Difference in Protein Expression Profile and Chemotherapy Drugs Response of Different Progression Stages of LNCaP Sublines and Other Human Prostate Cancer Cells

Abstract

Androgen ablation therapy is the primary treatment for metastatic prostate cancer. However, 80-90% of the patients who receive androgen ablation therapy ultimately develop recurrent tumors in 12-33 months after treatment with a median overall survival time of 1-2 years after relapse. LNCaP is a commonly used cell line established from a human lymph node metastatic lesion of prostatic adenocarcinoma. We previously established two relapsed androgen receptor (AR)-rich androgen-independent LNCaP sublines 104-R1 (androgen depleted for 12 months) and 104-R2 cells (androgen depleted for 24 months) from AR-positive androgen-dependent LNCaP 104-S cells. LNCaP 104-R1 and 104-R2 mimics the AR-positive hormone-refractory relapsed tumors in patients receiving androgen ablation therapy. Androgen treatment stimulates proliferation of 104-S cells, but causes growth inhibition and G1 cell cycle arrest in 104-R1 and 104-R2 cells. We investigated the protein expression profile difference between LNCaP 104-S vs. LNCaP 104-R1, 104-R2, PC-3, and DU-145 cells as well as examined the sensitivity of these prostate cancer cells to different chemotherapy drugs and small molecule inhibitors. Compared to 104-S cells, 104-R1 and 104-R2 cells express higher protein levels of AR, PSA, c-Myc, Skp2, BCL-2, P53, p-MDM2 S166, Rb, and p-Rb S807/811. The 104-R1 and 104-R2 cells express higher ratio of p-Akt S473/Akt, p-EGFR/EGFR, and p-Src/Src, but lower ratio of p-ERK/ERK than 104-S cells. PC-3 and DU-145 cells express higher c-Myc, Skp2, Akt, Akt1, and phospho-EGFR but less phospho-Akt and phospho-ERK. Overexpression of Skp2 increased resistance of LNCaP cells to chemotherapy drugs. Paclitaxel, androgen, and inhibitors for PI3K/Akt, EGFR, Src, or Bcl-2 seem to be potential choices for treatment of advanced prostate cancers. Our study provides rationale for targeting Akt, EGFR, Src, Bcl-2, and AR signaling as a treatment for AR-positive relapsed prostate tumors after hormone therapy.

Caffeic acid phenethyl ester induced cell cycle arrest and growth inhibition in androgen-independent prostate cancer cells via regulation of Skp2, p53, p21Cip1 and p27Kip1

“…Combination of higher dose of CAPE (20 or 40 μM) with LY294002 only showed additive but not synergistic suppressive effects. Expression of Bcl-2 in androgen-independent LNCaP 104-R1 cells is 65-fold higher that in parental androgen-dependent LNCaP 104-S cells at culture condition [21]. Bcl-2 expression may protect LNCaP 104-R1 cells from cell death under stress or drug treatment.…”

“…Compared to the parental LNCaP 104-S cells, 104-R1 cells is 28 fold more resistant to CAPE treatment. Under the culture condition, the protein abundance of Skp2, p21 Cip1 , p27 Kip1 , p53, total Akt, Akt1, Akt2, phospho-Akt S473, and phospho-Akt T308 are similar in 104-S and 104-R1 cells [21]. However, the protein level of Bcl-2 in 104-R1 cells is 65 fold higher than that in 104-S cells [21], and CAPE treatment caused very little inhibition on Bcl-2 level (Figure 12).…”

“…PC-3, LNCaP C4–2, 22Rv1, and DU-145 cells were purchased from Bioresource Collection and Research Center (Hsinchu city, Taiwan). PC-3, LNCaP C4–2, 22Rv1, and DU-145 cells were maintained in DMEM (Gibco/Invitrogen, Carlsbad, CA, U.S.A.) supplemented with 10% fetal bovine serum (FBS; Atlas Biologicals, Fort Collins, CO, U.S.A.), penicillin (100 U/ml), and streptomycin (100 μg/ml) as previously described [21]. …”

“…After 96 h of culture in the presence of different concentrations of CAPE, cells were processed as previously described [21, 26, 62]. Cell cycle profiles of LNCaP 104-R1, DU-145, 22Rv1, and LNCaP C4–2 cells were determined by flow cytometric analysis using a BD Facscan flow cytometer (BD Biosciences, San Jose, CA).…”

“…We discovered that CAPE treatment inhibited cell growth and induced G1 cell cycle arrest by suppressing c-Myc and Akt-related protein signaling networks in LNCaP 104-S and PC-3 cells [18–20]. However, the protein expression profile and response to treatment of chemotherapy drugs or kinase inhibitors was quite different between LNCaP 104-R1 and LNCaP 104-S cells [21]. We therefore used LNCaP 104-R1 cells as well as other CRPC cell lines 22Rv1, DU-145, and LNCaP C4–2 to determine the molecular mechanisms lying underneath of the anticancer effects of CAPE on CRPC cells.…”