Two mutations identified in the androgen receptor of the new human prostate cancer cell line MDA PCa 2a

Purpose: We have characterized the androgen receptor (AR) in a new human pr ostate cancer cell line, MDA PCa 2a, that has recently been established fro m a bone metastasis of a patient whose cancer exhibited androgen-independen t growth. Materials and Methods Androgen responsiveness of these cells was assessed b y measuring the effect of DHT and R1881 on cell growth and PSA secretion. S catchard analysis was used to characterize the affinity and abundance of AR protein. Using a PCR based strategy, genomic DNA of the entire coding regi on of AR gene was sequenced to identify possible mutations. Results: These cells express abundant AR (Nmax = 685 +/- 149 fmol./mg. prot ein), but the AR binding affinity (Kd) for DHT is only 25 nM, similar to 50 -fold lower affinity than the mutated AR in LNCaP prostate cancer cells (K- d = 0.5 nM) or the wildtype AR in MCF-7 breast cancer cells (Eh = 0.4 nM). Two mutations, L701H and T877A, were identified in the ligand binding domai n of the AR gene. Compared with LNCaP cells, the new cell line is significa ntly less responsive to DHT and R1881 as well as to other androgens such as testosterone, androstenedione, and DHEA. Similar to LNCaP cells, the Ligan d specificity of the AR in MDA PCa 2a cells appears to be relaxed and non-a ndrogens such as progesterone and estradiol act as agonists although with l ess potency than in LNCdP cells. Interestingly,in the absence of androgens, the new cell line expresses 15-fold higher baseline levels of PSA than LNC aP. Conclusions: Two mutations were identified in the AR gene of the MDA PCa 2a cell line that are likely responsible for the decreased androgen sensitivi ty and altered ligand specificity observed in these cells. Thus, this new c ell line with partial androgen responsiveness and PSA expression can serve as a functionally relevant model system of bone metastatic prostate cancer, and can be used to investigate the role of AR mutations in prostate cancer and its progression to androgen independence.