Zlatko P. Pavelic

Tumor angiogenesis in T1 oral cavity squamous cell carcinoma: Role in predicting tumor aggressiveness

Angiogenesis is necessary for tumor growth and metastasis. In breast and other cancers angiogenesis has been shown to correlate with tumor size, metastatic potential, and prognosis. Some studies of head and neck cancer have shown a similar correlation, although results are inconclusive. This study was performed to determine whether tumor angiogenesis can be used as a prognostic indicator for early oral cancers.

Prognostic Indicators for Squamous Cell Carcinoma of the Oral Cavity: A Clinicopathologic Correlation†

Fifty‐three patients with T1 squamous cell cancer of the floor of mouth and ventral surface of the tongue with a known clinical outcome were retrospectively analyzed and arbitrarily divided into “aggressive” and “nonaggressive” groups based on their clinical behavior. Various host and tumor factors were then evaluated in an attempt to determine whether the tumor behavior could have been predicted. The paraffin‐embedded tumor specimens were evaluated for tumor differentiation, tumor thickness and tumor invasion, microvessel density, and p53 expression. In addition, a composite morphologic grading score was obtained by combining cell differentiation, nuclear polymorphism, mitosis activity, depth of infiltration, type of infiltration, and lymphatic infiltration. No single technique appeared capable of identifying “aggressive” behavior, although possibly an evaluation of composite factors might show promise in the future.

The Loss of Heterozygosity in Retinoblastoma and p53 Suppressor Genes As a Prognostic Indicator for Head and Neck Cancer

Inactivation of tumor suppressor genes, including p53 and retinoblastoma(Rb), are commonly found in all cancers, including head and neck squamous cell carcinoma. Alterations at either p53 or Rb, however, are only weakly associated with tumor aggressiveness. In many cancers loss of heterozygosity (LOH) at multiple loci is associated with decreased survival. The polymerase chain reaction and highly informative microsatellite markers were used to compare DNA from matched sets of 63 head and neck squamous cell cancers and normal tissue for LOH at the p53 and Rb loci. At p53, 50 were informative, with LOH occurring in 19 (38%). Of the 57 that were informative at Rb, LOH occurred in 21 (37%). Of the 46 that were informative at both p53 and Rb, LOH occurred in 10 (22%) at both loci. When LOH for p53 and Rb individually was compared to stage, differentiation, and survival, there was no correlation. However, the patients with LOH at both loci had a significantly poorer survival (P = .009). This strongly supports the contention that simultaneous alterations of these two tumor suppressor genes favor tumor aggressiveness and can be used as a prognostic indicator.

Heterogeneity of c-myc expression in histologically similar infiltrating ductal carcinomas of the breast.

Anti-c-myc monoclonal antibody was used to evaluate the distribution of the c-myc protein in normal and tumor cells of infiltrating ductal carcinoma. A semiquantitative method for reporting immunohistochemical assay results (c-myc score) that enables correlations on a more quantitative basis was used in this study. HL-60 cells demonstrated the strongest nuclear staining when fixed in cold acetone (4° C) for 10 min. All 24 specimens of infiltrating ductal carcinomas of the breast and 7 of 11 samples of normal breast tissues studied revealed the presence of c-myc protein. The level of expression in normal breast tissue was much lower than that in breast cancer. Heterogeneity in expression was found within individual tumors and there were substantial differences in the level of expression among different tumors. The subcellular site of staining was predominantly nuclear, occasionally nuclear and cytoplasmic in the same cell, and rarely only cytoplasmic. All four patients with tumor cells located in close proximity to the ductal basement membrane and over-expressing c-myc protein had positive lymph nodes, suggesting that these tumors are more likely to metastasize.

The soft agar clonogenicity and characterization of cells obtained from human solid tumors by mechanical and enzymatic means

SummaryA two-step procedure for releasing cells from solid tumors has been applied to specimens of human melanoma, sarcoma, lung, colon, and breast carcinoma. The first population released mechanically has been compared with the population subsequently released enzymatically in tests of dye exclusion, ribonucleoside triposphate pool sizes, intactness of DNA, and clonogenicity in soft agar. While greater numbers of dye-excluding cells are released in the enzymatic step, and these cells have higher ribonucleoside triphosphate pools and more intact DNA, both populations contain clonogenic cells in approximately equal numbers.Several semisolid media were employed in tests of clonogenicity, and all methods employing an agar underlayer appeared satisfactory and approximately equivalent in cloning efficiency. The methyl cellulose upper layer system facilitated implanting of pooled colonies into nude mice, which resulted in growth in the nude host and marked increase in cloning efficiency when the cells were replanted into soft agar-methyl cellulose plates.A comparison of four different areas of individual tumor specimens was made with cells released enzymatically and measuring cell yield, dye exclusion, ATP pool size, and uptake and metabolism of 5-fluoropyrimidines. Only relatively small variations were seen from one area to the next, with trypan blue exclusion exhibiting the least variation, and metabolism of fluorinated pyrimidines showing the most.

