: canres.0603.2015.

2. Ahmad I, Morton JP, Singh LB, et al. ß-Catenin activation synergizes with PTEN loss to cause BC formation.Oncogene. 2011;30(2):178-189.

3. Puzio-Kuter AM, Castillo-Martin M, Kinkade CW, et al. Inactivation of p53 and Pten promotes invasive BC.Genes Dev. 2009;23(6):675-680.

4. Tsuruta H, Kishimoto H, Sasaki T, et al. Hyperplasia and carcinomas in Pten-deficient mice and reduced PTEN protein in human BC patients.Cancer Res.2006;66(17):8389-8396.

5. Thomas LR, Foshage AM, Weissmiller AM, Tansey WP. The MYC-WDR5 Nexus and Cancer.Cancer Res. 2015;75(19):4012-4015.

6. Stine ZE, Walton ZE, Altman BJ, Hsieh AL, Dang CV. MYC, Metabolism, and Cancer.Cancer Discov. 2015;5(10):1024-1039.

7. Oncomine™. Prescribing information. Compendia Bioscience: Ann Arbor, MI.

8. Su AI, Welsh JB, Sapinoso LM, et al. Molecular classification of human carcinomas by use of gene expression signatures.Cancer Res. 2001;61(20):7388-7393