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Second primary cancer (SPC) is a term used to describe a new primary cancer occurring in patients who had formerly been diagnosed with tumor. Even though SPCs appear to be related to primary cancers, they are actually entities that have arisen independently and not as a result of recurrence. This report is of the first case in literature of a patient hospitalized for the surgical treatment of 3 synchronous Head and Neck Cancers. A 66-year-old male was admitted to our hospital (Ospedale Degli Infermi-Biella, Italy) complaining about pharyngodynia. Three different lesions were identified through endoscopic examination and narrow band imaging: the first one on left tonsil, the second one on epiglottis, and the third one on right aryepiglottic fold. The case was subject to a multidisciplinary team analysis due to its complexity, then the surgery consisted in (1) CO 2 laser left tonsillectomy, associated with (2) CO 2 laser excision of the lesion on epiglottis free edge, and (3) CO 2 laser excision of right aryepiglottic fold lesion. Synchronous tumors are among the most defiant challenges for surgeons since no international guideline specifies differentiated strategies to be adopted in patients affected by synchronous Head and Neck Cancers, therefore surgical planning must be tailored differently from patient to patient, and many unsolved questions still concern clinical treatments to be adopted.

Background: Triple-negative breast cancer (TNBC) is a very heterogeneous disease. Several gene expression and mutation profiling approaches were used to classify it, and all converged to the identification of distinct molecular subtypes, with some overlapping across different approaches. However, a standardised tool to routinely classify TNBC in the clinics and guide personalised treatment is lacking. We aimed at defining a specific gene signature for each of the six TNBC subtypes proposed by Lehman et al. in 2011 (basal-like 1 (BL1); basal-like 2 (BL2); mesenchymal (M); immunomodulatory (IM); mesenchymal stem-like (MSL); and luminal androgen receptor (LAR)), to be able to accurately predict them., Methods: Lehman's TNBCtype subtyping tool was applied to RNA-sequencing data from 482 TNBC (GSE164458), and a minimal subtype-specific gene signature was defined by combining two class comparison techniques with seven attribute selection methods. Several machine learning algorithms for subtype prediction were used, and the best classifier was applied on microarray data from 72 Italian TNBC and on the TNBC subset of the BRCA-TCGA data set., Results: We identified two signatures with the 120 and 81 top up- and downregulated genes that define the six TNBC subtypes, with prediction accuracy ranging from 88.6 to 89.4%, and even improving after removal of the least important genes. Network analysis was used to identify highly interconnected genes within each subgroup. Two druggable matrix metalloproteinases were found in the BL1 and BL2 subsets, and several druggable targets were complementary to androgen receptor or aromatase in the LAR subset. Several secondary drug-target interactions were found among the upregulated genes in the M, IM and MSL subsets., Conclusions: Our study took full advantage of available TNBC data sets to stratify samples and genes into distinct subtypes, according to gene expression profiles. The development of a data mining approach to acquire a large amount of information from several data sets has allowed us to identify a well-determined minimal number of genes that may help in the recognition of TNBC subtypes. These genes, most of which have been previously found to be associated with breast cancer, have the potential to become novel diagnostic markers and/or therapeutic targets for specific TNBC subsets., (© 2022. The Author(s).)

Purpose: Struma ovarii (SO) is a highly specialized ovarian teratoma, consisting of thyroid tissue. Rarely, carcinomas histologically identical to their thyroid counterparts may occur, and are comprehensively defined as malignant struma ovarii (MSO). Their optimal management is controversial, and the molecular profile of the malignant counterpart in the ovary is incompletely known. In this study, the clinicopathological and molecular features of six MSO from different Italian Institutions were analysed, to explore genetic profiles of potential therapeutic interest., Methods: The histopathological features and immunoprofile (according to the known markers Galectin-3, HBME1, cytokeratin 19 and CD56) were reviewed. In addition, all cases underwent genetic analysis with a next-generation sequencing (NGS) hot spot cancer panel detecting mutations in 50 genes involved in cancerogenesis. RET/PTC rearrangements and TERT promoter alterations were also evaluated., Results: Papillary carcinoma in all similar to its thyroid counterpart was found in five of six cases, including classical (two tumors) and follicular variant (three tumors) types. The last case was a poorly differentiated carcinoma. An activating gene mutation, was detected in five of six cases, including two NRAS, two BRAF, and one JAK3 oncogene mutations. No alterations were found in the other panel genes, nor in TERT promoter, or in RET chromosomal regions., Conclusions: MSO is a rare condition. Papillary carcinoma is the predominant malignant type, sharing both histomorphological and molecular features of its thyroid counterpart. Interestingly, the single case of poorly differentiated carcinoma displayed a JAK3 mutation. The presence of such driving mutation could be of potential interest in guiding postoperative treatment.

