Patricia B. DeRose

Tumor angiogenesis as a prognostic factor in oral cavity tumors

BACKGROUND Lymph-node metastasis is the single greatest predictor of survival in patients with oral cavity cancers. Tumor angiogenesis has been correlated with metastasis in breast cancer and may have prognostic value in other tumors. PATIENTS AND METHODS Sixty-six patients with clinically node-negative oral cavity squamous cell cancers were reviewed. Samples were cut and stained for factor VIII. The percentage of area of tissue stained for factor VIII was quantitated by a computerized image analyzer. Tumor depth was measured with an ocular micrometer to the nearest 0.1 mm. Variables were statistically examined against regional recurrence. RESULTS The probability of metastasis (%) was 2 for tumor staining of < or = 10% and 93 for tumor staining > 10% (P < 0.0001). The tumor depth was < or = 4 mm in 10 and > 4 mm in 83 (P < 0.0001). Patients with < or = 4 mm and < or = 10% staining had a 2% rate of recurrence, and patients with > 4 mm and > 10% staining had a 100% rate of recurrence (P < 0.0001). CONCLUSION Although tumor thickness was suggestive of predictability, only angiogenesis was a statistically significant predictor of recurrence in a multivariate analysis (P < 0.0001). Angiogenesis showed a strong correlation with regional recurrence and may be used as an independent prognostic indicator.

Tumor angiogenesis as a prognostic factor in laryngeal cancer.

BACKGROUND Lymph node metastasis is the single greatest predictor of recurrence in laryngeal cancer. Prognostic factors are needed to target patients who may benefit from adjuvant therapy. Tumor angiogenesis correlates with metastasis in breast, bladder, and oral cavity cancer and may have prognostic value in other tumors. METHODS In order to examine the relationship of tumor angiogenesis to recurrence, 51 patients with squamous cell carcinoma of the larynx were reviewed. In a blinded design, previously sectioned slides were chosen for advanced tumor and highest vessel concentration. Samples were cut and immunocytochemically stained for CD-31 (an endothelial marker). A computer image analyzer quantitated the percent area of staining. Variables were statistically examined against recurrence. RESULTS Patients were stratified by percent tumor staining. Nodal involvement was seen in 9 (36%) patients with tumor staining < or = 20% and in 20 (77%) with tumor staining > 20% (P = 0.003). Patients with < or = 20% staining and without metastasis had a 13% rate of recurrence whereas patients with > 20% staining and without metastasis had a 67% rate of recurrence (P = 0.025). CONCLUSIONS Though nodal status was suggestive of predictability, only angiogenesis is a statistically significant predictor of recurrence in node negative patients (P = 0.025). Angiogenesis shows strong correlation with regional recurrence and may be used as an independent prognostic indicator to determine clinically node negative patients who may be at higher risk for metastasis and require adjuvant therapy.

Sex and androgenic steroid receptor expression in hepatic adenomas

Sex hormones and anabolic-androgenic steroids are implicated in the development and progression of hepatic adenomas (HA). We studied the expression of their receptors in HA and adjacent liver. Archival tissue sections of 27 HA (16 resections, four needle biopsies, seven aspirations) from 18 patients, and the adjacent liver, were immunostained with monoclonal antibody to estrogen receptor (ER, 1/80) (Dako, Carpinteria, CA), progesterone receptor (PR, 1/50) (BioGenex, San Ramon, CA), and androgen receptor (AR, 1/80) (BioGenex). An avidin-biotin complex technique was used with microwave antigen retrieval. Nuclear expression was assessed as 1+ to 3+ intensity, with semiquantitation of the percentage of nuclei immunopositive. Five percent or more nuclei immunopositive was regarded as positive. The 18 patients included 16 females of 34 years mean age (range, 16 to 49) with an available history of oral contraceptives in five; the two men were 24 and 30 years, with no history of androgenic steroids. ER, PR, and AR were present in seven (26%) (1+/-2+ intensity, 5% to 10% of nuclei) of HA, seven (26%) (1+/-2+ intensity, 5% to 30% of nuclei) and nine (33%) (1+/-3+ intensity, 5% to 80% of nuclei), respectively. In the adjacent liver in 11 cases, there were one (9%) ER, (2+ intensity, 5% of nuclei), four (36%) PR (1+/-2+ intensity, 5% to 20% of nuclei), and two (18%) AR (2+/-3+ intensity, 10% of nuclei). Receptors are present and may mediate the action of sex hormones or androgenic steroids on HA and adjacent liver, but in less than one third of patients. This may have therapeutic implications.