Insulin-like growth factor family in malignant haemangiopericytomas: the expression and role of insulin-like growth factor I receptor

Haemangiopericytoma is a rare soft tissue tumour originating from the contractile pericapillary cells. Relatively little is known about its molecular pathogenesis. To address this issue, the insulin‐like growth factor family (IGFs) was analysed in 19 tumours collected from a human tumour bank network. Seven of the tumours were associated with severe hypoglycaemia. Of these, six were retroperitoneal and one was located in the leg. 3 out of the 19 tumours (15·8 per cent) were positive for insulin‐like growth factor I (IGF I) mRNA and 11 were positive for IGF II mRNA (57·9 per cent). Almost 90 per cent of haemangiopericytomas expressed IGF I receptor (IGF IR) mRNA (17 out of 19), five (26·3 per cent) expressed IGF binding protein 1 (IGF BP1), three (15·8 per cent) expressed IGF BP2, and four (21 per cent) exhibited IGF BP3 mRNA. All of the 14 haemangiopericytomas examined with regard to specific receptor binding were IGF IR positive, ranging from 1·2 to 16·2 per cent. Binding was much higher in IGF I/IGF IR positive tumours (15·3±0·7) than in IGF I negative/IGF IR positive tumours (5·1±3·3). The potential role of IGF IR as a growth promoting factor in malignant haemangiopericytoma was studied using antisense oligonucleotides and monoclonal antibody αIR3 that specifically inhibit IGF IR synthesis or activity. 10 µM IGF IR antisense oligonucleotides significantly inhibited the growth of haemangiopericytoma cells in culture, by around 50 per cent; monoclonal antibody against IGF IR (αIR3) also significantly inhibited proliferation. The data suggest that IGF IR may play an important role in the genesis and progression of malignant haemangiopericytomas. Copyright

The Role of p53 Tumor Suppressor Gene in Human Head and Neck Tumorigenesis

Molecular genetics has led to new insights into diagnosis and treatment of human cancer. The alterations of tumor suppressor genes like retinoblastoma, p53 and others may have an important role in tumorigenesis. Mutations of p53 have been found in a majority of human malignancies including head and neck cancer. The distribution of p53 is different between types of tumors, suggesting environmental exposure as a cause active factor. The p53 mutation in head and neck tumors is an early event and appears to have a hot spot region at codons 238-248. While mutation and loss of heterozygosity at p53 are important in the genesis of head and neck cancer, other mechanisms such as binding of viral and cellular proteins to p53 are also likely to play a role.

Utility of anti-carcinoembryonic antigen monoclonal antibodies for differentiating ovarian adenocarcinomas from gastrointestinal metastasis to the ovary

The distinction between primary ovarian tumors and metastatic cancers to the ovary is frequently ambiguous. Recently, we reported that the D-14 monoclonal antibody (MAb), which is directed against a specific epitope of carcinoembryonic antigen (CEA), always reacts with colorectal adenocarcinomas and only rarely with neoplasms of non-gastrointestinal origin [J. Cancer Res. Clin. Oncol. 116, 51-56 (1990)]. We report here an analysis of the reactivity of four different anti-CEA MAbs with formalin-fixed tissue sections of human primary and metastatic colorectal and ovarian carcinomas. The four monoclonal antibodies employed were D-14, CEJ065, ZCEA1, and SP-625. D-14, CEJ065, and SP-625 MAbs reacted with essentially every colorectal adenocarcinoma. In contrast, ZCEA1 was the least reactive and 10 tumor samples showed no reactivity to this MAb. All four anti-CEA MAbs demonstrated scarce immunoreactivity with ovarian carcinomas and appear to be useful for distinguishing between ovarian carcinoma and colorectal metastasis to the ovary. Adenocarcinomas of the stomach and breast were also examined to determine CEA reactivity with the D-14 MAb, since these tumors need to be considered in the differential diagnosis of an ovarian mass. The majority of stomach adenocarcinomas were immunoreactive. In contrast, only 3 of 36 breast carcinomas were weakly immunoreactive, indicating that D-14 MAb is of no assistance in identifying breast carcinoma metastasis to the ovary.

C-MYC detection in bone marrow biopsies

In the past the fixation procedures employed in the immunohistostaining of paraffin-embedded tissues with anti-myc protein antibodies were unsatisfactory since they permitted the leakage of the protein from its normal nuclear localization in the cell. In the studies described here 3 different fixation methods were compared and it was shown that only one, the AMeX method, was suitable for studying c-myc expression in bone marrow biopsies.

The Role of the p53 Tumor Suppressor Gene in the Tumorigenesis of Inverting Papilloma of the Nose and Paranasal Sinuses

Inverting Papilloma (IP) is a rare neoplasm of the nose and paranasal sinuses. It is considered to be a premalignant lesion as there is a 7–21% incidence of squamous cell carcinoma (SCC) associated with IP. Although there have been many attempts to assign prognostic significance to various features of IP, there has not been a single reliable prognostic indicator identified. Recently it has been shown that mutations of the p53 tumor suppressor gene (TSG) are commonly involved in the process of cancer development. It has been assumed that cells which stain positive with p53 monoclonal antibody (MAb) contain mutant protein due to its lengthened half-life. To better understand the relationship of IP and carcinoma, we analyzed tumor specimens from 12 patients for p53 gene alterations using immunohistochemistry and DNA sequencing. Seven patients had IP without dysplasia, and five patients had IP with dysplasia or squamous cell carcinoma (SCC). All seven patients with IP only had tumors negative for p53 TSG. Three of five patients with IP and dysplasia or SCC stained positive for p53 TSG. No gene alterations of p53 TSG were detected in this study. The role and significance of p53 TSG in the tumorigenesis of IP is discussed based on these findings.