CD34+ Stromal Cells/Telocytes as a Source of Cancer-Associated Fibroblasts (CAFs) in Invasive Lobular Carcinoma of the Breast.

Several origins have been proposed for cancer-associated fibroblasts (CAFs), including resident CD34+ stromal cells/telocytes (CD34+SCs/TCs). The characteristics and arrangement of mammary CD34+SCs/TCs are well known and invasive lobular carcinoma of the breast (ILC) is one of the few malignant epithelial tumours with stromal cells that can express CD34 or αSMA, which could facilitate tracking these cells. Our objective is to assess whether tissue-resident CD34+SCs/TCs participate in the origin of CAFs in ILCs. For this purpose, using conventional and immunohistochemical procedures, we studied stromal cells in ILCs (n:42) and in normal breasts (n:6, also using electron microscopy). The results showed (a) the presence of anti-CD34+ or anti-αSMA+ stromal cells in varying proportion (from very rare in one of the markers to balanced) around nests/strands of neoplastic cells, (b) a similar arrangement and location of stromal cells in ILC to CD34+SCs/TCs in the normal breast, (c) both types of stromal cells coinciding around the same nest of neoplastic cells and (d) the coexpression of CD34 and αSMA in stromal cells in ILC. In conclusion, our findings support the hypothesis that resident CD34+SCs/TCs participate as an important source of CAFs in ILC. Further studies are required in this regard in other tumours.

Correlation of ultrasound findings with histology, tumor grade, and biological markers in breast cancer.

BACKGROUND AND PURPOSE: Ultrasound has been used successfully to differentiate benign and malignant breast lesions. The aim of this study was to investigate the correlation between ultrasound and prognostic indicators in breast cancer such as histological type, tumor grade, and biological markers. MATERIALS AND METHODS: Ultrasound findings (shape, margin, orientation, boundary, echo pattern, posterior acoustic feature, and presence of calcifications) of 458 breast cancers were analyzed and correlated with the tumor type, tumor grade, and biological markers by univariate and multivariate logistic regression analyses. The biological markers were estrogen receptor, progesterone receptor, and HER-2/neu. RESULTS: Invasive cancers displayed more frequently an irregular shape, a not parallel orientation, and a hypoechoic or complex echo pattern than carcinoma in situ cases (p < 0.05). Poorly differentiated invasive cancers had more frequently not circumscribed margins, an abrupt boundary, and a hypoechoic or complex echo pattern than moderately/well differentiated cancers (p < 0.05). Estrogen or progesterone receptor negative cancers more often displayed a hypoechoic or complex echo pattern and HER-2/neu positive cancers had more calcifications (p < 0.05). CONCLUSION: Ultrasound pattern is correlated with tumor type, tumor grade, and biological markers in breast cancers and it may be useful for prediction of prognosis.

Cytological features of lobular carcinoma of breast: how important are the intracytoplasmic lumina?

Lobular carcinoma is the most common pitfall in the cytodiagnosis of breast lesions. Of the cytological features described intracytoplasmic lumina are stressed by many authors as an important diagnostic feature. The present study aims at evaluating the importance of intracytoplasmic lumina in the cytodiagnosis of lobular carcinoma along with other cytological features. Fine needle aspirates from 11 cases of histologically proven classical lobular carcinomas of breast were reviewed. It was observed that the low cellularity, dispersed dyscohesive small cells, mild pleomorphism, nuclear moulding and Indian filing were the most important cytologic features which aid in the diagnosis of lobular carcinoma where as intracytoplasmic lumina were of little significance.

Lobular carcinoma with cytoplasmic granules.

We report on a case of invasive lobular carcinoma of the breast with a previously undescribed cytologic feature. Diff-Quik-stained cytologic preparations showed uniform single cells with prominent coarse cytoplasmic granules. Ultrastructurally, the granules showed features suggestive of autophagosomes and/or degenerative mitochondria. The cytologic differential diagnosis included granulocytic sarcoma, metastatic melanoma, extramedullary hematopoiesis, large granulocytic leukemia/lymphoma, and mast-cell tumor. Adjunctive studies were helpful in the diagnosis of carcinoma. Histologic study of the mastectomy specimen showed classic type of invasive lobular carcinoma.

