RT-112 Orthotopic Mouse Model Service for Bladder Cancer

The RT-112 orthotopic mouse model represents a differentiated, FGFR3-amplified luminal bladder cancer subtype, offering a clinically relevant preclinical system for testing FGFR inhibitors, intravesical agents, and differentiation-targeted therapies within a native urothelial microenvironment. Alfa Cytology builds and validates this model with meticulous attention to engraftment consistency, histopathological fidelity, and endpoint rigor—delivering pharmacodynamic datasets that support mechanistic insights and regulatory decision-making for your bladder cancer pipeline.

Overview of RT-112 Orthotopic Mouse Model for Bladder Cancer

The RT-112 cell line was established in 1973 from a grade II transitional cell carcinoma of the urinary bladder excised from a female patient. Genetically, RT-112 is defined by FGFR3 gene amplification and high-level protein overexpression, driven in part by an FGFR3–TACC3 fusion transcript that constitutively activates downstream RAS/MAPK and PI3K/AKT signaling. Unlike aggressive muscle-invasive lines, RT-112 retains wild-type TP53 and functional Rb, exhibits a near-diploid karyotype with low chromosomal instability, and maintains robust epithelial differentiation marked by E-cadherin, cytokeratin 20, and uroplakin II expression. Its luminal identity is further supported by high expression of the lineage transcription factors GATA3 and FOXA1.

Fig 1. Schematic illustration comparing the development of normal human bladder cancer to the development of bladder cancer in mice after this protocol is performed. (Lorenzatti Hiles, Guadalupe, et al., 2019)

When established orthotopically in immunodeficient hosts via intravesical instillation or intramural injection, RT-112 cells form well-differentiated, non-invasive to superficially invasive papillary tumors that recapitulate the architecture of low- to intermediate-grade human urothelial neoplasia. The model is exquisitely sensitive to FGFR-directed small-molecule inhibitors—including erdafitinib, infigratinib, and PD173074—making it the model of choice for preclinical validation of FGFR3-targeted therapeutics. Additionally, RT-112’s intact DNA damage response and functional p53 pathway render it suitable for evaluating agents that exploit these vulnerabilities, such as platinum-based chemotherapeutics and differentiation-modulating compounds. Tumor progression can be monitored through ultrasound imaging, and luciferase-engineered derivatives enable bioluminescence-based longitudinal quantification when higher throughput is desired.

Cell Line Information: RT-112

RT-112 (also designated RT112/84) is a well-characterized human bladder transitional cell carcinoma line that serves as a cornerstone model for FGFR3-driven, luminal-subtype bladder cancer research. Its relatively stable genome and preserved differentiation markers distinguish it from high-grade, mesenchymal bladder cancer models.

Our Services

Alfa Cytology offers end-to-end construction and management of the RT-112 orthotopic bladder cancer model, leveraging its unique FGFR3-driven biology to support targeted therapy development programs. Our scientists optimize intravesical instillation protocols—including poly-L-lysine preconditioning, controlled dwell times, and urethral clamping—to achieve consistent engraftment while preserving the differentiated, luminal tumor phenotype that defines RT-112’s clinical relevance. Each study is executed under IACUC-approved protocols with comprehensive histopathological validation, biomarker profiling, and pharmacodynamic readouts tailored to your compound’s mechanism of action.

Workflow of RT-112 Orthotopic Mouse Model Construction

Construction of the RT-112 orthotopic bladder tumor model follows a standardized pipeline optimized for this slow-growing, differentiation-preserving cell line. Both transurethral intravesical instillation and open surgical intramural injection routes are available, with the former preferred for evaluating intravesical agents and the latter for achieving precise intramural tumor localization. Given RT-112’s relatively low proliferative rate, extended culture periods and gentle handling are employed to maintain viability throughout the procedure.

