An -Acetylgalactosamino Dendron-Clearing Agent for High-Therapeutic-Index DOTA-Hapten Pretargeted Radioimmunotherapy.

Clearing agents (CAs) can rapidly remove nonlocalized targeting biomolecules from circulation for hepatic catabolism, thereby enhancing the therapeutic index (TI), especially for blood (marrow), of the subsequently administered radioisotope in any multistep pretargeting strategy. Herein we describe the synthesis and in vivo evaluation of a fully synthetic glycodendrimer-based CA for DOTA-based pretargeted radioimmunotherapy (DOTA-PRIT). The novel dendron-CA consists of a nonradioactive yttrium-DOTA-Bn molecule attached via a linker to a glycodendron displaying 16 terminal α-thio--acetylgalactosamine (α-SGalNAc) units (CCA α-16-DOTA-Y; molecular weight: 9059 Da).

Evaluation of CD20, CD22, and HLA-DR targeting for radioimmunotherapy of B-cell lymphomas.

Despite the promise of radioimmunotherapy using anti-CD20 antibodies (Ab) for the treatment of relapsed patients with indolent non-Hodgkin lymphoma (NHL), most patients treated with conventional doses of (131)I-tositumomab or (90)Y-ibritumomab eventually relapse. We did comparative assessments using conventional radioimmunotherapy targeting CD20, CD22, and HLA-DR on human Ramos, Raji, and FL-18 lymphoma xenografts in athymic mice to assess the potential for improving the efficacy of radioimmunotherapy by targeting other NHL cell surface antigens. Results of biodistribution studies showed significant differences in tumor localization consistent with variable antigenic expression on the different lymphoma cell lines.

A genetically engineered anti-CD45 single-chain antibody-streptavidin fusion protein for pretargeted radioimmunotherapy of hematologic malignancies.

Acute myelogenous leukemia (AML) currently kills the majority of afflicted patients despite combination chemotherapy and hematopoietic cell transplantation (HCT). Our group has documented the promise of radiolabeled anti-CD45 monoclonal antibodies (Ab) administered in the setting of allogeneic HCT for AML, but toxicity remains high, and cure rates are only 25% to 30% for relapsed AML. We now show the superiority of pretargeted radioimmunotherapy (PRIT) compared with conventional radioimmunotherapy using a recombinant tetravalent single-chain Ab-streptavidin (SA) fusion protein (scFv(4)SA) directed against human CD45, administered sequentially with a dendrimeric N-acetylgalactosamine-containing clearing agent and radiolabeled 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic (DOTA)-biotin.

Comparison of a tetravalent single-chain antibody-streptavidin fusion protein and an antibody-streptavidin chemical conjugate for pretargeted anti-CD20 radioimmunotherapy of B-cell lymphomas.

The efficacy of radioimmunotherapy (RIT) for patients with relapsed non-Hodgkin lymphoma (NHL) is limited by nonspecific delivery of radiation to normal tissues due to the long circulating half-life of radiolabeled anti-CD20 antibodies (Abs). Pretargeted RIT using a covalent conjugate of the 1F5 anti-CD20 Ab with streptavidin (SA) has been shown to augment the efficacy of RIT and decrease toxicity compared with a directly labeled 1F5 Ab. We have engineered a tetravalent singlechain 1F5 (scFv)4SA fusion protein and compared it to the 1F5-SA conjugate.

Intraperitoneal pretarget radioimmunotherapy with CC49 fusion protein.

Purpose: This study examined a pretarget radioimmunotherapy strategy for treatment of an i.p. tumor model (LS174T).

Pretargeted radioimmunotherapy of mesothelin-expressing cancer using a tetravalent single-chain Fv-streptavidin fusion protein.

Unlabelled: Mesothelin is a glycoprotein that is overexpressed in several human tumors, including mesotheliomas and ovarian cancers, and has been identified as a potential target for therapy. We evaluated the biodistribution and tumor-targeting ability of an antimesothelin tetravalent single-chain Fv-streptavidin fusion protein (SS1scFvSA) in mice.

Methods: SS1scFvSA was labeled with 125I or 111In for evaluation of internalization in vitro and for optimization of its biodistribution.

Single-chain Fv-streptavidin substantially improved therapeutic index in multistep targeting directed at disialoganglioside GD2.

Unlabelled: Multistep targeting can improve the therapeutic index of antibody-based targeting, particularly relevant to pediatric tumors where acute toxicity and late effects of treatment are major concerns. Neuroblastoma is uniquely suited for such investigations because of its abundance of surface ganglioside GD2.

Methods: 5F11scFv (scFv = single-chain variable fragment) was constructed from the variable regions of the heavy (V(H)) and kappa-light (V(L)) chain complementary DNA (cDNA) of anti-GD2 IgM hybridoma 5F11 and ligated to full-length streptavidin cDNA for expression in Escherichia coli.

