CD74 is an integral membrane protein that functions as a MHC class II chaperone. Moreover, it has recently been shown to have a role as an accessory-signaling molecule and has been implicated in malignant B-cell proliferation and survival.These biological functions combined with expression of CD74 on malignant B cells and limited expression on normal tissues implicate CD74 as a potential therapeutic target. The anti-CD74 monoclonal antibody LL1 has been humanized (hLL1milatuzumab or IMMU-115) and can provide the basis for novel therapeutic approaches to B-cell malignancies, particularly because this antibody shows rapid internalization into CD74 + malignant cells. This article reviews the preclinical evaluations of LL1, its humanized form, and isotope, drug, and toxin conjugates. These studies show that unconjugated hLL1and conjugates of hLL1constructs with radioisotopes, doxorubicin, and frog RNase have high antitumor activity innon^Hodgkin's lymphoma and multiple myeloma invitro andin tumor xenograft models. Singledose studies of hLL1 in monkeys showed no adverse effects but did decrease circulating B and T lymphocytes and natural killer cells. When evaluated in combination with rituximab, either equivalent or improved efficacy, compared with either antibody alone, was observed. CD74 is a new candidate target for the immunotherapy of neoplasms expressing this antigen, which can be exploited using either a naked antibody or conjugated to isotopes, drugs, or toxins. CD74 (invariant chain, Ii) is a type II transmembraneglycoprotein that associates with the MHC class II a and h chains and directs the transport of the ahIi (invariant chain) complexes to endosomes and lysosomes (1 -4). In addition, CD74 is involved in signaling pathway functioning as a survival receptor (5). These biological functions, combined with expression of CD74 on malignant B cells and limited expression on normal tissues, implicate CD74 as a potential therapeutic target. The anti-CD74 monoclonal antibody (mAb) LL1 has been humanized (hLL1 milatuzumab or IMMU-115) and can provide the basis for novel therapeutic approaches to B-cell malignancies. This article reviews the preclinical evaluations of LL1, its humanized form, and isotope, drug, and toxin conjugates. The evidence suggests that hLL1 is a promising candidate antibody for the therapy of CD74-expressing malignancies, such as non -Hodgkin's lymphoma (NHL) and multiple myeloma (MM). Due to its rapid internalization property, hLL1 also shows considerable promise as a drug, isotope, or toxin immunoconjugate. CD74 Molecule: Structure, Function, and ExpressionCD74 has 30 NH 2 -terminal, intracytoplasmic amino acid residues, a 26-amino acid hydrophobic transmembrane region, and a 160-amino acid extracytoplasmic domain containing two N-linked carbohydrate chains (1, 6). Once synthesized, CD74, DRa, and DRh begin to associate within the endoplasmic reticulum. This is believed to occur by sequential addition of DRa/h heterodimers to a trimeric core of CD74 molecules until a nine-subunit...

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Pigmented melanoma cells and cultured melanocytes express a differentiation-related glycoprotein designated as pigmentation-associated antigen (PAA) of Mr 70,000-80,000. As described previously, PAA was initially defined by reactivity with antibodies in the serum of a patient with melanoma. Here we describe the production and characterization of a mouse monoclonal antibody to PAA. This antibody (TA99, an IgG2a) was shown by sequential immunoprecipitation experiments to react with the same component as the human antibody. Ab TA99 immunoprecipitated PAA from lysates of cells radiolabeled with [35S]methionine, [3H]glucosamine, [3H]fucose, and [3H]mannose as well as 125I. Using Ab TA99, the distribution of PAA was examined in frozen sections of 19 normal tissues and quantitatively in 68 tissue culture cell lines. In frozen sections, only melanin-containing cells were positive, including epithelial cells in the basal layer of the epidermis, in which pigment originates from melanocytes by transfer of melanosomes, and pigmented cells of the eye. In tissue culture cell lines, only pigmented melanoma cells were positive. PAA is an intracellular antigen, with a distribution very similar to that of melanosomes. This evidence confirms the close association of PAA with melanin production, and suggests that PAA may be a melanosome component. PAA was shown to be different from tyrosinase, the enzyme involved in melanin synthesis, but it was found to be identical to the previously recognized glycoprotein, gp75, characteristic of pigmented melanomas and melanocytes.

The high-frequency blood group antigen Ok(a) is carried on a red cell membrane glycoprotein (gp) of 35-69 kDa that is widely distributed on malignant cells of different origins. Immunostaining of hemopoietic cells and a range of normal human tissues demonstrated a wide distribution of the Ok(a) gp that appears to be nonlineage-restricted, although certain tissues show differentiation-related expression. Ok(a) gp was purified from red cell membranes by immunoaffinity chromatography using mAb A103 and amino acid sequence analysis was performed. The N-terminal 30 amino acids are identical to the predicted sequence of M6 leukocyte activation antigen (M6), a member of the Ig superfamily (IgSF) with two IgSF domains. There are homologs in rat (MRC OX-47 or CE9), in mouse (basigin or gp42), and in chicken (HT7 or neurothelin). The molecular basis of the Ok(a) mutation was established by sequencing M6 cDNA derived from normal and Ok(a-) EBV-transformed B cell lines. A point mutation in the translated portion of M6 cDNA, G331AG-->AAG gives rise to a predicted E92-->K amino acid change in the first Ig-like domain of the Ok(a-) form of the protein. Transfection of mouse NS-0 cells with normal or Ok(a-) cDNA confirmed the identity of the protein and only the Ok(a-) transfectants failed to react with monoclonal anti-Ok(a) Ab.

The specificity of five mouse monoclonal anti-A blood group antibodies (Ab), four of which were produced by immunization with cultured human cancer cells and one with a synthetic antigen, has been determined by examining their reactivity with purified A glycolipids, erythrocyte glycolipids, oligosaccharides, ovarian cyst glycoproteins, and salivary glycoproteins. Two of the antibodies (HT29-36 and CB) reacted with all A variant structures tested and have a broad anti-A reactivity. Ab CLH6 did not agglutinate A erythrocytes and reacted preferentially with the type 1A structure. Ab S12 agglutinated all A1 erythrocytes and reacted best with simple, monofucosyl type 2 A structures, such as Aa-2, Ab-2, and A tetrasaccharide. Ab M2 has a novel, but complex, spectrum of reactivity. It reacts with type 3 and type 4 A chains and not with type 1 and type 2 A chains. It appears to recognize both an external A structure (formula; see text) (I) (found) in type 3 and type 4 chains) and also an internal structure (II) found in type 3 chains. Ab M2 agglutinates all A and AB erythrocytes but does not react with salivary glycoproteins.