We have recently isolated a human gene, ROX, encoding a new member of the basic helix-loop-helix leucine zipper protein family. ROX is capable of heterodimerizing with Max and acts as a transcriptional repressor in an E-box-driven reporter gene system, while it was found to activate transcription in HeLa cells. ROX expression levels vary during the cell cycle, being down-regulated in proliferating cells. These biological properties of ROX suggest a possible involvement of this gene in cell proliferation and differentiation. The ROX gene maps to chromosome 17p13.3, a region frequently deleted in human malignancies. Here we report the genomic structure of the human ROX gene, which is composed of six exons and spans a genomic region of less than 40 kb. In an attempt to identify possible inactivating mutations in the ROX gene in human breast cancer, we performed a single-strand conformation polymorphism analysis of its coding region in 16 sporadic breast carcinomas showing loss of heterozygosity in the 17p13.3 region. No mutations were found in this analysis. Five nucleotide polymorphisms were identified in the ROX gene, three of which caused an amino acid substitution. These nucleotide changes were present in the peripheral blood DNAs of both the patients and the control individuals. In vitro translated assays did not show a significant decrease in the ability of the ROX mutant proteins to bind DNA or to repress transcription of a driven reporter gene in HEK293 cells. Despite experimental evidence that ROX might act as a tumor suppressor gene, our data suggest that mutations in the coding region of ROX are uncommon in human breast tumorigenesis.

Colorectal and breast cancers account for a significant number of deaths due to malignant neoplasia. Laboratory medicine plays an important role in the diagnosis and management of these tumors through the application of histopathology, immunohistochemistry, and serologic identification of tumor markers. Approximately 5% to 10% of colorectal and breast cancers result from an inherited predisposition. The genes responsible for most genetically transmitted cancers have been identified, and the application of findings from molecular pathology are being evaluated. This article reviews the genetic changes that occur as a result of somatic mutation and inherited or germline mutations.

The short area of chromosome 17 is a frequent target for deletions in human tumors, including breast cancer. We have investigated by restriction fragment polymorphism analysis the pattern of loss of heterozygosity (LOH) at four loci on 17p13.1-17pter in a panel of 110 primary human breast carcinomas. A copy of the p53 gene was lost in 23% of the informative cases. Point mutations in the p53 gene were statistically associated with LOH at the same locus (p = 0.003) but not at other loci on 17p13.3-17pter. A second region bordered by the loci D17S5/D17S28 (17p13.3) and D17S34 (17pter) is also affected by LOH, independent of point mutations in the p53 gene. We propose the presence of a second tumor suppressor gene within this region. In support of this hypothesis is the significant association (p = 0.005) between LOH at the D17S5/D17S28, but not at the TP53 or D17S34 loci, and tumors having a high S-phase index.

Fibroblast growth factors (FGFs) are related polypeptides with mitogenic activity on cells of mesodermal and neuroectodermal origin. The Fgf-3 gene shares high homology with FGF-2 and its protein product can substitute FGF-2 as a growth factor. Other observations, however, indicate that Fgf-3 has specialized functions. We have investigated the effect of the expression and secretion of Fgf-3 on the growth and transformation of the human breast epithelial cell line MCF-10A. Overexpression of Fgf-3 stimulates proliferation of these cells in serum-free medium and induces anchorage-independent colony formation in soft agar. In contrast, these effects were not observed with purified FGF-1 and FGF-2 on either the parental or the Fgf-3-MCF-10A cells. Thus, Fgf-3 is distinct from FGF-1 and FGF-2 for its ability to induce cell proliferation and transformation of MCF-10A cells. This difference could be due, at least in part, to the expression of a specific set of FGF receptors with higher affinity for FGF-3 than FGF-1 or FGF-2.