Pretargeted alpha emitting radioimmunotherapy using (213)Bi 1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid-biotin.

Purpose: The use of an alpha emitter for radioimmunotherapy has potential advantages compared with beta emitters. When administered systemically optimal targeting of intact antibodies requires >24 h, therefore limiting the use of short-lived alpha emitters. This study investigated the biodistribution of bismuth-labeled biotin in A431 tumor-bearing mice pretargeted with antibody B3-streptavidin (B3-SA) and examined the therapeutic efficacy of the alpha emitter, (213)Bi-labeled biotin.

Biological comparison of 149Pm-, 166Ho-, and 177Lu-DOTA-biotin pretargeted by CC49 scFv-streptavidin fusion protein in xenograft-bearing nude mice.

The radiolanthanides (149)Pm, (166)Ho, and (177)Lu possess a range of half-lives and alpha(-) beta(-) energies for targeted radiotherapy of cancer. (149)Pm-, (166)Ho-, and (177)Lu-DOTA-biotin were pretargeted to LS174T colorectal tumors in nude mice with CC49 scFvSA antibody-streptavidin fusion protein. Tumor uptakes of (149)Pm (22.

In vivo evaluation of pretargeted 64Cu for tumor imaging and therapy.

Unlabelled: Pretargeting involves administration of a tumor-targeting monoclonal antibody (mAb) covalently linked to a molecule having a high-affinity binding site for a rapidly distributed radiolabeled effector molecule. The aim of this study was to compare pretargeting to a conventionally labeled antibody for tumor targeting of the intermediate-lived radionuclide (64)Cu, which has shown promise for PET imaging and radioimmunotherapy of cancer.

Methods: DOTA-biotin (where DOTA is 1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid) and the intact immunoconjugate DOTA-NR-LU-10 were labeled to high specific activities with (64)Cu, and the serum stabilities and target binding capabilities of each agent were assayed in vitro.

Pretarget radiotherapy with an anti-CD25 antibody-streptavidin fusion protein was effective in therapy of leukemia/lymphoma xenografts.

Although radioimmunotherapy with radiolabeled intact monoclonal antibodies has demonstrated efficacy in the treatment of lymphoma, it provides low tumor-to-normal-tissue radionuclide target ratios and unwanted prolonged radiation exposure to the bone marrow. To overcome these obstacles, the administration of the radionuclide was separated from that of the antibody by using an anti-IL-2 receptor alpha antibody single chain Fv-streptavidin fusion protein, followed by radiolabeled biotin to treat lymphoma or leukemia xenografted mice. This Pretarget approach provided extremely rapid and effective tumor targeting, permitting the use of short-lived alpha-emitting radionuclides.

Comparison of anti-CD20 and anti-CD45 antibodies for conventional and pretargeted radioimmunotherapy of B-cell lymphomas.

Radiolabeled anti-CD20 antibodies produce responses in 60% to 95% of patients with relapsed non-Hodgkin lymphoma (NHL); however, absorbed radiation ratios between tumors and normal organs are relatively low, and many patients have relapses. In this study we compared the abilities of anti-CD45 (BC8) and anti-CD20 (1F5) antibodies to target human Ramos lymphoma xenografts in athymic mice. When direct radioiodination was performed with conventional methods, BC8 delivered 2- to 4-fold more radioiodine to tumors than 1F5, with tumor-to-normal organ ratios as high as 20:1 using radiolabeled BC8 compared with a maximal ratio of 9.

Radioimmunotherapy of A431 xenografted mice with pretargeted B3 antibody-streptavidin and (90)Y-labeled 1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid (DOTA)-biotin.

We investigated the biodistribution of (88)Y/(111)In-labeled 1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid (DOTA)-biotin and therapy with (90)Y-labeled DOTA-biotin in tumor-bearing mice after B3-streptavidin antibody conjugate (B3-SA) pretargeting. B3 antibody, recognizing Lewis(y) antigen, was conjugated to streptavidin (B3-SA). For pretargeting, 400 micro g of the B3-SA was injected i.

Pretargeting radioimmunotherapy of a murine model of adult T-cell leukemia with the alpha-emitting radionuclide, bismuth 213.

We used a pretargeting technique to treat a nonobese diabetic/severe combined immunodeficient murine model of human adult T-cell leukemia with an anti-Tac antibody-streptavidin (HAT-SA) conjugate, which recognizes CD25, followed by bismuth 213 ((213)Bi)-1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid (DOTA)- biotin. In the 3-step pretargeting radioimmunotherapy protocol, HAT-SA (140 or 400 microg) was administered intravenously (i.v.