Bcl‐2: Bax and Bcl‐2: Bcl‐x ratios by image cytometric quantitation of immunohistochemical expression in ovarian carcinoma: correlation with prognosis

Bcl-2 is a proto-oncogene which is involved in prolonging cell survival by inhibiting programmed cell death. Bax and bcl-x are members of the bcl-2 family; when overexpressed, they can counteract the ability of bcl-2 to inhibit apoptosis. This suggests a model in which the ratios of bcl-2 to bax and bcl-x can be used to determine response to therapy and prognosis. The expression of bcl-2, bax and bcl-x was studied in 50 ovarian carcinomas. The percentage of positive area immunostained (PPA) in the nucleus and cytoplasm of each ovarian carcinoma was quantitated in 15 high power fields by image cytometry. The ratios were obtained by dividing the PPA of bcl-2 by the PPA of bax and bcl-x. 17 of 50 ovarian carcinomas (34%) stained positively for bcl-2, 39 for bax (78%) and 47 for bcl-x (94%). Although there is no significant statistical correlation between expression of bcl-2, bax or bcl-x and grade (P = 0.15; P = 0. 47; P = 0.56), stage (P = 0.71; P = 0.6; P = 0.42), and overall or disease-free survival (P = 0.26; P = 0.55; P = 0.16), increased bcl-2 expression was demonstrated in patients with shortened overall and disease-free survival. Also, increased expression of bax and bcl-x was associated with increased overall and disease-free survival. Bcl-2:bax and bcl-2:bcl-x ratios less than 1 are associated with survival advantage, although not statistically significant (P = 0.83; P = 0.93). Image cytometric measurement of bcl-2, bax, and bcl-x expression is feasible. There is a tendency for their expression to correlate with prognosis in ovarian carcinomas.

Prognostic indicators in male breast carcinoma.

Abstract: Twenty‐nine male breast cancers (MBC) were studied to determine the relationship between expression of several prognostic factors and clinical outcome. Immunohistochemistry employing a labeled streptavidin‐biotin method was used to detect the presence of estrogen (ER) and progesterone receptors (PR), cathepsin D (CD), c‐erbB‐2 oncoprotein, epidermal growth factor receptor (EGFR), and p53; results were visually semiquantitated. DNA ploidy was evaluated by image analysis (CAS 200) of 5 μm fixed embedded Feulgenstained tissue sections. For proliferating cell nuclear antigen (PCNA), nuclear immunostain was quantitated as percentage positive nuclear area (PPNA) by image cytometry (CAS 200). The frequency of expression was ER, 26/29 (89.7%); PR, 19/29 (65.5%); CD, 25/29 (86.2%); c‐erbB‐2, 5/29 (17.2%); EGFR, 4/29 (13.8%); and p53, 9/29 (31%). Twenty‐one (72.4%) were aneuploid; the mean PPNA for PCNA was 37.87% (control 13%). Of 20 patients, 10 (50%) MBC had lymph node metastases; 6 (21%) had distant metastases to lung (1) and bone (5). Five of the patients died of MBC. Excluding the patients with only ductal carcinoma in situ, the 1‐and 5‐year survival rates were 90.5% and 56.3%, respectively. In this comprehensive study of a large number of available prognostic markers, their frequency (with the exception of higher ER and CD) and prognostic significance were similar to that in female breast carcinoma. Among clinical and standard pathologic unfavorable prognostic indicators, age ≥ 62 years was significant (p = .004). Trends toward reduced survival were associated with axillary lymph node metastases (p = .145), ER negativity (p = .058), PR negativity (p = .116), and aneuploid DNA content (p = .201).

DNA index and ploidy distinguish normal human parathyroids from parathyroid adenomas and primary hyperplastic parathyroids

