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Mass spectral analyses of labile DOTA-NHS and heterogeneity determination of DOTA or DM1 conjugated anti-PSMA antibody for prostate cancer therapy.

Lu SX, Takach EJ, Solomon M, Zhu Q, Law SJ, Hsieh FY

DMPK, Drug Safety and Disposition, Millennium Pharmaceuticals, Inc., Cambridge, Massachusetts 02139, USA.

Prostate specific membrane antigen (PSMA) is a well-characterized glycoprotein overexpressed on the surface of prostate cancer cells. The novel radiopharmaceutical 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA) radiolabeled with Yttrium (90Y) or Indium (111In) conjugated with anti-PSMA genetically engineered humanized monoclonal antibody (huJ591) has been investigated to target prostate cancer cells. The immunoconjugate of huJ591 with the analog of the cytotoxic drug maytansine, DM1 (N2'-deacetyl-N2'-(3-mercapto-1-oxopropyl)-maytansine) has also been developed at Millennium Pharmaceuticals. Activation of the DOTA molecule, resulting in 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid mono-(N-hydroxysuccinimidyl) ester (DOTA-NHS), allows conjugation with the anti-PSMA antibody through lysine residues in the antibody. The objectives of the study were to characterize the unstable chemical properties of DOTA-NHS before bioconjugation with huJ591, evaluate the binding profiles of DOTA to huJ591, and calculate trace metal elements (which may disturb 90Y or 111In labeling efficacy to the DOTA-huJ591 conjugate). A novel LC/MS/MS (Liquid Chromatography/Mass Spectrometry/Mass Spectrometry) quantitation method was developed to monitor the stability of DOTA-NHS in solid form and its bioconjugation chemistry reactions. Meanwhile, metal analysis was quantified by Inductively Coupled Plasma Mass Spectrometry (ICP/MS) to estimate the amounts of trace metals in DOTA-NHS and ensure radiolabeling efficiency of the conjugate at the radiopharmacy. MALDI-TOF MS (Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry) was used to identify levels of DOTA or DM1conjugation in DOTA-huJ591 and DM1-huJ591 conjugates, respectively.

Published 3 March 2005 in J Pharm Sci, 94(4): 788-97.
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