Subsequent Development Of Metastases Research Articles

Non-canonical EphA2 activation underpins PTEN-mediated metastatic migration and poor clinical outcome in prostate cancer

BackgroundThe key process of mesenchymal to amoeboid transition (MAT), which enables prostate cancer (PCa) transendothelial migration and subsequent development of metastases in red bone marrow stroma, is driven by phosphorylation of EphA2S897 by pAkt, which is induced by the omega-6 polyunsaturated fatty acid arachidonic acid. Here we investigate the influence of EphA2 signalling in PCa progression and long-term survival.MethodsThe mechanisms underpinning metastatic biopotential of altered EphA2 signalling in relation to PTEN status were assessed in vitro using canonical (EphA2D739N) and non-canonical (EphA2S897G) PC3-M mutants, interrogation of publicly available PTEN-stratified databases and clinical validation using a PCa TMA (n = 177) with long-term follow-up data. Spatial heterogeneity of EphA2 was assessed using a radical prostatectomy cohort (n = 67).ResultsNon-canonical EphA2 signalling via pEphA2S897 is required for PCa transendothelial invasion of bone marrow endothelium. High expression of EphA2 or pEphA2S897 in a PTENlow background is associated with poor overall survival. Expression of EphA2, pEphA2S897 and the associated MAT marker pMLC2 are spatially regulated with the highest levels found within lesion areas within 500 µm of the prostate margin.ConclusionEphA2 MAT-related signalling confers transendothelial invasion. This is associated with a substantially worse prognosis in PTEN-deficient PCa.

Role of Multidetector CT Imaging in the Risk Stratification of Gastrointestinal Stromal Tumors (GISTs)–A Retrospective Analysis

Abstract Background Gastrointestinal stromal tumors (GISTs) are the most common gastrointestinal mesenchymal neoplasms which can arise from any part of the gastrointestinal tract (GIT) or an extraintestinal location. Size and the organ of origin are the major imaging inputs expected from the radiologist. However, it is worthwhile to find out which imaging characteristics on MDCT correlate with risk stratification. This knowledge would help the clinician in treatment planning and prognostication. The aim of this retrospective study is to evaluate the various MDCT imaging characteristics of GISTs and find out which parameters have significant association with risk and subsequent development of metastasis on follow-up whenever it was possible. Materials and Methods This is a retrospective study conducted on 45 histopathologically proven cases of GIST from two institutions by searching from the digital archives. The following imaging parameters were analyzed: maximum size in any plane, organ of origin, shape (round, ovoid or irregular), margin (well-defined or ill-defined), surface (smooth or lobulated), percentage of necrosis, growth pattern, enhancement characteristics–both intensity (mild, moderate or significant) and pattern (homogenous vs. heterogenous), calcification, infiltration into adjacent organs, and presence of metastasis at presentation or on follow-up. Results CT morphological parameters of significance in risk stratification as per our study include tumor necrosis, predominant cystic change, irregular and lobulated shape/surface characteristics, and adjacent organ infiltration.The parameters which were associated with development of metastasis were size > 5 cm, necrosis > 30%, and the presence of adjacent organ infiltration. Conclusion The radiologist has an important role in ascertaining the size of tumor as well as the organ of origin accurately to guide the clinician in risk calculation and subsequent prognostication. In addition, certain CT characteristics mentioned above, namely, tumor size, significant necrosis/cystic changes, irregular/lobulated contour, and invasion of adjacent organs, help in risk stratification and in predicting metastasis/poor prognosis.

Continuous endoglin (CD105) overexpression disrupts angiogenesis and facilitates tumor cell metastasis

Endoglin (CD105) is an auxiliary receptor for members of the TFG-β superfamily. Whereas it has been demonstrated that the deficiency of endoglin leads to minor and defective angiogenesis, little is known about the effect of its increased expression, characteristic of several types of cancer. Angiogenesis is essential for tumor growth, so high levels of proangiogenic molecules, such as endoglin, are supposed to be related to greater tumor growth leading to a poor cancer prognosis. However, we demonstrate here that endoglin overexpression do not stimulate sprouting or vascularization in several in vitro and in vivo models. Instead, steady endoglin overexpression keep endothelial cells in an active phenotype that results in an impairment of the correct stabilization of the endothelium and the recruitment of mural cells. In a context of continuous enhanced angiogenesis, such as in tumors, endoglin overexpression gives rise to altered vessels with an incomplete mural coverage that permit the extravasation of blood. Moreover, these alterations allow the intravasation of tumor cells, the subsequent development of metastases and, thus, a worse cancer prognosis.