The expression of three epidermal growth factor (EGF)-related peptides, transforming growth factor alpha (TGF-alpha), amphiregulin (AR) and cripto-1 (CR-1), was examined by immunocytochemistry (ICC) in 68 primary infiltrating ductal (IDCs) and infiltrating lobular breast carcinomas (ILCs), and in 23 adjacent non-involved human mammary tissue samples. Within the 68 IDC and ILC specimens, 54 (79%) expressed immunoreactive TGF-alpha, 52 (77%) expressed AR and 56 (82%) expressed CR-1. Cytoplasmic staining was observed with all of the antibodies, and this staining could be eliminated by preabsorption of the antibodies with the appropriate peptide immunogen. Cytoplasmic staining with all of the antibodies was confined to the carcinoma cells, since no specific immunoreactivity could be detected in the surrounding stromal or endothelial cells. In addition to cytoplasmic reactivity, the AR antibody also exhibited nuclear staining in a number of the carcinoma specimens. No significant correlations were found between the percentage of carcinoma cells that were positive for TGF-alpha, AR or CR-1 and oestrogen receptor status, axillary lymph node involvement, histological grade, tumour size, proliferative index, loss of heterozygosity on chromosome 17p or overall patient survival. However, a highly significant inverse correlation was observed between the average percentage of carcinoma cells that expressed AR in individual tumours and the presence of a point-mutated p53 gene. Likewise, a significantly higher percentage of tumour cells in the ILC group expressed AR as compared with the average percentage of tumour cells that expressed AR in the IDC group. Of the 23 adjacent, non-involved breast tissue samples, CR-1 could be detected by ICC in only three (13%), while TGF-alpha was found in six (26%) and AR in ten (43%) of the non-involved breast tissues. These data demonstrate that breast carcinomas express multiple EGF-related peptides and show that the differential expression of CR-1 in malignant breast epithelial cells may serve as a potential tumour marker for breast cancer.

A series of 121 human breast tumors was screened for point mutations in exons 5 through 8 of the p53 gene, by SSCP analysis. On the same tumor samples, the S-phase index (SPI) was determined by the incorporation of BUdR in fresh tissue. p53 mutations were observed in 29% of the cases. The frequency of point mutations for the individual exons was: exon 5, 10.0%; exon 6, 9.9%; exon 7, 7.1% and exon 8, 5.5%. Two mutations detected by SSCP were confirmed by sequencing the p53 cDNA. The presence of a p53 mutation, irrespective of its location, correlates (p = 0.003) with a high SPI. This association appears to primarily reflect mutations in exon 5 (p = 0.0002) and exon 6 (p = 0.05), since mutations in exons 7 and 8 failed to show any association. These results indicate that mutations in the p53 gene identify highly proliferating tumors, and that the position of the p53 mutation may have different effects upon the proliferative activity of tumor cells in vivo.

The prognostic value of breast cancer proliferative activity was evaluated in 385 women operated for primary, non-metastasised mammary carcinoma. Cell kinetics was measured using two immunohistochemical techniques. Cells in S-phase of cell cycle were labelled in vitro by incubation of fresh tissue fragments with 5-bromo 2-deoxyuridine (BrdU), a thymidine analogue. Nuclei of cells in active DNA synthesis were stained by an anti-BrdU monoclonal antibody (Mab). Cells in interphase and mitosis were detected with Ki-67, a Mab that is known to react with a nuclear antigen present in G1/S/G2/M phases of cell cycle, but not in resting cells. This reagent provides a means of evaluating the growth fraction of neoplastic cells. BrdU was incorporated in a proportion of tumour cells ranging from 0.1 to 65.5% (median 6.8%). In the panel of tumours presented in this report the median percentage of Ki-67 positive cells (Ki-67 score) was 9.0% (range 0.1-77%). The relationship between disease-free survival (DFS), BrdU labelling index, Ki-67 score and 13 different clinico-pathological variables was investigated by multivariate analysis, using the Cox proportional hazards model. Axillary node status (P = 0.009) and Ki-67 score (P = 0.038) emerged as independent prognostic factors. Nodal status and tumour growth fraction allowed division of patients into groups at different risk of relapse: tumours with a proliferative index below the median value showed a lower recurrence rate than tumours with a high proliferative activity (P < 0.001). In particular, no relapse occurred in pN0 patients bearing carcinomas with a Ki-67 labelling < 9.0% 4 years after surgery. These findings suggest that the evaluation of proliferative activity in breast cancer enhances the probability of correctly predicting outcome after surgery and could be of assistance in the planning of adjuvant therapies.

