[1306] WNT-β-Catenin Is Upregulated in Very Aggressive Tumor Pattern of Invasion

Tatyana Isayeva, Margaret Brandwein-Gensler. UAB, Birmingham, AL

Background: The WNT pathway is a critical to proliferation, differentiation, oncogenesis and is upregulated in many cancers. Among the downstream effects, WNT-β-catenin is involved in tumor invasion. We developed 4 cell lines from oral/oropharyngeal SCC, which are unique as they are matched to different worst patterns of invasion (WPOI) in the corresponding resections. These histopathological WPOI were developed and validated by us as outcome prognosticators.
Design: Cell lines were derived from fresh resections, cancer cells were sorted from cancer-associated fibroblasts by flow cytometry and grown in keratinocyte media/keratinocyte growth supplements. HPV was determined by RT-PCR with HPV16/18 E6/E7 primers, with appropriate controls. Invasion was measured with the BioCoat Matrigel chamber, 24h after stimulation with TGFß. Cells that invaded across the chamber membrane were counted after fixation and staining. UAB-1, UAB-4, and the corresponding frozen samples from the 2 resections were assessed for WNT signalling with the RT2 96 well RT-PCR kit (SABiosciences).
Results: Table 1 shows the clinicopathologic features for these cells.

Table 1: UAB Cell Lines

	UAB-1	UAB-2	UAB-3	UAB-4
Pattern of invasion, corresponding resection	WPOI3	WPOI3	WPOI5	WPOI5
HPV16/18	Pos/Neg	Neg/Neg	Neg/Neg	Neg/Neg
Site	Orophx	Oral	Oral	Oral
Invasion Assay	3.0	14.0	32.7	40.7

Mean invading tumor cell counts after TGFβ stimulation, normalized for control (no stimulation)

UAB-1 was HPV16+/HPV18-, all other cells were HPV. Transwell assays showed increased invasion in WPOI5 cells compared with WPOI3 cells after stimulation.

RT-PCR results shows up-regulation of WNT-pathway in tumor-2/UAB-4, compared to tumor-1/UAB-1.

Table 2: WNT in UAB-1 & UAB-4

Gene	Tumor-1 (WPOI3)	UAB-1 derived from Tumor-1	Tumor-2 (WPOI5)	UAB-4 derived from Tumor-2
WISP1	0	4.2	1.6	6.7
WNT1	0	1.3	5.1	15.2
WNT6	1.4	5.9	4.9	39.1
WNT7A	0	3.6	4.9	36.9
WNT10A	0	0	2.8	14.7
WNT11	0	2.5	5.9	35.8
CTNNB1	0	6.5	3.5	9.8

RT-PCR, C(t) normalized for mean of 5 internal controls


Conclusions: Cell lines derived from WPOI5 carcinomas (aggressive pattern of invasion) have greater invasive characteristics, compared to cell lines from WPOI3 (nonaggressive) tumors. Aggressive tumor invasion is associated with upregulated WNT signalling. These cell lines will serve as a mechanistic model of invasion in ongoing studies.
Category: Head & Neck

Wednesday, March 21, 2012 9:30 AM

Poster Session V # 166, Wednesday Morning