Nanomolar concentration of blood-soluble drag-reducing polymer inhibits experimental metastasis of human breast cancer cells.

Metastasis is the leading cause of cancer mortality. Extravasation of cancer cells is a critical step of metastasis. We report a novel proof-of-concept study that investigated whether non-toxic blood-soluble chemical agents capable of rheological modification of the near-vessel-wall blood flow can reduce extravasation of tumor cells and subsequent development of metastasis. Using an experimental metastasis model, we demonstrated that systemic administration of nanomolar concentrations of so-called drag-reducing polymer dramatically impeded extravasation and development of pulmonary metastasis of breast cancer cells in mice. This is the first proof-of-principle study to directly demonstrate physical/rheological, as opposed to chemical, way to prevent cancer cells from extravasation and developing metastasis and, thus, it opens the possibility of a new direction of adjuvant interventional approach in cancer.

Abstract 4275: Exploring the metastatic potential of exosomal NM23 signaling using a triple negative breast cancer model in mice

Abstract Decades of research has established the idea that cancer cells of the primary tumor prime the metastatic microenvironment prior to establishing metastases at distant sites in the body. While this process is thought to facilitate neocolonization of dislodged primary tumor cells, the mechanisms by which cancer cells communicate to their remote targets remain largely unknown. Our lab has previously shown that triple negative breast cancer cells release exosomes carrying NM23, a nucleoside diphosphate kinase (eNDPK) implicated in promoting angiogenesis and pro-metastatic events extracellularly. To further elucidate the role of eNDPK in breast cancer signaling, we are developing a novel murine model that will allow us to examine its effects in vivo, as well as establish a timeline for the occurrence of metastases. To establish our metastasis model in mice, we first showed that injecting human MDA-MB-231 cells into the mammary fat pad results in the formation of a primary tumor and subsequent development of metastases in the lung. Treating mice with inhibitors of NDPK activity reduces the size of the primary tumor and prevents secondary tumor formation in the lung. To mimic the pulmonary tumor microenvironment, we have isolated endothelial cells from the lungs of 6-8 day old mouse pups using magnetic beads conjugated to the endothelial cell markers CD54, CD102, or CD106. Purity of endothelial cells was confirmed by immunofluorescence staining. In preparation for in vivo experiments that demonstrate eNDPK targeting to lung endothelial cells, we have 3D printed biocompatible polylactic acid (PLA) tubular inserts that are capable of supporting lung endothelial cells encompassed in a gelatinous cell scaffold. Inserts containing lung endothelial cells are surgically introduced into the subcutaneous tissue of adult SCID mice. Human MDA-MB-231 cells expressing GFP-tagged tetraspanin are then injected orthotopically into the mammary fat pads of mice. After a period of 7-21 days, the inserts are excised and analyzed for the presence of GFP-labeled exosomes containing eNDPK and reorganization of the insert microenvironment. Using this model for metastasis, we will be able to confirm the involvement of eNDPK in cell-cell communication between primary tumor cells and their targets in vivo. The significance of uncovering the mechanism(s) by which cancer cells metastasize is emphasized by the fact that recurrence of cancer at distant sites is associated with the most negative outcome in women diagnosed with breast cancer. Implication of eNDPK signaling in metastasis will lead to future research in developing novel small molecule inhibitors. Further, eNDPK can be used as a biomarker for the beginning stages of metastatic breast cancer, replacing current unreliable early detection methods and improving long-term survival outcomes. Citation Format: Suzann Duan, Senny Nordmeier, Iain L.O. Buxton. Exploring the metastatic potential of exosomal NM23 signaling using a triple negative breast cancer model in mice. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4275.

