title: 111In-LLP2A-DOTA Polyethylene Glycol-Targeting {alpha}4{beta}1 Integrin: Comparative Pharmacokinetics for Imaging and Therapy of Lymphoid Malignancies.
abstract: N-[[4-[[[(2-ethylphenyl)amino]carbonyl]amino]phenyl]acetyl]-N(epsilon)-6-[(2E)-1-oxo-3-(3-pyridinyl-2-propenyl)]-l-lysyl-l-2-aminohexanedioyl-(1-amino-1-cyclohexane)carboxamide (LLP2A) is a high-affinity, high-specificity peptidomimetic ligand (inhibitory concentration of 50% = 2 pM) that binds the activated alpha4beta1 integrin found on a variety of malignant lymphoid cell lines. To better determine whether this ligand holds promise for imaging and therapy in lymphoid malignancies, 6 LLP2A derivatives, as LLP2A-1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (LLP2A-DOTA) and LLP2A-DOTA-polyethylene glycol (LLP2A-DOTA-PEG), were designed, synthesized, and radiolabeled with (111)In. Comparative pharmacokinetic studies in mice with Raji B-cell lymphoma xenografts were then complemented by small-animal PET of the lead molecular LLP2A format using (64)Cu-LLP2A-11-bis(carboxymethyl)-1,4,8,11-tetraazabicyclo[6.6.2]hexadecane ((64)Cu-LLP2A-CB-TE2A).
  
  
  METHODS
  LLP2A-DOTA and LLP2A-CB-TE2A were prepared using solid-phase synthesis; LLP2A-DOTA-PEG(2,000), LLP2A-DOTA-PEG(5,000), LLP2A-DOTA-PEG(10,000), (LLP2A-DOTA)(2)PEG(10,000), and (LLP2A-DOTA)(4)PEG(10,000) were prepared by PEGylation. (111)In radiolabeling of DOTA and (64)Cu radiolabeling of CB-TE2A conjugates yielded 370-1,850 and 3,700-7,400 kBq/microg (10-50 and 100-200 microCi/microg), respectively. The pharmacokinetics of the six (111)In radioconjugates were studied in vivo using biodistribution data (4 and 24 h) and whole-body autoradiography (24 h) in mice with Raji tumor xenografts. (64)Cu-LLP2A-CB-TE2A was imaged (4 and 24 h) on a small-animal PET scanner in the same mouse model.
  
  
  RESULTS
  The highest tumor uptake in pharmacokinetic studies was obtained with LLP2A-DOTA and (LLP2A-DOTA)(4)-PEG(10,000). For (111)In-LLP2A-DOTA (1 nM) at 4 and 24 h after injection, ratios of tumor to blood and tumor to nontumor (normal) organ (T/NT) were 8 to 35:1 for all organs or tissue except the spleen, marrow, and kidney, which were between 2:1 and 1:1. Tetravalent (LLP2A-DOTA)(4)-PEG(10,000) (1.1 nM) had tumor uptake similar to the univalent LLP2A-DOTA but higher liver, marrow, and kidney uptake. The excellent T/NT of LLP2A was also demonstrated by small-animal PET with (64)Cu-LLP2A-CB-TE2A at both 4 and 24 h after injection; obvious spleen targeting was apparent, but little kidney or liver activity was observed.
  
  
  CONCLUSIONS
  Of the conjugates investigated, the univalent, non-PEGylated ligand (111)In-LLP2A-DOTA exhibited the best T/NT ratios and showed the greatest potential for imaging of alpha4beta1 in human lymphoma. Furthermore, this univalent non-PEGylated LLP2A format, as (64)Cu-LLP2A-CB-TE2A, demonstrated excellent tumor targeting by small-animal PET and warrants further investigation as an agent for the study of alpha4beta1 expression in human lymphoid malignancies.
authors.affiliation: Division of Hematology and Oncology, Department of Internal Medicine, UC Davis School of Medicine, Sacramento, California, USA. sjdenardo@ucdavis.edu
authors.forename: Sally J
authors.initials: SJ
authors.lastname: Denardo
authors.forename: Ruiwu
authors.initials: R
authors.lastname: Liu
authors.forename: Huguette
authors.initials: H
authors.lastname: Albrecht
authors.forename: Arutselvan
authors.initials: A
authors.lastname: Natarajan
authors.forename: Julie L
authors.initials: JL
authors.lastname: Sutcliffe
authors.forename: Carolyn
authors.initials: C
authors.lastname: Anderson
authors.forename: Li
authors.initials: L
authors.lastname: Peng
authors.forename: Riccardo
authors.initials: R
authors.lastname: Ferdani
authors.forename: Simon R
authors.initials: SR
authors.lastname: Cherry
authors.forename: Kit S
authors.initials: KS
authors.lastname: Lam
mesh_terms: D000818:Animals; D002051:Burkitt Lymphoma / Q000000981:diagnostic imaging* / Q000378:metabolism* / Q000532:radiotherapy; D045744:Cell Line, Tumor; D016503:Drug Delivery Systems / Q000379:methods; D005260:Female; D006801:Humans; D039041:Integrin alpha4beta1 / Q000378:metabolism*; D008657:Metabolic Clearance Rate; D051379:Mice; D008810:Mice, Inbred C57BL; D008819:Mice, Nude; D009928:Organ Specificity; D009942:Organometallic Compounds / Q000493:pharmacokinetics* / Q000627:therapeutic use; D011092:Polyethylene Glycols / Q000493:pharmacokinetics* / Q000627:therapeutic use; D011877:Radionuclide Imaging; D019275:Radiopharmaceuticals / Q000138:chemical synthesis / Q000493:pharmacokinetics; D014018:Tissue Distribution
chemical_list: D039041:Integrin alpha4beta1; D009942:Organometallic Compounds; D019275:Radiopharmaceuticals; D011092:Polyethylene Glycols