• Origin: The MC38 cell line is derived from C57BL6 murine colon adenocarcinoma cells. The cell line is a commonly used murine model for colorectal carcinoma.
• Background Information: The human epidermal growth factor receptor 2 (HER2), known as erbB-2, or proto-oncogene Neu, is a receptor tyrosine-protein kinase encoded by the ERBB2 (HER2) gene on chromosome 17q12. ErbB2 does not bind ligand, yet appears to be the major signaling partner for other ERBB receptors by forming heteromeric complexes with ErbB1, ErbB3, or ErbB4. Upon ligand-induced dimerization, the receptors autophosphorylate on specific tyrosine residues in their cytoplasmic tails. These residues provide docking sites for phosphotyrosine binding, cytoplasmic signaling molecules that activate numerous intracellular signaling pathways. HER2 overexpresses in a broad number of cancer types, such as bladder, breast, cervical, cholangio, colorectal, endometrial, esophageal, gastric, head and neck, liver, lung, ovarian, and salivary gland cancers. Especially, amplification and overexpression of HER2 occurs in 25% to 30% of the instances of human breast cancer and are associated with a poor prognosis. Several HER2-targeting therapies such as trastuzumab, pertuzumab, T-DM1, DS8201 and RC48 have been approved worldwide for patients with HER2-positive tumors.
• Gene targeting strategy: By using the unique gene editing technology of Biocytogen, the exogenous promoter, human HER2 coding sequence, and terminator sequence were modified into B-hHER2 MC38 plus cell lines.
• Tumorigenicity: Confirmed in B-hHER2 mice, B-h4-1BB/hHER2 mice.
• Application: The B-hHER2 MC38 plus tumor models can be used for preclinical evaluation of antibody drugs targeting human HER2.
• Notes: B-hHER2 MC38 plus cells can not establish tumors in wild-type C57BL/6 mice, they can only establish tumors in HER2 humanized mice.

HER2 expression analysis in B-hHER2 MC38 plus cells by flow cytometry. Single cell suspension from B-hHER2 MC38 plus #1-F12 cultures was analyzed with anti-human HER2 antibody (Biolegend, 324406) by flow cytometry. Human HER2 was detected on the surface of B-hHER2 MC38 plus and human cell line NCI-N87.

HER2 expression evaluated on B-hHER2 MC38 plus tumor cells by flow cytometry.

B-hHER2 MC38 plus cells were subcutaneously transplanted into B-hHER2 mice (n=5). Upon conclusion of the experiment, tumor cells were harvested and analyzed with anti-human HER2 antibody (Biolegend, 324406) by flow cytometry. As shown, human HER2 was highly expressed on the surface of tumor cells. Therefore, B-hHER2 MC38 plus cells can be used for in vivo efficacy studies evaluating novel HER2 therapeutics.

Subcutaneous tumor growth of B-hHER2 MC38 plus cells. B-hHER2 MC38 plus cells (5x10⁵) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into B-hHER2 mice (female, 8-10-week-old, n=5). Tumor volume and body weight were measured twice a week. (A) Average tumor volume. (B) Body weight. Volume was expressed in mm³ using the formula: V=0.5 X long diameter X short diameter². Results indicate that B-hHER2 MC38 plus cells were able to establish tumors in vivo and can be used for efficacy studies. Values are expressed as mean ± SEM.

B-hHER2 MC38 plus tumor growth curves from individual mice. B-hHER2 MC38 plus cells (5x10⁵) and wild-type MC38 cells (5x10⁵) were subcutaneously implanted into B-hHER2 mice (female, 8-10-week-old, n=5). Results indicate that B-hHER2 MC38 plus cells were able to establish tumors in vivo and can be used for efficacy studies. Values are expressed as mean ± SEM.

Antitumor activity of anti-human HER2 ADC (Trastuzumab analog-MMAE, in-house) in B-h4-1BB/hHER2 mice. (A) Anti-human HER2 ADC inhibited B-hHER2 MC38 plus tumor growth in B-h4-1BB/hHER2 mice. Murine colon cancer B-hHER2 MC38 plus cells were subcutaneously implanted into homozygous B-h4-1BB/hHER2 mice (female, 8-week-old, n=6). Mice were grouped when tumor volume reached approximately 100-150 mm³, at which time they were intraperitoneally injected with anti-human HER2 ADC Trastuzumab analog-MMAE (in-house) indicated in panel. (B) Body weight changes during treatment. As shown in panel A, 10mg/kg anti-human HER2 ADC was efficacious in controlling tumor growth in B-h4-1BB/hHER2 mice, demonstrating that B-hHER2 MC38 plus provide a powerful preclinical model for in vivo evaluation of anti-human HER2 ADC. Values are expressed as mean ± SEM.