Non-invasive lobular carcinoma within a fibroadenoma, a preoperatively diagnosed case.

Breast cancer within a fibroadenoma is rare and usually diagnosed postoperatively from pathological specimens. This paper reports a 54-year-old female with non-invasive carcinoma within a fibroadenoma, diagnosed preoperatively. She underwent a medical examination and mastopathy was suspected. On physical examination a mass 2 cm in diameter was palpated in the left breast. Ultrasonography showed a mass with smooth margins and uniform internal echoes, but cytology showed malignancy. Mammography showed a round mass with distinct margins and no calcification. As fibroadenoma, diagnosed by ultrasonography and mammography, and breast cancer, diagnosed by cytology, were not consistent results several core biopsies were performed. Needle biopsy showed proliferation of atypical epithelial cells; breast cancer within a fibroadenoma was diagnosed. MRI showed a circular mass with distinct, smooth margins and in a dynamic study, the mass showed irregular staining and the presence of early staining. Left lumpectomy and dissection of the left axillary lymph nodes was performed. Histological examination showed non-invasive lobular carcinoma occurring within a fibroadenoma.

Cytokeratin 8 immunostaining pattern and E-cadherin expression distinguish lobular from ductal breast carcinoma.

Immunohistochemistry using antibodies to cytokeratin 8 can serve as a valuable diagnostic tool for the differentiation of lobular from ductal carcinomas of the breast. In contrast with ductal carcinomas, which exhibit a peripheral-predominant immunostaining pattern, adjacent tumor cells "molding" to each other, lobular carcinomas exhibit a ring-like perinuclear immunostaining pattern, creating a "bag of marbles" appearance with neighboring tumor cells. This immunostaining pattern is stable even in the tumors that otherwise do not exhibit characteristic histomorphologic features (i.e., solid or pleomorphic type of a lobular carcinoma) and tumors that mimic growth patterns characteristic of the respective other tumor type (i.e., targetoid or single-file growth pattern in a ductal carcinoma). Furthermore, we demonstrate that ductal carcinomas express E-cadherin in a similar peripheral-predominant immunostaining pattern (33/33 cases), while all 15 lobular carcinomas were negative for E-cadherin, suggesting a role for E-cadherin in the architectural organization of the cytoskeletal scaffolding within the tumor cells.

Malignant phyllodes tumor in the right breast and invasive lobular carcinoma within fibroadenoma in the other: case report.

CONTEXT: The malignant variety of the phyllodes tumor is rare. The occurrence of invasive lobular carcinoma within fibroadenoma is rare as well. DESIGN: Case report. CASE REPORT: A 58-year-old black female patient was referred to the Mastology unit of the Department of Gynecology, Federal University of São Paulo / Escola Paulista de Medicina, in February 1990, presenting an ulcerated tumor in the right breast with fast growth over the preceding six months. She was a virgin, with meno-pause at the age of 45 years and had not undergone hormone replacement treatment. The physical examination showed, in her right breast, an ulcerated tumor of 20 x 30 cm which was not adher-ent to the muscle level, multilobular and with fibroelastic consistency. The axillary lymph nodes were not palpable. The left breast showed a 2 x 3 cm painless, movable nodule, with well-defined edges, and fibroelastic consistency. We performed left-breast mammography, which showed several nodules with well-defined edges, the largest being 2 x 3 cm and exhibiting rough calcification and grouped microcalcifications within it. The patient underwent a frozen biopsy that showed a malignant variant of the phyllodes tumor in the right breast and fibroadenoma in the left one. After that, we performed a total mastectomy in the right breast and an excision biopsy in the left one. Paraffin study confirmed the frozen biopsy result from the right breast, yet we observed that in the interior of the fibroadenoma that was removed on the left, there was a focal area of invasive lobular carcinoma measuring 0.4 cm. The patient then underwent a modi-fied radical mastectomy with total axillary lymphadenectomy. None of the 21 dissected lymph nodes showed evidence of metastasis. In the follow-up, the patient evolved asymptomatically and with normal physical and laboratory examination results up to July 1997.

