title: Mass spectral analyses of labile DOTA-NHS and heterogeneity determination of DOTA or DM1 conjugated anti-PSMA antibody for prostate cancer therapy.
abstract: 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.
authors.affiliation: DMPK, Drug Safety and Disposition, Millennium Pharmaceuticals, Inc., Cambridge, Massachusetts 02139, USA.
authors.forename: Sharon X
authors.initials: SX
authors.lastname: Lu
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Takach
authors.forename: Marjorie
authors.initials: M
authors.lastname: Solomon
authors.forename: Qing
authors.initials: Q
authors.lastname: Zhu
authors.forename: Say-Jong
authors.initials: SJ
authors.lastname: Law
authors.forename: Frank Y
authors.initials: FY
authors.lastname: Hsieh
mesh_terms: D000906:Antibodies / Q000737:chemistry*; D000972:Antineoplastic Agents, Phytogenic / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D006573:Heterocyclic Compounds, 1-Ring / Q000032:analysis*; D006801:Humans; D007167:Immunotherapy; D000922:Immunotoxins / Q000737:chemistry* / Q000627:therapeutic use; D008297:Male; D013058:Mass Spectrometry; D020782:Matrix Metalloproteinases / Q000737:chemistry*; D008453:Maytansine / Q000737:chemistry*; D008970:Molecular Weight; D019999:Pharmaceutical Solutions; D017430:Prostate-Specific Antigen / Q000737:chemistry* / Q000276:immunology*; D011471:Prostatic Neoplasms / Q000628:therapy*; D011786:Quality Control; D012015:Reference Standards; D015203:Reproducibility of Results; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013388:Succinimides / Q000032:analysis*; D015021:Yttrium Radioisotopes
chemical_list: D000906:Antibodies; D000972:Antineoplastic Agents, Phytogenic; C502492:DOTA-NHS; D006573:Heterocyclic Compounds, 1-Ring; D000922:Immunotoxins; D019999:Pharmaceutical Solutions; D013388:Succinimides; D015021:Yttrium Radioisotopes; D008453:Maytansine; D017430:Prostate-Specific Antigen; C417183:Dm1 matrix metalloproteinase; D020782:Matrix Metalloproteinases