A75: Modern concepts of cellular and molecular mechanisms of breast cancer and their prognostic significance

Breast cancer is one of the most common cancers in women. The incidence of breast cancer in 2014 in the Republic of Mordovia was 69.9 per 100,000 female populations. Breast cancer occurs when excessive expression of oncoproteins switches in the case of transformation of proto-oncogene in PRADI. In primary breast tumors, mutations and the expression of the three oncogenes Her2/neu, C-mys, Int-2, as well as in supressonyh genes – the p53 gene and the retinoblastoma gene RB are the most common. Several studies found that oncogene – C-mys was expressed in 16.8% of primary breast cancer cases and in 35% cases with subsequent development of metastases. Proteins that stimulate the phosphorylation of mitogen-activated protein kinasescan activateunder the influence of growth factors. Development of breast cancer is regulated by a complex interaction of many hormones and growth factors. Currently, one of the leading theories of developing breast cancer is the increased hormonal stimulation of proliferative processes in the development of neoplasia. One of the manifestations of hormonal imbalance in tumor during a regular decrease in blood competitive inhibitor of the biological effects of estrogen – progesterone, which is in correlation with the stage of the spread of neoplasia. The role of the overproduction of estrogen in the pathogenesis of breast cancer is confirmed by the fact that in women who underwent oophorectomy before the age of 38 years, the risk of breast cancer development is 1.5 times less than in those who did not have such operation. Excessive accumulation of lipid peroxidation products in the area of neoplasia activates mechanisms violation of intercellular interaction that caused the destruction of the lipid components of membranes. The study group included 112 patients treated at the State Institution of Health of the Republic of Mordovia “National Oncology Center”. To identify the nature of tumors, all patients underwent immunohistochemical analysis. Androgen receptors were found in 48% of breast cancer cases, the expression level of androgen receptor in the tumor was much lower than the expression level of estrogen and progesterone receptors. Low levels of progesterone receptor expression in breast cancer cells were combined with high levels of expression of Ki-67 antigen, HER-2 oncoprotein in tumor cells. Patients with HER-2 (3+) and (2+) had more frequent multiple metastases in lymph nodes compared to patients with HER-2 (0) and (1+) phenotypes. Maximum expression of HER-2 oncoprotein in tumor cells indicated high metastatic potential and poor prognosis. It may be concluded that the cellular and molecular mechanisms of breast cancer are complex. Therefore, carcinogenesis has a “multistep” nature and at least two or more mutations in the cells of the same clone – parent and child are required to generate malignant tumors. Thus, the development of oncogenic transformation does not necessarily mean the process of tumor formation.

Complete Regression of Melanocytic Nevi: Correlation Between Clinical, Dermoscopic, and Histopathologic Findings in 13 Patients

BackgroundThe complete regression of melanocytic tumors, confirmed by histology, has rarely been reported in the literature. It is very difficult to determine the malignant or benign nature of a regressed tumor, and on occasions, the only indication of malignancy is the subsequent development of metastasis. Material and methodsWe performed a descriptive study of melanocytic nevi that had undergone complete, histologically confirmed regression prior to excision in the dermatology department of our hospital over a period of 3 years. We included only lesions in which dermoscopy performed prior to regression showed features that suggested benignity. We assessed various clinical, dermoscopic, histologic, and immunohistochemical features. ResultsThe mean time to complete regression was 6.4 months. The main dermoscopic patterns observed were reticular and mixed reticular/globular. Unlike what is generally seen in melanomas, the main histologic finding was the presence of fine or lamellar fibrosis. In all cases, there was a predominance of CD8+ T cells. ConclusionsThe clinical, dermoscopic, and histologic features of the melanocytic nevi studied suggest the existence of a highly characteristic form of tumor regression characterized by very rapid regression and the involvement of a cytotoxic mechanism.

Regresión completa de nevos melanocíticos: correlación clínica, dermatoscópica e histológica de una serie de 13 casos

BackgroundThe complete regression of melanocytic tumors, confirmed by histology, has rarely been reported in the literature. It is very difficult to determine the malignant or benign nature of a regressed tumor, and on occasions, the only indication of malignancy is the subsequent development of metastasis. Material and methodsWe performed a descriptive study of melanocytic nevi that had undergone complete, histologically confirmed regression prior to excision in the dermatology department of our hospital over a period of 3 years. We included only lesions in which dermoscopy performed prior to regression showed features that suggested benignity. We assessed various clinical, dermoscopic, histologic, and immunohistochemical features. ResultsThe mean time to complete regression was 6.4 months. The main dermoscopic patterns observed were reticular and mixed reticular/globular. Unlike what is generally seen in melanomas, the main histologic finding was the presence of fine or lamellar fibrosis. In all cases, there was a predominance of CD8+ T cells. ConclusionsThe clinical, dermoscopic, and histologic features of the melanocytic nevi studied suggest the existence of a highly characteristic form of tumor regression characterized by very rapid regression and the involvement of a cytotoxic mechanism.

