Monica Autiero

Differential antibody reactivity and CD4 binding of the mammary tumor marker protein GCDFP-15 from breast cyst and its counterparts from exocrine epithelia.

Analysis of biopsies from breast cancer patients demonstrated that GCDFP‐15 (gross cystic disease fluid protein‐15) is a specific immunocytochemical marker of primary and secondary apocrine breast tumors. The protein has an amino acid sequence identical to SABP (secretory actin‐binding protein), to PIP (prolactin‐inducible protein) and to gp17, a protein isolated from human seminal plasma. The latter was found to bind to CD4, a T‐cell co‐receptor involved in antigen recognition, thereby inhibiting the ability of the receptor to interact with the HIV‐1 envelope protein gp120. We compare here the ability of independently purified GCDFP‐15, SABP and gp17 and of recombinant PIP both to cross‐react with a panel of monoclonal antibodies (MAbs) raised against GCDFP‐15 or gp17, respectively, and to bind to CD4. We show that, although the various factors share the ability to bind to the panel of antibodies used, differences in the pattern of MAb recognition can be demonstrated. By comparing the kinetic constants for binding of GCDFP‐15 and gp17 to CD4 by biosensor technology, significant differences in binding affinities were observed between the 2 factors, thus reflecting structural differences. Surface plasmon resonance analysis also showed that anti‐GCDFP‐15 and anti‐gp17 antibodies inhibit the binding of CD4 to GCDFP‐15 and gp17, respectively, to different extents. Our data thus indicate that, while the various forms of the protein are encoded by the same cDNA, tissue specificities due to post‐translational modifications exist. This information may be relevant for developing more sensitive and accurate tests for the use of GCDFP‐15 as a diagnostic mammary tumor marker and, most importantly, raises the possibility that GCDFP‐15 may constitute a breast tumor‐specific antigen. Int. J. Cancer 78:76–85, 1998.© 1998 Wiley‐Liss, Inc.

Intragenic amplification and formation of extrachromosomal small circular DNA molecules from the PIP gene on chromosome 7 in primary breast carcinomas.

The PIP gene is expressed in exocrine glands and, in pathologic conditions, in breast cysts and breast cancers exhibiting apocrine features. It is localized on the long arm of chromosome 7, a region frequently alterated in mammary tumors. We previously described an abnormal restriction pattern of the PIP gene in 33% of prostate carcinomas analyzed. Here, we analyze the structure of the PIP gene in primary breast carcinomas. We report that part of the 3′ end, including exon 3, intron C, two‐thirds of exon 4 and a small portion of intron B, is amplified and involved in the formation of extrachromosomal spcDNA molecules in 3/14 (21.4%) breast cancers analyzed. The involvement of a well‐defined intragenic region of a gene in the formation of spcDNA appears to be unprecedented. Since spcDNA has been suggested to serve as an enhancer of genetic instability, the PIP gene may be the target of genomic variability processes in breast cancer.

Abnormal restriction pattern of PIP gene associated with human primary prostate cancers.

The PIP gene, localized in the 7q34 region that contains a number of fragile sites such as FRA 7H and FRA TI, codes for gp17/PIP, a protein secreted by breast apocrine tumors. We analyzed the integrity of this gene in 20 tumors of the urogenital tract. We found rearranged EcoRI fragments in 5 of 15 primary prostate carcinomas. No rearrangement was found in normal prostates derived from five patients undergoing prostatocystectomy during treatment of bladder cancers. By Southern blot hybridization with PIP gene exon-specific probes, the rearrangements were mapped at or near the 3 end of the gene. These abnormalities were found, not only in the neoplastic cells invading the prostatic tissues, but also in seminal vesicles without histologic tumoral features. These data suggest a critical role of the PIP gene or neighboring genes in prostate cancer.

Downstream Sequence Adjacent to AUG Affects Translation of Chloramphenicol Acetyl Transferase in Eukaryotic Cells

The CAT gene is widely used as a reporter in eukaryotic systems because of the efficient translation of its mRNA. We report here that a sequence occurring in the CAT mRNA at +15 nucleotides from CAT AUG is essential for translation. This sequence includes a stem-loop structure, which, however, exhibits a calculated stability significantly lower than that required for a hairpin to act as an enhancer of translation in vitro. Replacement of this region with the corresponding sequence from mRNAs that are normally translated in eukaryotic systems drastically reduced translation of CAT in COS cells, although the consensus sequence around the AUG, known to be required for high-level translation initiation, was conserved. These observations may be relevant for the exploitation of the CAT reporter system for analysis of the mechanisms of translation initiation by means of fusion constructs.