We have undertaken a systematic study of primary human breast tumor DNA to identify and characterize frequently occurring somatic mutations. Loss of heterozygosity (LOH) has been the most frequent mutation in our panels of primary breast tumor DNA. It is currently thought that LOH reveals recessive mutations within the affected region of the genome. One goal of our studies has been to physically define the target genes revealed by LOH in primary breast tumors. We have focused our efforts on chromosome 17, finding five regions of the chromosome which are independently affected by LOH in breast tumors. Two apparent target loci are on chromosome 17p; one is the TP53 gene. The other is an as-yet undefined locus telomeric to the TP53 gene. Loss of expression of the nme1 gene on chromosome 17q in tumors was linked to patients with a poor prognosis (p = 0.018). Although a significant trend (p = 0.05) was found between LOH of the nme1 gene and loss of nme1 expression, no point mutations were found within the coding region of the nme1 gene by single strand conformational polymorphism (SSCP) or nucleotide sequence analysis. These and other results suggest to us that there may be potential tumor suppressor genes both centromeric and telomeric to the nme1 locus on chromosome 17q.

The capacity of breast tumor cells to proliferate is considered a potential prognostic factor together with other histopathologic parameters. The authors determined the proliferation index on a large panel of human primary breast tumors by measuring the levels of incorporation of bromodeoxyuridine (BrdU) by fresh tumor specimens in culture. Previous analysis showed that the percentage of cells entering the S-phase of the cell cycle strongly correlates with tumor grade, tumor size, and estrogen and progesterone receptor status. The capacity of tumor cells to proliferate might be associated with specific genetic mutations in primary tumors. To test this hypothesis, a panel of 96 human breast carcinomas, for which the BrdU labeling index (LI) was known, were tested for loss of heterozygosity (LOH) or increased copy number (ICN) at chromosomes 1q, 3p, 13q, 17p, and 18q. On chromosome 17p, LOH and ICN were observed in 27% and 12%, respectively, of the informative breast tumors. The LOH on chromosome 17p was significantly associated with tumors having an elevated BrdU proliferation index (P = 0.022). No association (P = 0.45) was observed between BrdU LI and tumor size (T2 + T3 compared with T1), tumor grade, and lymph node status. Increased copy number on chromosome 17p, LOH or ICN on 1q, and LOH on 13q14, 18q, and 3p also showed no significant correlation with cell kinetic parameters. These data are consistent with the presence of a gene or genes on chromosome 17p13 near the YNZ22.1 locus whose normal functioning is necessary for controlling breast tumor cells proliferation in vivo.

Twenty-six primary breast tumors were examined for mutations in the p53 tumor suppressor gene by an RNase protection assay and nucleotide sequence analysis of PCR-amplified p53 complementary DNAs. Each method detected p53 mutations in the same three tumors (12%). One tumor contained two mutations in the same allele. Single strand conformation polymorphism analysis of genomic DNA and complementary DNA proved more sensitive in the detection of mutations. Combining this technique with the other two a total of 12 mutations in the p53 gene were demonstrated in 11 tumors (46%), and a polymorphism at codon 213 was detected in another tumor. Loss of heterozygosity on chromosome 17p was detected by Southern blot analysis in 30% of the tumor DNAs. Not all of the tumors containing a point mutation in p53 also had loss of heterozygosity of the remaining allele, suggesting that loss of heterozygosity may represent a later event.