Dilation of Malignant Strictures in Endoscopic Ultrasound Staging of Esophageal Cancer and Metastatic Spread of Disease

Background. Dilation of malignant strictures in endoscopic ultrasound (EUS) staging of esophageal cancer is safe, but no data exists regarding the subsequent development of metastases. Aim. Compare the rates of metastases in esophageal cancer patients undergoing EUS staging who require esophageal dilation in order to pass the echoendoscope versus those who do not. Methods. We reviewed consecutive patients referred for EUS staging of esophageal cancer. We evaluated whether dilation was necessary in order to pass the echoendoscope, and for the subsequent development of metastases after EUS at various time intervals. Results. Among all patients with similar stage (locally advanced disease, defined as T3, N0, M0 or T1-3, N1, M0), there was no difference between the dilated and nondilated groups in the rates of metastases at 3 months (14% versus 10%), P = 1.0, 6 months (28% versus 20%), P = 0.69, 12 months (43% versus 40%), P = 1.0, or ever during a mean followup of 15 months (71% versus 55%), P = 0.48. Conclusions. Dilation of malignant strictures for EUS staging of esophageal cancer does not appear to lead to higher rates of distant metastases.

Elevated osteonectin/SPARC expression in primary prostate cancer predicts metastatic progression

The majority of prostate cancers (CaP) are detected in early stages with uncertain prognosis. Therefore, an intensive effort is underway to define early predictive markers of CaP with aggressive progression characteristics. In order to define such prognostic markers, we performed comparative analyses of transcriptomes of well- and poorly differentiated (PD) tumor cells from primary tumors of patients (N=40) with 78 months of mean follow-up after radical prostatectomy. Validation experiments were carried out at transcript level by quantitative real-time reverse transcriptase-PCR (RT-PCR) (N=110) and at protein level by immunohistochemistry (N=53) in primary tumors from an independent patient cohort. Association of a biochemical network of 12 genes with SPARC gene as a central node was highlighted with PD phenotype. Of note, there was remarkable enrichment of NKXH_NKXH_HOX composite regulatory elements in the promoter of the genes in this network suggesting a biological significance of this gene-expression regulatory mechanism in CaP progression. Further, quantitative expression analyses of SPARC mRNA in primary prostate tumor cells of 110 patients validated the association of SPARC expression with poor differentiation and higher Gleason score. Most significantly, higher SPARC protein expression at the time of prostatectomy was associated with the subsequent development of metastasis (P=0.0006, AUC=0.803). In summary, we propose that evaluation of SPARC in primary CaP has potential as a prognostic marker of metastatic progression.

Mammalian Target of Rapamycin (mTOR) Regulates Cellular Proliferation and Tumor Growth in Urothelial Carcinoma

Mammalian target of rapamycin (mTOR) signaling has been associated with aggressive tumor growth in many cancer models, although its role in urothelial carcinoma (UCC) has not been extensively explored. Expression of phosphorylated mTOR (P-mTOR) and a downstream target, ribosomal S6 protein (P-S6), was identified in 74% (90/121) and 55% (66/121) of muscle-invasive UCCs, respectively. P-mTOR intensity and %positive cells were associated with reduced disease-specific survival (P = 0.04, P = 0.08, respectively). Moreover, P-mTOR intensity corresponded to increased pathological stage (P < 0.01), and mTOR activity was associated with cell migration in vitro. In addition, mTOR inhibition via rapamycin administration reduced cell proliferation in UCC cell lines RT4, T24, J82, and UMUC3 in a dose-dependent manner to 6% of control levels and was significant at 1 nmol/L in J82, T24, and RT4 cells (P < 0.01, P < 0.01, P = 0.03, respectively) and at 10 nmol/L in UMUC3 cells (P = 0.03). Reduced proliferation corresponded with reduced P-S6 levels by Western blot, and effects were ablated by pretreatment of cells with mTOR-specific siRNA. No effects of rapamycin on apoptosis were identified by TUNEL labeling or PARP cleavage. Administration of rapamycin to T24-xenografted mice resulted in a 55% reduction in tumor volume (P = 0.03) and a 40% reduction in proliferation (P < 0.01) compared with vehicle-injected mice. These findings indicate that mTOR pathway activation frequently occurs in UCC and that mTOR inhibition may be a potential means to reduce UCC growth.

