A review and proposed nomenclature for major proteins of the milk-fat globule membrane.

Summary Background Chemotherapy is the standard of care for advanced stages of non-small-cell lung cancer (NSCLC). TG4010 is a targeted immunotherapy based on a poxvirus (modified vaccinia virus Ankara) that codes for MUC1 tumour-associated antigen and interleukin 2. This study assessed TG4010 in combination with first-line chemotherapy in advanced NSCLC. Methods 148 patients with advanced (stage IIIB [wet] or IV) NSCLC expressing MUC1 by immunohistochemistry, and with performance status 0 or 1, were enrolled in parallel groups in this open-label, phase 2B study. 74 patients were allocated to the combination therapy group, and received TG4010 (10 8 plaque forming units) plus cisplatin (75 mg/m 2 on day 1) and gemcitabine (1250 mg/m 2 on days 1 and 8) repeated every 3 weeks for up to six cycles. 74 patients allocated to the control group received the same chemotherapy alone. Patients were allocated using a dynamic minimisation procedure stratified by centre, performance status, and disease stage. The primary endpoint was 6-month progression-free survival (PFS), with a target rate of 40% or higher in the experimental group. Analyses were done on an intention-to-treat basis. This study is completed and is registered with ClinicalTrials.gov, number NCT00415818. Findings 6-month PFS was 43·2% (32 of 74; 95% CI 33·4–53·5) in the TG4010 plus chemotherapy group, and 35·1% (26 of 74; 25·9–45·3) in the chemotherapy alone group. Fever, abdominal pain, and injection-site pain of any grade according to National Cancer Institute Common Toxicity Criteria were more common in the TG4010 group than in the chemotherapy alone group: 17 of 73 patients (23·3%) versus six of 72 (8·3%), 12 (16·4%) versus two (2·8%), and four (5·5%) versus zero (0%), respectively. The most common grade 3–4 adverse events were neutropenia (33 [45·2%] of patients in the TG4010 plus chemotherapy group vs 31 [43·1%] in the chemotherapy alone group) and fatigue (18 [24·7%] vs 13 [18·1%]); the only grade 3–4 events that differed significantly between groups were anorexia (three [4·1%] vs 10 [13·9%]) and pleural effusion (none vs four [5·6%]). 38 of 73 patients (52·1%) in the TG4010 plus chemotherapy group and 34 of 72 (47·2%) in the chemotherapy alone group had at least one serious adverse event. Interpretation This phase 2B study suggests that TG4010 enhances the effect of chemotherapy in advanced NSCLC. A confirmatory phase 2B–3 trial has been initiated. Funding Transgene SA, Advanced Diagnostics for New Therapeutic Approaches (ADNA)/OSEO.

https://www.thelancet.com/pdfs/journals/lanonc/PIIS1470-2045(11)70259-5.pdf

Therapeutic vaccination with TG4010 and first-line chemotherapy in advanced non-small-cell lung cancer: a controlled phase 2B trial

Mucins are a family of high molecular weight, highly glycosylated glycoproteins found in the apical cell membrane of human epithelial cells from the mammary gland, salivary gland, digestive tract, respiratory tract, kidney, bladder, prostate, uterus and rete testis. Increased synthesis of the core protein and alterations in the carbohydrates attached to these glycoproteins are believed to play important roles in the function and proliferation of tumour cells. Aberrant glycosylation leads not only to the production of novel carbohydrate structures, but also to the exposure of the core peptide. These novel epitopes may be candidates for diagnosis or therapy, by using either synthetic mucin fragments as vaccines, or monoclonal antibody-based reagents which detect these structures.

Mucins: structure, function, and associations with malignancy.

The isolation and characterization of complementary DNAs (cDNAs) which code for an epithelial antigen aberrantly expressed in human breast tumor tissue are described here. The only information regarding the primary structure of this potentially important antigen has been a 20-amino-acid repeat motif. We now report the complete amino acid sequences of different forms of the human epithelial tumor antigen as deduced from the nucleotide sequence of isolated non-repeat cDNAs. The diversity of protein forms is generated by a series of alternative splicing events that occur in the regions located upstream and downstream to a central tandem repeat array. Isolated cDNAs coding for the upstream region show that differential usage of alternative splice acceptor sites may generate two protein forms containing putative signal peptides of varying hydrophobicities. The complexity of possible antigen forms is further compounded by alternative splicing events occurring in the region 3' to the repeat array. The isolated cDNAs 3' to the tandem repeats indicate that whereas one mRNA transcript is colinear with the gene, and defines an open reading frame (ORF) containing 160 amino acids downstream to the repeat array, a second cDNA correlates with a mRNA that is generated by a series of splicing events. The deduced amino acid sequence of the spliced cDNA contains an ORF that is identical for 149 amino acids downstream to the repeat array with the amino acid sequence of the unspliced cDNA. At this point it diverges and continues for an additional 179 amino acids. The sequence contains a highly hydrophobic 28-amino-acid peptide, located towards the carboxyl terminus, that may correspond to a transmembrane region. The cDNAs and deduced amino acid sequences, presented here, define the complete amino acid sequences of the epithelial tumor antigen and demonstrate the existence of multiple protein forms that probably localize to different cellular and extracellular compartments.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1990.tb15511.x