Transforming growth factor alpha (TGF alpha) is one growth factor that has been circumstantially implicated in regulating the autocrine growth of breast cancer cells. Expression of TGF alpha can be modulated by activated cellular protooncogenes such as ras and by estrogens. For example, the epidermal growth factor (EGF)-responsive normal NOG-8 mouse and human MCF-10A mammary epithelial cell lines can be transformed with either a point-mutated c-Ha-ras protooncogene or with a normal or point-mutated c-neu (erbB-2) protooncogene. In ras transformed NOG-8 and MCF-10A cells but not in neu transformed cells there is a loss in or an attenuated response to the mitogenic effects of EGF. This response may be due in part to an enhanced production of endogenous TGF alpha that is coordinately and temporally linked to the expression of the activated ras gene and to the acquisition of transformation-associated properties in these cells. TGF alpha mRNA and TGF alpha protein can also be detected in approximately 50-70% of primary human breast tumors. In addition, approximately 2- to 3-fold higher levels of biologically active and immunoreactive TGF alpha can also be detected in the pleural effusions from breast cancer patients as compared with the TGF alpha levels in the serous effusions of noncancer patients. Over-expression of a full-length TGF alpha cDNA in NOG-8 and MCF-10A cells is capable of transforming these cells. Finally, expression of TGF alpha mRNA and production of biologically active TGF alpha protein is also found in normal rodent and human mammary epithelial cells.

Transforming growth factor-alpha (TGF alpha) has been implicated in the autocrine growth control of a number of different rodent and human tumor cells, including breast cancer cells. Although TGF alpha has been detected in a limited number of normal tissues, its distribution and physiological function in the mammary gland are relatively unknown. TGF alpha mRNA expression was detected by in situ hybridization with a labeled TGF alpha antisense RNA probe and quantitated by application of computer-assisted digital image processing in both the ductal and alveolar epithelial cells in the virgin rat and nulliparous and parous human mammary glands. During pregnancy and lactation, the level of TGF alpha mRNA expression in the ductal and alveolar epithelial cells increased two- to threefold, while a heterogeneous yet strong expression of TGF alpha mRNA could also be detected in approximately 10-15% of the surrounding stromal cells in the pregnant mammary gland.

Transforming growth factor alpha (TGF alpha) mRNA expression was measured by Northern blot analysis in 18 human, primary, infiltrating, ductal breast carcinomas. Expression of a 4.8-kilobase TGF alpha mRNA transcript was detected in nine of 18 tumors. No evidence was observed of any gross amplifications or major rearrangements of the TGF alpha gene in the breast carcinoma specimens. Biologically active and immunoreactive TGF alpha was measured in the pleural effusions or in the ascitic fluids from 37 noncancer and 63 cancer patients. The TGF alpha activity detected ranged from 0.2 to 26 ng/mL in most effusions from both groups. However, 29 of 63 (46%) of the effusions from cancer patients exhibited TGF alpha levels that were 6 ng/mL or higher, whereas only seven of 37 (19%) of those from noncancer patients exceeded this level (P less than .03). In particular, effusions obtained from breast cancer patients showed a significantly higher level of TGF alpha, compared with those from noncancer patients (P less than .001). Effusions from 14 cancer patients also contained elevated levels of two tumor-associated antigens, CEA and/or TAG-72, and within this group, nine also had elevated levels of TGF alpha.

To determine whether the enhanced expression of transforming growth factor alpha (TGF alpha) is sufficient to induce the neoplastic transformation of an immortalized population of mammary epithelial cells, we cotransfected NOG-8 cells, a cloned mouse mammary epithelial cell line, with a simian virus 40-human TGF alpha cDNA expression vector plasmid and a pSV2neo plasmid. After cotransfection, nine G418-resistant NOG-8 colonies were cloned and expanded. All clones were subsequently analyzed for TGF alpha mRNA expression by northern blot analysis, TGF alpha secretion, anchorage-dependent growth in serum-free medium, anchorage-independent growth in soft agar, and tumorigenicity in nude mice. Three TGF alpha-transfected NOG-8 clones expressed high levels of a specific TGF alpha mRNA, secreted elevated levels of TGF alpha into the culture medium (177-595 ng/10(8) cells/48 h), exhibited an enhanced growth rate, grew aggressively as colonies in soft agar, and formed undifferentiated, invasive carcinomas in nude mice. A neutralizing mouse monoclonal antibody generated against the low molecular weight human TGF alpha peptide was able to inhibit colony formation in soft agar by TGF alpha-transfected NOG-8 clones that produced high levels by TGF alpha. This inhibition suggested that TGF alpha acted through an external autocrine loop. NOG-8 cells and NOG-8 cells transfected with a pSV2neo plasmid alone secreted very low levels of TGF alpha, failed to grow as colonies in soft agar and did not form tumors in nude mice. These results demonstrate that overexpression of a human TGF alpha cDNA in immortalized, nontransformed mouse mammary epithelial cells can induce a transformed phenotype in vitro and can facilitate tumor formation in vivo.