Malignant phyllodes tumor in the right breast and invasive lobular carcinoma within fibroadenoma in the other: case report

CONTEXT: The malignant variety of the phyllodes tumor is rare. The occurrence of invasive lobular carcinoma within fibroadenoma is rare as well. DESIGN: Case report. CASE REPORT: A 58-year-old black female patient was referred to the Mastology unit of the Department of Gynecology, Federal University of Sao Paulo / Escola Paulista de Medicina, in February 1990, presenting an ulcerated tumor in the right breast with fast growth over the preceding six months. She was a virgin, with meno-pause at the age of 45 years and had not undergone hormone replacement treatment. The physical examination showed, in her right breast, an ulcerated tumor of 20 x 30 cm which was not adher-ent to the muscle level, multilobular and with fibroelastic consistency. The axillary lymph nodes were not palpable. The left breast showed a 2 x 3 cm painless, movable nodule, with well-defined edges, and fibroelastic consistency. We performed left-breast mammography, which showed several nodules with well-defined edges, the largest being 2 x 3 cm and exhibiting rough calcification and grouped microcalcifications within it. The patient underwent a frozen biopsy that showed a malignant variant of the phyllodes tumor in the right breast and fibroadenoma in the left one. After that, we performed a total mastectomy in the right breast and an excision biopsy in the left one. Paraffin study confirmed the frozen biopsy result from the right breast, yet we observed that in the interior of the fibroadenoma that was removed on the left, there was a focal area of invasive lobular carcinoma measuring 0.4 cm. The patient then underwent a modi-fied radical mastectomy with total axillary lymphadenectomy. None of the 21 dissected lymph nodes showed evidence of metastasis. In the follow-up, the patient evolved asymptomatically and with normal physical and laboratory examination results up to July 1997

AgNORs in breast tumours.

There is evidence that the quantitative distribution of AgNOR proteins is a proliferation-related parameter that can be used as a prognostic index in tumour pathology. In breast cancer, some authors found a significant prognostic correlation of AgNOR protein quantity, whereas other did not. However, in all the reports dealing with AgNOR area (as opposed to count) this parameter was always turned out to be an independent prognostic indicator. The present study tests the significance of AgNOR proteins in a large series of primary breast carcinomas, exploring the associations between the AgNOR protein amount, as evaluated by image cytometry, and the other well-established prognostic markers commonly considered for breast cancer, along with patients' survival. Our results demonstrated a highly significant association between AgNOR protein quantity and tumour prognosis. Moreover, when the AgNOR area values were entered in multivariate analysis together with the other predictive parameters commonly considered in breast carcinomas, they showed an independent prognostic value together with Ki67-labelling index (LI), N-status and tumour size. Considering node-negative and -positive cases separately, the AgNOR protein area and Ki67-LI both come out as a independent predictors only in the latter group: the short follow-up time of our series (36 months median) could be responsible for this discrepancy.

Immunohistochemical localization of metallothionein in human breast cancer in comparison with cathepsin D, stromelysin-1, CD44, extracellular matrix components, P53, Rb, C-erbB-2, EGFR, steroid receptor content and proliferation.

Metallothionein (MT) is a low molecular weight, cysteine-rich, zinc-binding protein that may have a function in cellular repair processes, growth and differentiation. Using a monoclonal antibody (E9) to metallothionein, we investigated the immunohistochemical expression of MT in routinely fixed and paraffin-embedded tissue from 98 cases of female breast carcinomas. The MT expression was studied in comparison with the expression of the basement membrane (BM) antigens (type IV collagen, laminin), fibronectin, cathepsin D, adhesion molecule CD44, p53 protein, the pRb, c-erbB-2 oncoprotein, EGFR, stromelysin-1, proliferation indices (Ki-67, PCNA), steroid receptor content as well as with other conventional clinicopathological parameters of breast cancer. Strong MT expression was observed in the majority of tumour cells in 18.4% of tumours, focal MT positivity in 13.3% and almost complete lack of MT expression in 68.4% of cases (mean value 33.36 +/- 26.36). The MT expression in carcinoma cells was strongly associated with the DCIS component of the tumour (p < 0.0001). High values of MT were correlated with low steroid receptor status (p = 0.08 for ER receptor and p = 0.019 for PgR receptor content). MT positive cases were correlated with stromelysin-1 expression (p = 0.059) and cathepsin D (p = 0.058). These findings suggest that MT expression is characteristic of the early phase of breast carcinogenesis, possibly regulated by hormones, and could be a new potential prognostic marker in breast cancer.