Deep proteome profiling of sera from never-smoked lung cancer patients

Previous studies on the serum proteome are hampered by the huge dynamic range of concentration of different protein species. The use of Equalizer Beads coupled with a combinatorial library of ligands has been shown to allow access to many low-abundance proteins or polypeptides undetectable by classical analytical methods. This study focused on never-smoked lung cancer, which is considered to be more homogeneous and distinct from smoking-related cases both clinically and biologically. Serum samples obtained from 42 never-smoked lung cancer patients (28 patients with active untreated disease and 14 patients with tumor resected) were compared with those from 30 normal control subjects using the pioneering Equalizer Beads technology followed by subsequent analysis by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Eighty-five biomarkers were significantly different between lung cancer and normal control. The application of classification algorithms based on significant biomarkers achieved good accuracy of 91.7%, 80% and 87.5% in class-prediction with respect to presence or absence of disease, subsequent development of metastasis and length of survival (longer or shorter than median) respectively. Support vector machine (SVM) performed best overall. We have proved the feasibility and convenience of using the Equalizer Beads technology to study the deep proteome of the sera of lung cancer patients in a rapid and high-throughput fashion, and which enables detection of low abundance polypeptides/proteins biomarkers. Coupling with classification algorithms, the technologies will be clinically useful for diagnosis and prediction of prognosis in lung cancer.

Density of DC-LAMP+ mature dendritic cells in combination with activated T lymphocytes infiltrating primary cutaneous melanoma is a strong independent prognostic factor

As the most potent antigen presenting cells, dendritic cells (DCs) play key roles in the immune response against tumors. Their density in the tumor tissue has been associated with prognosis in patients with various cancers. However, few studies have been aimed at the presence and maturation state of DCs in cutaneous melanoma, with regard to their potential clinical correlates. In this study, the density of DCs expressing CD1a and the maturation marker DC-LAMP was determined by immunohistochemistry in primary tumor samples from 82 patients with cutaneous malignant melanoma. Intratumoral and peritumoral cell densities were analyzed in relation to tumor thickness and the subsequent development of metastases, as well as to patients' survival. CD1a(+) DCs were found both infiltrating melanoma cell nests and in the surrounding stroma, while DC-LAMP(+) mature DCs were generally confined to the peritumoral areas, associated with lymphocytic infiltrates. DC density values significantly correlated with the number of activated (CD25(+) or OX40(+)) T lymphocytes (p < 0.001). The degree of infiltration by CD1a(+) and DC-LAMP(+) DCs showed strong inverse correlation with the thickness of melanomas (p < 0.001). High peritumoral density of mature DCs was associated with significantly longer survival (p = 0.0195), while density of CD1a(+) cells had a prognostic impact of borderline significance (p = 0.0610). Moreover, combination of high peritumoral CD1a(+) or DC-LAMP(+) cell density with high number of CD25(+) or OX40(+) lymphocytes identified patient subgroups with more favorable survival compared to other subgroups. A multivariate survival analysis involving DC and activated T-cell densities alone and in combinations, as well as traditional prognostic factors, identified high DC-LAMP(+) cell/high OX40(+) cell density and Breslow index as independent predictors of good prognosis. These results suggest that the presence of CD1a(+) DCs primarily depends on the thickness of melanomas, without direct relationship with the patients' survival. On the other hand, the density of mature DCs, especially in association with that of activated T cells, proved of prognostic importance, suggesting that these parameters could be considered as signs of a functional immune response associated with better outcome of the disease.