Gene amplification is a relatively frequent event in human malignant tumors and is believed to have an important function in neoplastic transformation and tumor progression. Our attention has been focused on the amplification and the expression of the int-2 gene for several reasons: (1) In the mouse mammary tumorigenesis int-2 is frequently activated by MMTV proviral integration. (2) The human homolog of int-2, located on chromosome 11q13, is frequently amplified in human primary tumors and is comprised in an amplification unit encompassing the hst gene, which is often coamplified; the amplification at the 11q13 locus in breast carcinomas correlates with a poor outcome of the disease. (3) int-2 and hst belong to the basic FGF gene family. All these observations raise the possibility that the human int-2 gene plays an active role in the neoplastic process, but this will prove to be true only if int-2 is expressed in human tumors. In the present study we used RNA:RNA in situ hybridization and Northern blot analysis to show that int-2 gene is expressed in a number of human carcinomas amplified at the same locus.

NOG-8 ras cells are a normal mouse mammary epithelial cell line transfected with a plasmid containing a glucocorticoid-inducible mouse mammary tumor virus long terminal repeat linked to the activated c-Ha-ras protooncogene. After addition of dexamethasone, there is a rapid induction (within 1-3 h) of p21ras protein that is concomitant with a parallel induction of the c-Ha-ras specific mRNA. After 4-6 days of dexamethasone treatment, NOG-8 ras cells are able to grow as colonies in semisolid medium. Between 9 and 12 days of dexamethasone treatment, there is a 5- to 6-fold increase of transforming growth factor alpha (TGF alpha) activity in the conditioned medium from NOG-8 ras cells. A 60-65% reduction in epidermal growth factor cell surface receptors on NOG-8 ras cells also occurs during this time interval. A 3- to 4-fold increase of the expression of a specific TGF alpha mRNA can be detected within 2 days of dexamethasone treatment, preceding the increase in TGF alpha protein found in the conditioned medium. Exogenous TGF alpha is able to stimulate in a dose-dependent fashion the anchorage-dependent and anchorage-independent growth of NOG-8 ras cells to a level comparable to that observed in dexamethasone treated ras-transformed NOG-8 ras cells. These results suggest that the enhanced expression of TGF alpha after induction of an activated ras protooncogene may be necessary for the anchorage-independent growth and subsequent morphological changes and the enhanced growth rate observed in ras-transformed mammary epithelial cells.

The prolactin, or lactogenic hormone, receptor has been purified (approximately 80%) from lactating mouse liver and human term placenta by the nondenaturing zwitterionic detergent 3-[(3-cholamidopropyl)-dimethylammonio]-1-propane sulfonate and a prolactin affinity column. The isolated "core-binding unit" has a molecular weight of 37,000 +/- 2,000 daltons. It retains the specificity for lactogenic hormones and binds prolactin with an affinity (Ka = 2 to 6 X 10(9) M-1) similar to that of the receptor as it occurs in its membranous environment (Ka = 3 to 5 X 10(9) M-1). Whether this "core-binding unit" exists on the cell surface in a cryptic or active form is influenced greatly by its association with other membrane proteins and the concentration of phosphatidylcholine within its local membranous environment.

A primary culture of serous cystadenocarcinoma of the ovary was used to study the expression of intermediate filament proteins and the deposition of basal lamina proteins. It was found that cells grown on type I and IV collagens or in collagen gels failed to express vimentin, which was readily demonstrable in cultures of the same cells grown on plastic or glass. Furthermore cells grown in collagen gels formed colonies demonstrating a cystic architecture. Unlike what is commonly observed on glass or plastic, where laminin and fibronectin are deposited as disorganized fibrils in the extracellular space, in or on collagen these proteins appear solely at the interface between the epithelial cells and matrix. The results suggest that the extracellular matrix influences the cytoskeletal organization of the intermediate filaments and determines cell polarity. They confirm that collagen substrates permit epithelial cell cultures to progress toward a more differentiated state.