KC-3099

CHOK1-TMEFF2-High Cell Line

34126

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Background of CHOK1-TMEFF2-High Cell Line

TMEFF2 (Transmembrane protein with EGF-like and two follistatin-like domains 2), also known as TENB2 or TPEF, is a type-I transmembrane proteoglycan primarily expressed in the prostate and central nervous system. It plays a complex dual role in cell signaling, acting as a neuroprotective survival factor in the brain while modulating the AKT and MAPK pathways in malignancies. In prostate cancer, TMEFF2 is significantly overexpressed and androgen-regulated, making it a critical diagnostic biomarker and therapeutic target. Current drug development focuses on Antibody-Drug Conjugates (ADCs) and bispecific T-cell engagers (TCEs); for instance, recent clinical trials for candidates like JNJ-70218902 (a T-cell redirecting antibody) have explored its potential in treating metastatic castration-resistant prostate cancer (mCRPC).

Specifications

Catalog Number	KC-3099
Cell Line Name	CHOK1-TMEFF2-High Cell Line
NCBI/UniProt Accession Number	NM_016192.4
Clone Number	3#
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line expressing exogenous human TMEFF2 gene in high level
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% RPMI1640 + 20% FBS + 10% DMSO
Propagation Medium	RPMI1640 + 10% FBS + 10μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-2 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

CHOK1-TMEFF2-High Cell Line was generated using a lentiviral vector expressing the human TMEFF2 sequence.

Characterization

Figure 1: Characterization of human TMEFF2 overexpression in the CHOK1-TMEFF2 stable high level clone using FACS.

Figure2: Characterization of endogenous human TMEFF2 expression in CHOK1 using FACS.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 + 10% FBS + 10μg/mL Puromycin)in a 37°C water bath.
2. Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
3. Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
4. Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
5. Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
6. Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
7. Incubate the flask at 37°C, 5% CO₂ incubator.
8. Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-2 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI1640 + 20% FBS + 10% DMSO) fresh immediately before use.
2. Keep the freezing medium on ice and label cryovials.
3. Transfer cells to a sterile, conical centrifuge tube, and count the cells.
4. Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
5. Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
6. Aliquot 1 mL of the cell suspension into each cryovial.
7. Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
8. Transfer vials to liquid nitrogen for long-term storage.

References

1. Masood, Motasim et al. “TMEFF2: A Transmembrane Proteoglycan with Multifaceted Actions in Cancer and Disease.” Cancers vol. 12,12 3862. 21 Dec. 2020, doi:10.3390/cancers12123862
2. Gao, Lingling et al. “TMEFF2 is a novel prognosis signature and target for endometrial carcinoma.” Life sciences vol. 243 (2020): 116910. doi:10.1016/j.lfs.2019.116910
3. Xie, Sidi et al. “TMEFF2 promoter hypermethylation is an unfavorable prognostic marker in gliomas.” Cancer cell international vol. 21,1 148. 4 Mar. 2021, doi:10.1186/s12935-021-01818-x
4. Calvo, Emiliano et al. “A first-in-human study of JNJ-70218902, a bispecific T-cell-redirecting antibody against TMEFF2 in metastatic castration-resistant prostate cancer.” The oncologist vol. 30,1 (2025): oyae313. doi:10.1093/oncolo/oyae313