Small Hypoattenuating Hepatic Lesions at Contrast-enhanced CT: Prognostic Importance in Patients with Breast Cancer

To retrospectively determine the prognostic importance of small hypoattenuating hepatic lesions at contrast material-enhanced computed tomography (CT) in patients with breast cancer. This retrospective study was approved by the committee on human research. Written informed consent was not required. The authors retrospectively identified 153 patients with breast cancer who underwent serial abdominal CT and who did not have definite liver metastases present at initial CT. The mean age was 56 years (age range, 27-93 years). Two readers independently recorded the presence, size, and number of small (15 mm or less in diameter) hypoattenuating hepatic lesions at initial CT. Another reader independently recorded the presence or absence of definite hepatic metastases at final CT. The association between the presence, size, and number of small hypoattenuating hepatic lesions at initial CT and the subsequent development of metastases was analyzed by using Kaplan-Meier analysis. One or more small hypoattenuating hepatic lesions were seen at initial CT in 54 of 153 patients (35%). After a median follow-up of 584 days (range, 16-1827 days), definite hepatic metastases developed in 43 of 153 patients (28%), including 15 of 54 patients (28%) with hypoattenuating lesions at initial CT and 28 of 99 patients (28%) without hypoattenuating lesions at initial CT. Findings from the Kaplan-Meier analysis showed no association between the presence (P = .56), size (P = .55), or number (P = .30) of small hypoattenuating hepatic lesions at initial CT and the subsequent development of hepatic metastases. In patients with breast cancer who do not have definite hepatic metastases at initial examination, there is no evidence that small hypoattenuating hepatic lesions seen at initial CT contribute to an increased risk of subsequently developing hepatic metastases.

Conjugated Linoleic Acid Content in Breast Adipose Tissue of Breast Cancer Patients and the Risk of Metastasis

The association between the level of conjugated linoleic acid (CLA) in breast adipose tissue at the time of diagnosis and the subsequent development of metastasis was examined in a cohort of 209 patients presenting with an initially localized breast cancer. CLA level in breast adipose tissue was used as a qualitative biomarker of its past dietary intake. Biopsies of adipose tissue were obtained at the time of initial surgery. A CLA-enriched fraction was prepared by high performance liquid chromatography and CLA measured as a percentage of total fatty acids, using capillary gas chromatography. Mean CLA level was low (0.44% of total fatty acids) and the range between patients was narrow (0.19-0.85). With a median follow-up time of 7.5 yr, 45 patients developed metastases. A Cox proportional hazard regression model was used to identify prognostic factors. We did not find any significant association between CLA level in adipose fat and either the prognostic factor (tumor size, nodal status, histoprognostic grade, mitotic index, and estrogen or progesterone receptors) or the risk of metastasis or death. We concluded that CLA are unlikely to be involved in survivorship. However, the hypothesis that a higher intake of CLA might have a protective effect on the risk of metastasis cannot be ruled out from these data, since the level of CLA in breast cancer patients' adipose tissue is likely to be too low and the range of CLA distribution too narrow for any protection to be detectable.

Simultaneous measurement of serum 5-S-cysteinyldopa, circulating intercellular adhesion molecule-1 and soluble interleukin-2 receptor levels in Japanese patients with malignant melanoma.

Serum 5-S-cysteinyl dopa (5-S-CD), circulating intercellular adhesion molecule-1 (cICAM-1) and soluble interleukin-2 receptor (sIL-2R) have each been reported as useful markers for melanoma progression. To assess the clinical relevance of these three markers, we simultaneously assayed their serum levels in 30 Japanese melanoma patients. Pre-surgical serum levels of 5-S-CD, cICAM-1 and sIL-2R were elevated in six, 13 and five patients respectively. These abnormal values returned to normal after curative surgery in most of the patients, suggesting a direct relationship to the presence of the primary tumour. Pre-surgical values of these three markers, either individually or in combination, did not predict the development of subsequent metastases. The sequential measurements of the three markers in eight patients who had relapse after surgery showed that serum 5-S-CD is the most useful marker for disease progression, although it is dependent on the melanin-producing ability of individual recurrent tumours. sIL-2R seemed to reflect in vivo activation of the host immune system and was a good indicator for predicting occult metastasis in selected cases. Circulating ICAM-1 levels were less relevant to the clinical disease course in our cases, although they tended to increase strikingly after liver metastasis. Our results in this limited number of cases show that the significance of the three markers varied with the individual and suggest that the simultaneous measurement of these markers may facilitate the early detection of metastases and proper therapeutic intervention.