DNA analysis and S-phase fraction determination by flow cytometric analysis of infiltrating lobular carcinoma of the breast.

Flow cytometric analysis was performed on 50 infiltrating labular carcinomas (ILCs) of the breast from 50 patients with clinical follow-up (average duration, 4.1 yr) who had been treated between 1976 and 1991. The patients were classified as alive with no evidence of disease, alive with disease, or dead of disease. Ploidy and S-phase fraction (SPF) were compared with clinical outcome, histologic pattern (classical versus variant patterns), nuclear grade 1 or 2, axillary lymph node status, tumor size, percentage of signet ring cells, and estrogen receptor status. There was no association between aneuploid or diploid ILC and disease recurrence (of those patients classified as alive with no evidence of disease plus dead of disease, 4 (40%) of 10 were aneuploid and 15 (38%) of 40 were diploid), survival (of those classified as alive with no evidence of disease plus alive with disease, 9 (90%) of 10 were aneuploid and 36 (90%) of 40 were diploid), or any of the other factors evaluated. However, ILCs with a high SPF were more likely to recur than those with a low SPF (a high SPF was found in 9 [56%] of 16; a low SPF was found in 9 [30%] of 30), but this relationship was not statistically significant (P = 0.08). When only diploid ILCs were considered, there was a statistically significant association between high SPF and recurrence (a high SPF was found in 9 [64%] of 14, a low SPF was found in 6 [24%] of 25, P = 0.033). After the stratification of the diploid ILCs by the stage of disease, this relationship persisted only in Stage 1 (recurrence: a high SPF was found in 3 [75%] of 4; a low SPF was found in 1 [9%] of 11, P = 0.033). There was no association between SPF and any of the other factors evaluated. Our study indicates that ILC is usually diploid and that SPF may be a prognostic indicator that is limited to Stage 1 disease.

Distribution of thrombospondin and integrin alpha V in DCIS, invasive ductal and lobular human breast carcinomas. Analysis by electron microscopy.

The ultrastructural distribution of thrombospondin (TSP) and its cell surface receptor, integrin alpha V, was studied in two cases of human breast carcinoma: one of ductal carcinoma in situ (DCIS) with an invasive component, and one of invasive lobular carcinoma. In DCIS, moderate immunolabelling for TSP and integrin alpha V was observed in the rough endoplasmic reticulum and at the plasma membrane of intraductal carcinoma cells. TSP was also associated with extracellular matrix collagen fibrils surrounding in situ carcinoma cells. In the invasive part of this ductal carcinoma, most of the malignant cells were negative for TSP, while integrin alpha V was moderately expressed in these cells. In sharp contrast, typical strands of invasive lobular carcinoma cells in "Indian file" showed moderate TSP immunostaining in the rough endoplasmic reticulum and strong immunoreactivity for TSP at the plasma membrane and in the extracellular matrix. Moderate to strong immunoreactivity for integrin alpha V was also observed in invasive lobular carcinoma cells. Because of the role of TSP during cancer cell invasion, the different expression patterns of TSP in invasive ductal versus lobular carcinoma may well reflect biological differences between these two types of breast carcinoma and could account for the peculiar diffuse invasive behaviour of breast lobular carcinoma cells.

Tumour vascularity and basement membrane structure in breast cancer as related to tumour histology and prognosis.

A series of 202 breast cancer biopsy specimens were analysed immunohistochemically for collagen IV to demonstrate basement membrane (BM) structures and blood vessels within tumour tissue. Integrity of the BM was graded into four categories and the number of vascular channels per square millimetre of tumour tissue were counted. Defective BM structures were significantly related to high grade, lack of tubule formation, invasive disease, high S-phase fraction and variability in nuclear size and shape. High vascular channel density was related to poor tumour differentiation and a high proliferation rate of cancer cells as well as to the absence of tubule formation, inconspicuous intraductal growth and low progesterone receptor content. High vascular density and defective BM structures were signs of poor prognosis and short recurrence-free survival in the entire cohort and also in local tumours. In multivariate analysis, the vascular density had independent prognostic value, as did the diameter, axillary lymph node status and mitotic rate. The counting of vascular channels within the tumour provides additional prognostic information in breast cancer, in contrast to analysis of the BM integrity which shows hardly any prognostic information additional to that provided by the special histological features, e.g. tubule formation and intraductal growth pattern.

