We are starting to see publications that address the reproducibility issues from the start. Using more rigorous protocols, having two different teams run parallel studies, doing 'em over with different lots of chips, etc. IMO, these will go a long way towards dealing with the skepticism about the reliability of the SELDI method, and towards supporting the stock price. But again, until the new system is launched and selling, we're not going to see significant appreciation. However, studies like these below should give prospective purchasers of the new systems additional confidence in their new toy, thus supporting the launch.
>>Dis Markers. 2003-2004;19(4-5):185-95.
Serum protein expression profiling for cancer detection: Validation of a SELDI-based approach for prostate cancer.
Grizzle WE, Adam BL, Bigbee WL, Conrads TP, Carroll C, Feng Z, Izbicka E, Jendoubi M, Johnsey D, Kagan J, Leach RJ, McCarthy DB, Semmes OJ, Srivastava S, Srivastava S, Thompson IM, Thornquist MD, Verma M, Zhang Z, Zou Z.
University of Alabama at Birmingham, Birmingham, AL, USA.
Multiple studies have reported that analysis of serum and other bodily fluids using surface enhanced laser desorption/ionization time of flight mass spectroscopy (SELDI-TOF-MS) can identify a "fingerprint" or "signature" of spectral peaks that can separate patients with a specific disease from normal control patients. Ultimately, classification by SELDI-TOF-MS relies on spectral differences in position and amplitude of resolved peaks. Since the reproducibility of quantitation, resolution and mass accuracy of the SELDI-TOF-MS, or any high throughput mass spectrometric technique, has never been determined this method has come under some skepticism as to its clinical usefulness. This manuscript describes a detailed design of a three-phase study to validate the clinical usefulness of SELDI-TOF-MS in the identification of patients with prostatic adenocarcinoma (PCA). At the end of this validation study, the usefulness of the general SELDI-TOF-MS approach to identifying patients with PCA will be demonstrated and how it compares with PCA diagnosis by measuring prostate specific antigen.<<
>>Prostate. 2004 Sep 1;60(4):325-31.
Proteomic analysis of dunning prostate cancer cell lines with variable metastatic potential using SELDI-TOF.
Gretzer MB, Chan DW, Van Rootselaar CL, Rosenzweig JM, Dalrymple S, Mangold LA, Partin AW, Veltri RW.
The Brady Urological Institute, Johns Hopkins Medical Institutions, Baltimore, Maryland.
BACKGROUND: Surface enhanced laser desorption and ionization-time-of-flight (SELDI-TOF) is an evolving proteomic technology for improving biomarker discovery that allows for rapid and sensitive analysis of complex protein mixtures generated from body fluids, cells, and/or tissues. SELDI-based profiling identifies unique, differentially expressed proteins relating to specific cancer-related disease states. We utilized SELDI-TOF following pre-processing with molecular separation and chemical fractionation of cell membrane extracts from three Dunning rat prostate cancer cell lines of varying metastatic potential to search novel proteins that are differentially expressed. METHODS: Dunning rat cell sublines of variable (%) metastatic potential; G (0%), AT-1 (20%), and Mat-Ly-Lu (100%) were cultured in two different laboratories. Cell lysis was performed in a homogenation buffer (320 mM sucrose/50 mM Tris/0.5 mM PSMF) using Dounce homogenation. After centrifugation, the membrane pellet was washed 2x and then solublized in 2% CHAPS/8 M urea. This sample was further processed using positive pressure molecular ultrafiltration at 30 kDa or precipitation with 50% ammonium sulfate. Next, each sample was applied to an IMAC3-Ni ProteinChip (Ciphergen Biosystems, Freemont, CA) and analyzed using Ciphergen's Protein Biology System with protein peak analysis software. RESULTS: SELDI-TOF analysis differentiated the three Dunning rat cell sublines based upon protein concentration normalized profiles between 5,000 and 20,000 Da. The preparations from the three cells lines showed clear differences when the extracts from the metastatic sublines (AT-1 and MLL) were compared to the benign subline (G) for proteins with molecular weights of 9 kDa (decrease), 12 kDa (significant decrease), 14 kDa (decrease), and 17 kDa (significant gain). After pre-processing extracts with ammonium sulfate and molecular ultrafiltration, the molecular profile changes from one subline to the next became more apparent. Our results were reproducible using multiple runs including from Dunning cells cultured in a separate laboratory, and using different lots of SELDI ProteinChips. CONCLUSIONS: The application of SELDI-TOF to a series of Dunning rat prostate cancer cell lines illustrated apparent changes in protein profiles among the three cell lines with known differences in metastatic biologic activity. SELDI-TOF identified four reproducible changes in protein expression in the AT1 and MLL metastatic cell sublines. Three of the expression changes were manifested as decreases, but one protein (17 kDa) was over-expressed in the AT1 and MLL cell lines. Emphasis will be placed on the isolation, purification, and characterization of the 17 kDa over-expressed protein and its potential role in PCa metastasis.<<
Power3 is about to run trials for a breast cancer test. Thinking prostate might be the next target for CIPH, but this is just a slightly educated guess right now. CIPH has the best commercialization terms with Hopkins and with Eastern Virginia Medical School, both of which are well along in pursuing prostate cancer biomarkers with SELDI.
Cheers, Tuck |