Prognosis of salivary adenocarcinomas

52 salivary adenocarcinomas of the years 1965-1980 from the files of the Salivary Gland Registry, Institute of Pathology, University of Hamburg, were evaluated retrospectively with regard to clinical follow up and cytochemically assessed nuclear DNA content. The age distribution showed a peak from the 6th to 8th decade (range 3 to 87 years). The m:f ratio was 1:1.36, the mean age was 59.3 years. Over 80% of the tumours were located in the major salivary glands. The clinical course was characterized by metastases present at initial diagnosis (16 cases), subsequent development of metastases (9 cases), local recurrence (15 cases) or death from tumour (10 cases) and was related to differentiation, grade 3 tumours showing the worse clinical courses. In 37 cases, nuclear DNA content was determined by a single scanning cytophotometry device. 28 cases were diploid, 9 were atypical. The clinical course was significantly related to the histogram type, atypical tumours showing a dismal prognosis.

Lack of correlation between metastasis of human rectal carcinoma and the absence of stromal fibronectin.

In a retrospective study we have used an immunoperoxidase procedure to localize the glycoprotein fibronectin in human rectal carcinomas, concentrating on tumour invading thick-walled extramural veins. Fibronectin was present in 29 out of 38 cases, in connective tissue stroma, and was not in direct association with the tumour cells, except in areas of necrosis. We found no correlation between the presence or absence of stromal fibronectin and (1) the degree of cellular differentiation within the tumour, (2) tumour progression (Dukes' classification) (3) the subsequent development of metastases and (4) patient longevity. OUr results do not support the conclusions from in vitro studies (Smith et al., 1979) that the metastatic potential of carcinomas may be partly determined by the ability of tumour cells to synthesize pericellular fibronectin.

Metastatic squamous cell carcinoma of the lip.

Squamous cell carcinoma of the lip is a common lesion that is not always acknowledged as a potentially lethal disease. We reviewed the cases of 27 patients with metastatic squamous cell carcinoma of the lip. Our data confirm that patients in an older age group are at greater risk for development of metastases than the general population of patients with squamous cell carcinoma of the lip. We also found a universally grave prognosis for patients who exhibit fixation of tumor to the mandible or erosion of the mandible. We were unable to confirm a greater risk for development of metastases among more advanced (T3 and T4) lesions. A number of our patients had T1 and T2 primary lesions. We also could not establish a relationship between a patient's delay before seeking medical assistance and subsequent development of metastases.

Prognosis in inoperable Stage III carcinoma of the breast

One hundred and eighty-four patients with inoperable Stage III breast cancer presenting to the Guy's Hospital Breast Unit between 1961–1973 were treated initially by radiotherapy alone. The response rate was 60% but duration of response and survival were short. Seventy-two per cent of patients did not have prolonged control of local disease and 63% developed distant metastases. The association between certain prognostic variables and response to radiotherapy, subsequent development of metastases and survival was analysed statistically. The duration of response to radiotherapy showed no significant associations with any prognostic variable. Subsequent distant metastases occurred less often in patients responding to radiotherapy, having a subsequent mastectomy or if the duration of symptoms before presentation was long, but more frequently if the primary tumours were diffuse. Survival was shorter in patients who were early postmenopausal, had a short duration of symptoms or had diffusc primary tumours. Improved survival was associated with a good response to radiotherapy and, unexpectedly, with deep fixation of the primary tumour. Prognosis was not significantly associated with size of primary tumour or involvement of skin or lymph nodes. The effectiveness of combining variables in predicting prognosis is described. A further group of 30 patients, unsuitable for radiotherapy were treated primarily by additive endocrine therapy and had a median survival of 14 months. The clinical course of Stage III breast cancer is variable, there being two extremes: a slowly-growing, non-metastasising form and a more common, rapidly-growing, metastasising form. For prognosis to be improved, systemic therapy as part of the primary management of this disease may be necessary. Prognostic variables should be considered in the design and assessment of future clinical trials.