Antitumor activity of anti-human HER2 ADC (Trastuzumab analog-MMAE, in-house) in B-h4-1BB/hHER2 mice. Murine colon cancer B-hHER2 MC38 plus cells were subcutaneously implanted into homozygous B-h4-1BB/hHER2 mice (female, 8-week-old, n=6). Mice were grouped when tumor volume reached approximately 100-150 mm³, at which time they were intraperitoneally injected with anti-human HER2 ADC Trastuzumab analog-MMAE (in-house) indicated in panel.

Antitumor activity of anti-human HER2 ADC DS8201 in B-h4-1BB/hHER2 mice. (A) Anti-human HER2 ADC inhibited B-hHER2 MC38 plus tumor growth in B-h4-1BB/hHER2 mice. Murine colon cancer B-hHER2 MC38 plus cells were subcutaneously implanted into homozygous B-h4-1BB/hHER2 mice (female, 9-week-old, n=6). Mice were grouped when tumor volume reached approximately 100 mm³, at which time they were intraperitoneally injected with anti-human HER2 ADC DS8201 (purchased from Daiichi Sankyo) or Trastuzumab analog-MMAE (in-house) indicated in panel. (B) Body weight changes during treatment. As shown in panel A, Both anti-human HER2 ADC were efficacious in controlling tumor growth in B-h4-1BB/hHER2 mice, demonstrating that B-hHER2 MC38 plus provide a powerful preclinical model for in vivo evaluation of anti-human HER2 ADC. Values are expressed as mean ± SEM.

Antitumor activity of anti-human HER2 ADC (Trastuzumab analog-MMAE, in-house) in B-h4-1BB/hHER2 mice. Murine colon cancer B-hHER2 MC38 plus cells were subcutaneously implanted into homozygous B-h4-1BB/hHER2 mice (female, 9-week-old, n=6). Mice were grouped when tumor volume reached approximately 100 mm³, at which time they were intraperitoneally injected with anti-human HER2 ADC DS8201 (purchased from Daiichi Sankyo) or Trastuzumab analog-MMAE (in-house) indicated in panel. (B) Body weight changes during treatment.

Antitumor activity of anti-human CD3/HER2 BsAbs (provided by a client) in B-hCD3E/HER2 mice. (A) Anti-human CD3/HER2 antibody BsAbs inhibited B-hHER2 MC38 plus tumor growth in B-hCD3E/HER2 mice. Murine colon cancer B-hHER2 MC38 plus cells were subcutaneously implanted into homozygous B-hCD3E/HER2 mice (female, 7-9-week-old, n=6). Mice were grouped when tumor volume reached approximately 100 mm³, at which time they were intraperitoneally injected with anti-human CD3/HER2 BsAbs Ab1 (provided by a client) indicated in panel. (B) Body weight changes during treatment. As shown in panel A, anti-human CD3/HER2 BsAbs Ab1 (provided by a client) was efficacious in controlling tumor growth in B-hCD3E/HER2 mice, demonstrating that the B-hCD3E/HER2 mice provide a powerful preclinical model for in vivo evaluation of anti-human CD3/HER2 BsAbs. Values are expressed as mean ± SEM.

Antitumor activity of anti-human CD3/HER2 BsAbs (provided by a client) in B-hCD3E/HER2 mice. (A) Anti-human CD3/HER2 antibody BsAbs inhibited B-hHER2 MC38 plus tumor growth in B-hCD3E/HER2 mice. Murine colon cancer B-hHER2 MC38 plus cells were subcutaneously implanted into homozygous B-hCD3E/HER2 mice (female, 7-9-week-old, n=6). Mice were grouped when tumor volume reached approximately 100 mm³, at which time they were intraperitoneally injected with anti-human CD3/HER2 BsAbs Ab1 (provided by a client) indicated in panel.