Human epithelial tumor antigen cDNA sequences. Differential splicing may generate multiple protein forms.

Mucins are the structural components of the mucus gels that protect the respiratory, gastrointestinal, and reproductive tracts. These polydisperse glycoproteins (250,000 to 20,000,000 D) are approximately 80% carbohydrate on a mass basis and have a high intrinsic viscosity due to their large size and extreme hydrophilicity. Mucin oligosaccharides, the structures responsible for this hydrophilicity, are heterogeneous in size and structure but are chiefly O-linked, i.e., they initiate from N-acetylgalactosamine residues attached to threonine and serine residues of the polypeptide backbone. Our understanding of the structure of mucins has advanced rapidly in the last few years with the isolation and sequencing of cDNA clones that encode mucin polypeptide backbones. All currently well-characterized mucins have been found to contain extended arrays of tandemly repeated peptides rich in potential O-glycosylation sites. Less is known about the unique sequences that flank the tandem repeat arrays of secretory mucins, but currently available information indicates that these flanking regions contain cysteine-rich stretches that participate in mucin oligomer formation. Thus, secretory mucins appear to consist of oligomers containing heavily glycosylated domains flanked by unique sequences required for polymerization. Progress has also been made in characterizing the genes that encode mucins. At least four human mucin genes are known at present, although many others may remain to be discovered. Moreover, much work remains before we gain an understanding of the mechanisms involved in the expression of mucin genes and their tissue-specific regulation.

Mucin genes and the proteins they encode : structure, diversity, and regulation

A monoclonal antibody, H23, that specifically recognizes a breast-tumor-associated antigen, was used to isolate a cDNA insert that codes for the antigenic epitope. Nucleotide sequencing of this cDNA, as well as a longer 850-bp cDNA insert, shows that they are composed of 60-bp (G + C)-rich tandem repeating units. The coding strand was determined and codes for a proline-rich 20-amino-acid repeat motif. A comparison of the highly conserved repeat unit with the deduced flanking amino acid sequences demonstrates conservation of specific subregions of the repeat consensus within the flanking amino acids. Hybridization of the 60-bp cDNA probe with RNAs extracted from a variety of primary and metastatic human tumors yields relatively high levels of hybrid with the breast carcinomas, as compared to lower hybrid levels with RNAs from other epithelial tumors. RNA extracted from breast tissue adjacent to the tumor or from benign breast tumors, demonstrates low or undetectable levels of hybridization. Probing Southern blots with the 60-bp repeat shows that the tumor antigen is highly polymorphic and contains a variable number of tandem repeats (VNTRs). The VNTR nature of the gene was confirmed by probing Southern blots with unique genomic sequences that are physically linked to an isolated gene fragment that also contains the tandem repeat array. Mouse cells transfected with this gene fragment produce tumor antigen that is readily detected by H23 monoclonal antibodies. The allelic forms seen in 10 different primary human tumors demonstrate 100% concordance with the various mRNA species expressed. These studies are extended to the protein forms detected by immunoblot analyses that show both a correlation of the expressed tumor antigen species with the allelic forms as well as significantly increased expression in breast cancer tissue. The above studies unequivocally establish the over-expression of a VNTR gene coding for an epithelial tumor antigen in human breast cancer tissue.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1990.tb15512.x

A transcribed gene, containing a variable number of tandem repeats, codes for a human epithelial tumor antigen. cDNA cloning, expression of the transfected gene and over-expression in breast cancer tissue.

Sheila Zrihan

Papers

A transcribed gene, containing a variable number of tandem repeats, codes for a human epithelial tumor antigen. cDNA cloning, expression of the transfected gene and over-expression in breast cancer tissue.