BACKGROUND The goal of this study was to identify factors that might aid in diagnosis and intraoperative management of hyperparathyroidism. METHODS We analyzed biopsy specimens of 242 parathyroids from 159 patients by use of flow cytometry and image cytometry (ICM) for DNA index (DI), defined as the content of nuclear DNA compared with that expected for a DNA diploid standard, for proliferative index (PI), and for ploidy (diploid versus aneuploid or tetraploid). RESULTS True normal and normal parathyroids from patients with solitary adenomas were uniformly diploid. Abnormal ploidy (aneuploidy or tetraploidy) was identified frequently in adenomas and occasionally in hyperplasias with the exception that multiple endocrine neoplasia (MEN) biopsy specimens were uniformly diploid. DI for adenomas was similar to that for hyperplasias, and DI of both was higher than for normal glands. ICM-DI correlated positively with flow cytometry-DI and patient age and inversely with serum parathyroid hormone. PI was relatively low in all groups but was higher for hyperplasias versus normal parathyroids from patients with solitary adenomas and MEN versus non-MEN. PI correlated inversely with patient age. CONCLUSIONS DI by ICM differentiates normal from abnormal parathyroids. DI might influence extent of resection in two- and three-gland hyperplasia and selection of the most appropriate gland for autografting and cryopreservation in patients with four-gland hyperplasia.

IMAGE CYTOMETRIC COMPARISON OF PROLIFERATING CELL NUCLEAR ANTIGEN AND MIB-1 STAINING IN HEPATOCELLULAR CARCINOMA AND ADJACENT LIVER TISSUE

Studies of cellular proliferation may aid in elucidation of mechanisms of hepatocarcinogenesis and provide potential prognostic information in cases of hepatocellular carcinoma. Increased proliferation has been reported in the liver tissue adjacent to hepatocellular carcinoma and may be a risk factor for recurrence after tumor resection. We quantitated proliferating cell nuclear antigen (PCNA) clone PC10 and MIB-1 nuclear immunostain by image cytometry in fixed, paraffin-embedded tissue sections of normal, cirrhotic, and dysplastic livers with and without co-existent hepatocellular carcinoma and in the hepatocellular carcinoma. Fifteen high power fields were analyzed in each case using the CAS 200 image analyzer and results were expressed as the percent positive nuclear area. Cirrhotic liver showed significantly less proliferation adjacent to hepatocellular carcinoma than without hepatocellular carcinoma. There was a similar, but not significant, tendency for dysplastic liver. Increasing mean proliferation was noted from normal to cirrhotic to dysplastic liver to low grade to high grade hepatocellular carcinoma. These findings were similar using both PC10 and MIB-1. There tended to be slightly more staining with PC10 than with MIB-1, but the mean percent positive nuclear area was significantly greater with PC10 only in high grade hepatocellular carcinoma. Comparison of individual cases, however, yielded significant correlation between the two antibodies in the low grade and high grade hepatocellular carcinomas, but not in the benign livers. These findings demonstrate that image cytometric quantitation of PC10 and MIB-1 immunostain may be comparable measures of cellular proliferation and that there is no increase in proliferation in the hepatic tissue adjacent to hepatocellular carcinoma.

Melanoma Growth Stimulatory Activity in Primary Malignant Melanoma: Prognostic Significance

Malignant melanoma (MM) cells do not require all exogenous growth factors of normal melanocytes. It is hypothesized that they make their own growth factors including melanoma growth stimulatory activity (MGSA). Cultured melanoma cells respond to MGSA with increased growth and angiogenesis suggesting a role for MGSA in MM proliferation, differentiation, and progression. We assessed the prognostic significance of MGSA expression in 37 primary MM immunostained for MGSA. Immunostains were graded for intensity (0–3+), percentage of cells immunostained, and location of immunostain (intraepidermal, junctional, or dermal). In addition, 31 melanocytic and 23 dysplastic nevi were similarly studied for MGSA expression. All MM showed the presence of immunostain, 6 (16%) 1+, 12 (32%) 2+, and 19 (51%) 3+. Six (16%) had immunostain in ≤50% tumor cells, 31 (84%) in >50%. A significant number of MM showed >50% tumor cells staining at the dermal–epidermal junction compared with intraepidermal staining (P < .0001). Intensity and amount of immunostain did not correlate with Clarks or Breslows level. During a mean follow-up of 60 months (range: 5–101) on 27 patients, there were 4 local recurrences, 6 distant metastases, and 10 deaths. MGSA expression was not of prognostic significance with regard to survival (overall, disease free), or local recurrence or distant metastasis in primary MM. MGSA expression was similar in benign melanocytic and dysplastic nevi. Strong diffuse expression was noted in the junctional component of all junctional and most compound nevi. The dermal component consistently expressed less or no (in 45% of intradermal nevi) MGSA. MGSA expression does not correlate with prognosis in MM. Increased expression of MGSA at the dermal–epidermal junction in nevi and MM may indicate a role for MGSA in early local growth, before development of atypia.