KC-5644

HEK293-ERBB3-EGFR Cell Line

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Home » HEK293-ERBB3-EGFR Cell Line

Background of HEK293-ERBB3-EGFR Cell Line

ERBB3 (Erb-B2 Receptor Tyrosine Kinase 3) is a Protein Coding gene. Human epidermal growth factor receptor-3 (ERBB3) is a member of the ERBB receptor tyrosine kinases (RTKs) and is expressed in many malignancies. It is activated by binding to ligands Neuregulin-1 and Neuregulin-2. Since ERBB3 lacks intrinsic kinase activity, signal transduction occurs through formation of heterodimers with EGFR, ERBB2, and ERBB4. ERBB3 is a signaling specialist since it has six binding sites for the p85 SH2 adapter subunit of phosphoinositide 3' kinases. Along with other ERBB receptors, ERBB3 is associated with regulating normal cell proliferation, apoptosis, differentiation, and survival, and has received increased research attention for its involvement in cancer therapies. ERBB3 expression or co-expression levels have been investigated as predictive factors for cancer prognosis and drug sensitivity. Diseases associated with ERBB3 include Lethal Congenital Contracture Syndrome 2 and Visceral Neuropathy, Familial, 1, Autosomal Recessive.
Epidermal Growth Factor Receptor (EGFR) is a transmembrane glycoprotein that constitutes one of the four members of ErbB family of tyrosine kinase receptors. Activation of EGFR leads to autophosphorylation of receptor tyrosine kinase that initiates a cascade of downstream signaling pathways involved in regulating cellular proliferation, differentiation, and survival. EGFR is abnormally activated by various mechanisms like receptor overexpression, mutation, ligand-dependent receptor dimerization, ligand-independent activation and is associated with the development of variety of human cancers.

Specifications

Catalog NumberKC-5644
Cell Line NameHEK293-ERBB3-EGFR Cell Line
Clone Number45#
Host Cell LineHEK293
DescriptionStable HEK293 clone expressing exogenous human ERBB3 and EGFR gene
QuantityTwo vials of frozen cells (≥2-106/vial)
StabilityStable in culture over a minimum of 10 passages
ApplicationDrug screening and biological assays
Freezing Medium70% DMEM + 20% FBS + 10% DMSO
Propagation MediumDMEM + 10% FBS + 150μg/mL Hygromycin B
Selection MarkerHygromycin B
MorphologyEpithelial
SubcultureSplit saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 105 cells/mL
Incubation37 °C with 5% CO2
StorageLiquid nitrogen immediately upon receiving
Doubling TimeApproximately 30 hours
Mycoplasma StatusNegative

Cell Line Generation

HEK293-ERBB3-EGFR cell line was generated using a lentiviral vector expressing the ERBB3 and EGFR sequence.

Characterization

Figure 1: Characterization of ERBB3 and EGFR overexpression in the HEK293-ERBB3-EGFR stable clone using FACS(Kyinno, Cat#KB-1106).

Figure 2: Characterization of ERBB3 and EGFR in the HEK293 stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (DMEM supplemented with 10% FBS and 150μg/mL Hygromycin B)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 1-3 × 105 cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% DMEM + 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. Chen Y, Lu A, Hu Z, Li J, Lu J. ERBB3 targeting: A promising approach to overcoming cancer therapeutic resistance. Cancer Lett. 2024 Sep 1;599:217146. doi: 10.1016/j.canlet.2024.217146. Epub 2024 Aug 2. PMID: 39098760.
2. Nguyen DQ, Hoang DH, Nguyen Vo TT, Huynh V, Ghoda L, Marcucci G, Nguyen LXT. The role of ErbB3 binding protein 1 in cancer: Friend or foe? J Cell Physiol. 2018 Dec;233(12):9110-9120. doi: 10.1002/jcp.26951. Epub 2018 Aug 4. PMID: 30076717.
3. Sabbah DA, Hajjo R, Sweidan K. Review on Epidermal Growth Factor Receptor (EGFR) Structure, Signaling Pathways, Interactions, and Recent Updates of EGFR Inhibitors. Curr Top Med Chem. 2020;20(10):815-834. doi: 10.2174/1568026620666200303123102. PMID: 32124699.
4. Sigismund S, Avanzato D, Lanzetti L. Emerging functions of the EGFR in cancer. Mol Oncol. 2018 Jan;12(1):3-20. doi: 10.1002/1878-0261.12155. Epub 2017 Nov 27. PMID: 29124875; PMCID: PMC5748484.
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