1. Cell Expansion and Pre-Implantation Quality Control: RT-112 cells are expanded in RPMI 1640 complete medium and harvested during the exponential growth phase, typically 2–5 days post-passage. Because RT-112 is difficult to trypsinize, an extended 5–10-minute trypsin/EDTA treatment is applied to achieve complete detachment without compromising viability. A single-cell suspension is prepared, viability is confirmed by trypan blue exclusion (>85% required), and mycoplasma PCR testing is performed on each batch.
2. Host Selection and Surgical Preparation: Female athymic nude mice, 6–8 weeks of age, are selected as hosts due to RT-112’s human origin and immunogenicity. Mice are acclimatized for a minimum of one week, then anesthetized with isoflurane (3% induction, 1.8% maintenance) on a heated surgical platform. The lower abdomen is clipped and prepared aseptically, and ophthalmic ointment is applied to prevent corneal desiccation during the procedure.
3. Bladder Preconditioning (Intravesical Route): The bladder is manually emptied by gentle abdominal compression. A lubricated 24-gauge catheter is advanced transurethrally to approximately 1 cm depth. Fifty microliters of 0.1% poly-L-lysine is instilled and allowed to dwell for 15 minutes to disrupt the urothelial glycosaminoglycan barrier and enhance RT-112 cell adhesion—a step particularly critical for this slow-adhering, differentiated line.
4. Tumor Cell Instillation or Intramural Injection: For intravesical delivery, RT-112 cells are resuspended in complete growth medium (not PBS) to preserve viability, and 2×10⁶ cells in 50 µL are slowly instilled through the catheter. A lightweight surgical clamp is applied around the urethral meatus, and the suspension is retained for 1.5 hours under continuous anesthesia to maximize attachment. For the open surgical route, a 1-cm lower midline incision is made, the bladder is exteriorized, and a 1:1 mixture of Matrigel and RT-112 cell suspension (~2×10⁶ cells in 10 µL) is injected intramurally using a 30-gauge needle guided by curved forceps.
5. Post-Procedure Care and Recovery: Following intravesical instillation, the clamp and catheter are removed, and the bladder is allowed to empty spontaneously. After open surgery, the bladder is returned to the abdominal cavity, the wall is closed with absorbable sutures, and the skin is secured with wound clips. Mice receive appropriate analgesia per IACUC guidelines and are monitored in a heated recovery chamber until fully ambulatory. Body weights are recorded daily for the first three days and twice weekly thereafter.
6. Tumor Monitoring and Terminal Analysis: Tumor growth is monitored every 3–7 days via high-frequency micro-ultrasound to visualize bladder wall thickness and tumor morphology. For luciferase-tagged RT-112 variants, bioluminescence imaging is performed following intraperitoneal luciferin injection to quantify tumor burden non-invasively. At study endpoint, bladders are harvested, weighed, and processed for H&E staining, IHC analysis of FGFR3, Ki-67, E-cadherin, and cleaved caspase-3, and correlation of imaging data with histopathological findings.

Fig 2. RT-112 Orthotopic Mouse Model construction workflow.

Case Study-RT-112 Orthotopic Mouse Model Development

In a preclinical evaluation program, Alfa Cytology utilized the RT-112 orthotopic model to assess the efficacy of an FGFR3-targeted therapeutic candidate in a differentiated bladder tumor context. Following poly-L-lysine preconditioning and intravesical instillation, tumors were allowed to establish for two weeks, after which high-frequency ultrasound confirmed uniform tumor presence across the cohort. Animals were randomized into vehicle control and treatment groups, with dosing initiated once tumors reached a measurable size. Throughout the treatment window, tumor dimensions were tracked via serial ultrasound imaging, and body weights were monitored to assess treatment tolerability. At study termination, excised bladders underwent comprehensive histopathological evaluation, including H&E staining and IHC profiling of FGFR3, proliferation markers, and apoptotic indices. The integrated dataset revealed treatment-associated reductions in tumor cellularity and FGFR3 pathway activity, providing the sponsor with mechanistic evidence to support further preclinical development of their FGFR-directed agent.

Why Choose Alfa Cytology?

Engaging Alfa Cytology for your RT-112 orthotopic bladder cancer study ensures access to a biologically faithful, FGFR3-driven model platform managed by scientists with specialized expertise in luminal bladder cancer biology and intravesical delivery optimization.

• Differentiation-Preserving Protocols: Our gentle handling and optimized preconditioning methods maintain RT-112’s epithelial, luminal phenotype throughout the study, ensuring histopathological fidelity to human low-grade urothelial tumors.
• FGFR3 Expertise: We have deep experience with FGFR-driven models, enabling precise pharmacodynamic readouts and biomarker correlations for FGFR inhibitor programs.
• Flexible Implantation Routes: Both transurethral intravesical and open intramural techniques are available, selected to match your therapeutic route and study objectives.
• Multimodal Longitudinal Monitoring: High-frequency ultrasound and optional bioluminescence imaging provide real-time tumor growth data with minimal animal stress.
• Comprehensive Endpoint Portfolio: Terminal assessments include bladder weight, H&E histopathology, FGFR3 IHC, proliferation and apoptosis markers, and DNA damage response profiling.
• Regulatory-Aligned Reporting: All data are generated under IACUC-approved protocols with full traceability, supporting IND-enabling and publication-ready documentation.

Contact Us

Looking to evaluate your FGFR-targeted or intravesical therapeutic candidate in a differentiated, clinically relevant bladder cancer model? Reach out to us today to discuss how the RT-112 orthotopic model can advance your preclinical program. Our scientific team will collaborate with you to design a customized study plan, define optimal endpoints, and deliver a comprehensive data package aligned with your development milestones.

For research use only. Not intended for any clinical use.