Wilms3

The Wilms3 cell line was established from a primary Wilms tumor in a pediatric patient, characterized by a somatic WT1 mutation. Unlike many other Wilms tumor cell lines, Wilms3 harbors a heterozygous frameshift mutation in the WT1 gene (c.1293-1294insA, p.V432SfsX87), leading to the production of a truncated WT1 protein. This partial loss of WT1 function is associated with the development of tumors that display a stromal or mesenchymal phenotype. However, the WT1 mutation in Wilms3 is not homozygous, which adds complexity to its study, as it retains some WT1 function that can influence tumor biology differently compared to cell lines with complete WT1 loss.

Wilms3 also carries a mutation in the CTNNB1 gene, specifically affecting threonine 41 (p.T41A), which plays a critical role in the Wnt signaling pathway. This mutation stabilizes β-Catenin, preventing its degradation and leading to the constitutive activation of the Wnt pathway. The persistent activation of Wnt signaling drives cell proliferation and contributes to tumorigenesis in Wilms3, making it a key model for studying the impact of CTNNB1 mutations in the context of a partially functional WT1 background.

Phenotypically, Wilms3 cells exhibit a mesenchymal-like morphology, expressing vimentin and lacking cytokeratin, consistent with the stromal characteristics observed in the original tumor. These cells show limited differentiation potential, with the ability to undergo some mesenchymal differentiation under specific conditions. Proteomic analyses of Wilms3 have revealed the activation of several receptor tyrosine kinases (RTKs), including PDGFRβ and AXL, which support cell survival and proliferation. Additionally, downstream signaling pathways such as MAPK and PI3K/AKT are activated, reinforcing the malignant properties of Wilms3 cells.

One unique aspect of Wilms3 is its partial WT1 functionality, which provides a distinct perspective on how WT1 mutations contribute to Wilms tumor biology when the mutation is not complete. The interplay between WT1 and Wnt signaling in Wilms3 offers a valuable opportunity to study the nuanced roles these pathways play in tumor development. Overall, Wilms3 serves as an important model for investigating the molecular mechanisms underlying Wilms tumor in the presence of partial WT1 loss and constitutive Wnt pathway activation.

37°C, 5% CO₂, humidified atmosphere.

1. Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit.

2. Upon receipt, either store the cryovial immediately at temperatures below -150°C to ensure the preservation of cellular integrity, or proceed to step 3 if immediate culturing is required.

3. For immediate culturing, swiftly thaw the vial by immersing it in a 37°C water bath with clean water and an antimicrobial agent, agitating gently for 40-60 seconds until a small ice clump remains.

4. Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening.

5. Carefully open the disinfected vial and transfer the cell suspension into a 15 ml centrifuge tube containing 8 ml of room-temperature culture medium, mixing gently.

6. Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium.

7. Gently resuspend the cell pellet in 10 ml of fresh culture medium. For adherent cells, divide the suspension between two T25 culture flasks; for suspension cultures, transfer all the medium into one T25 flask to promote effective cell interaction and growth.

8. Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes.

Mycoplasma contamination is excluded using both PCR-based assays and luminescence-based mycoplasma detection methods.

To ensure there is no bacterial, fungal, or yeast contamination, cell cultures are subjected to daily visual inspections.

For long-term preservation, place vials in vapor-phase liquid nitrogen at about −150 to −196 °C. Storage at −80 °C is acceptable only as a short interim step before transfer to liquid nitrogen.

Cryopreserved cell lines are shipped on dry ice in validated, insulated packaging with sufficient refrigerant to maintain approximately −78 °C throughout transit. On receipt, inspect the container immediately and transfer vials without delay to appropriate storage.

At Cytion, we stand behind the quality and viability of our products. We guarantee that our cells will be viable at the start of culture for 60 days after shipment from our facilities. We assure that all materials conform to the specifications described in the Certificate of Analysis and/or Product Sheet.

For full details, please refer to our Warranty Statement.