Prognostic, quantitative histopathologic variables in lobular carcinoma of the breast.

BACKGROUND: A retrospective investigation of 53 consecutively treated patients with operable lobular carcinoma of the breast, with a median follow-up of 6.6 years, was performed to examine the prognostic value of quantitative histopathologic parameters. METHODS: The measurements were performed in routinely processed histologic sections using a simple, unbiased technique for the estimation of the three-dimensional mean nuclear volume (vv(nuc)). In addition, quantitative estimates were obtained of the mitotic index (MI), the nuclear index (NI), the nuclear volume fraction (Vv(nuc/tis)), and the mean nuclear profile area (aH(nuc)). RESULTS: Estimates of vv(nuc), MI, NI, and Vv(nuc/tis) were of significant or marginally significant prognostic value in univariate analyses, whereas no prognostic significance was attributed to estimates of aH(nuc). In multivariate Cox analyses, the clinical stage of disease, vv(nuc), MI, and NI were of significant independent, prognostic value. On the basis of the multivariate analyses, a prognostic index with highly distinguishing capacity between prognostically poor and favorable cases was constructed. CONCLUSION: Quantitative histopathologic variables are of value for objective grading of malignancy in lobular carcinomas. The new parameter--estimates of the mean nuclear volume--is highly reproducible and suitable for routine use. However, larger and prospective studies are needed to establish the true value of the quantitative histopathologic variables in the clinical management of patients with breast cancer.

Progesterone induction of mammary carcinomas in BALB/c female mice. Correlation between progestin dependence and morphology.

We have demonstrated that medroxyprogesterone acetate (MPA), when administered in high doses, induces mammary carcinomas in virgin female BALB/c mice. Since one of the possible explanations for this effect was its progestagenic effects, we decided to investigate whether progesterone (Pg) alone could also induce mammary adenocarcinomas in our model and if MPA at doses lower than those used to establish the model was also carcinogenic. A total of 136 mice were subdivided into 3 groups: Group 1, 44 mice were implanted s.c. with 40 mg Pg silastic pellets at the beginning of the experiment, and 6 months later with a 20 mg Pg pellet; Group 2, 45 mice were similarly treated with MPA pellets; Group 3, 47 mice were inoculated s.c. with 40 mg MPA every three months. At the end of 20 months, 9 animals had developed mammary tumors in Group 1, 18 in Group 2 and 34 in Group 3 (actuarial incidence = 28%, 58%, and 98%, respectively); tumor latency was similar in all groups: 46.2 +/- 13.1, 51.3 +/- 9.9, and 50.1 +/- 2.1 weeks, respectively. Seven (Group 1), 14 (Group 2), and 25 (Group 3) tumors were transplanted into syngeneic mice to determine progestin dependence. All tumors, except one from Group 1, were histologically characterized. In Group 1 (Pg 60 mg), 4 tumors (67%) were infiltrating lobular carcinomas and 2 were ductal carcinomas (33%). In Group 2 (MPA 60 mg), 2 tumors (14%) were lobular and 12 were ductal adenocarcinomas (86%) (Group 1 vs Group 2: p < 0.05), whereas in Group 3 (MPA 160 mg), 8 were lobular carcinomas (32%) and 17 were ductal carcinomas (68%). In syngeneic passages all lobular tumors behaved as progestin independent (PI) and ductal tumors as progestin dependent (PD). All ductal tumors, except one, expressed estrogen receptors (ER) and progesterone receptors (PR), whereas receptor expression was variable in lobular carcinomas. It can be concluded that Pg induces mostly lobular, PI mammary tumors in BALB/c female mice. The fact that most MPA-induced tumors are ductal and PD suggests that the two hormones use different carcinogenic pathways.