Thirty-Day Mortality Following Systemic Anticancer Therapy: Evaluating Risk Factors Without Selection Bias in a Real-World, Population-Based Cohort From 2009 to 2019.

AIMS: Risk factors for systemic anticancer therapies (SACTs) administered close to death derived from existing quality indicators are not directly applicable in the clinic, because they condition on future events, which leads to selection bias. This study aimed to adapt a previously suggested indicator for its use in a clinical context and to evaluate it in a real-world, population-based cohort of cancer patients. MATERIALS AND METHODS: An improved version of the '30-day mortality after SACT' indicator suggested by Wallington et al. (Lancet Oncol 2016; 17:1203-16) was defined. All SACTs (n = 16 622) for all patients (n = 10 213) treated for common malignancies between 2009 and 2019 in the North Denmark Region were included. The results for the improved and Wallington's indicators were calculated and compared. RESULTS: Overall, the association between clinical variables and 30-day mortality following SACT was similar for both indicators, except for the 75+ years age group. However, Wallington's indicator showed varying absolute risk when comparing values for quarterly and yearly observation intervals. The improved and Wallington's indicators showed large differences between curative (1.0% and 1.1%, respectively) and palliative SACTs (9.1% and 11.7%, respectively). For palliative SACTs, different types of malignancy presented with large variations for the improved indicator, ranging from above 10% for gastroesophageal, pancreatic and lung cancers to below 4% for prostate cancers. The value of the improved indicator was significantly lower in the last years of the study period compared with the 2009-2011 period. The type of malignancy was also associated with significant differences. CONCLUSIONS: We defined an indicator adapted to the clinical context evaluating 30-day mortality following SACT. This indicator can be used to identify risk factors to help with clinical decision-making. A significant downward trend was observed in the 30-day mortality following palliative SACT over an 11-year period.

Adjuvant platinum-based chemotherapy in non-small cell lung cancer: The role of relative dose-intensity and treatment delay.

BACKGROUND: The study investigated the association of the relative dose-intensity (RDI) of cisplatin and timing of adjuvant platinum-based chemotherapy (APC) with survival for stage I-III non-small cell lung cancer (NSCLC) patients. MATERIAL AND METHODS: Real-life data of patients treated with APC (four cycles of cisplatin and vinorelbine) between 2007 and 2014 was included to analyse the association between disease-free survival (DFS) and overall survival (OS) with RDI (ratio of received to planned dose-intensity). High RDI was defined as cisplatin RDI of > 75% and low RDI ≤ 75%. RESULTS: Out of 198 patients, 166 were eligible. Low RDI was administered to 72 (43%) patients. In multivariate analysis, those patients had a significantly higher risk of recurrence (HR: 1.87, 95%CI 1.13-3.09, p = 0.01) and death (HR: 1.91, 95%CI 1.32-3.23, p = 0.01) versus patients in the high RDI group. The risk of death was significantly higher in patients with PS 1 treated with low versus high RDI (HR: 2.72, 95%CI: 1.22-6.09, p = 0.014). The risk of recurrence was higher for patients with squamous cell carcinoma of low versus high RDI (HR: 3.82, 95%CI: 1.01-14.4, p = 0.048). No impact of delayed APC beyond six weeks from surgery on neither DFS (HR: 0.78, 95%CI: 0.46-1.33, p = 0.36) nor OS (HR 0.67, 95%CI: 0.40-1.15, p = 0.15) was observed. CONCLUSION: Low cisplatin RDI ≤ 75% of APC, but not extended time from surgery to APC onset > six weeks, was associated with significantly shorter survival in NSCLC patients.

Fractional exhaled nitric oxide as a potential biomarker for radiation pneumonitis in patients with non-small cell lung cancer: A pilot study.

INTRODUCTION: The aim of the study was to investigate repetitive fractional exhaled nitric oxide (FeNO) measurements during high-dose radiation therapy (HDRT) and to evaluate the use of FeNO to predict symptomatic radiation pneumonitis (RP) in patients being treated for non-small cell lung cancer (NSCLC). MATERIALS AND METHODS: A total of 50 patients with NSCLC referred for HDRT were enrolled. FeNO was measured at baseline, weekly during HDRT, one month- and every third month after HDRT for a one-year follow-up period. The mean FeNO(visit 0-6) was calculated using the arithmetic mean of the baseline and weekly measurements during HDRT. Patients with grade ≥ 2 of RP according to the Common Terminology Criteria for Adverse Events (CTCAE) were considered symptomatic. RESULTS: A total of 42 patients completed HDRT and weekly FeNO measurements. Grade ≥ 2 of RP was diagnosed in 24 (57%) patients. The mean FeNO(visit 0-6) ±â€¯standard deviation in patients with and without RP was 15.0 ±â€¯7.1 ppb (95%CI: 12.0-18.0) and 10.3 ±â€¯3.4 ppb (95%CI: 8.6-11.9) respectively with significant differences between the groups (p = 0.0169, 95%CI: 2.3-2.6). The leave-one-out cross-validated cut-off value of the mean FeNO(visit 0-6) ≥ 14.8 ppb was predictive of grade ≥ 2 RP with a specificity of 71% and a positive predictive value of 78%. CONCLUSIONS: The mean FeNO(visit 0-6) in patients with symptomatic RP after HDRT for NSCLC was significantly higher than in patients without RP and may serve as a potential biomarker for RP.

Longitudinal alterations in nutrient intake and food pattern in patients with non-small cell lung cancer during anti-neoplastic treatment: a cohort study.

BACKGROUND: Unintentional weight loss is frequently observed in cancer patients. Nutritional therapy is essential, and dietary counselling is the first step. The present study aimed to explore the nutrient intake and food patterns in weight-stable and weight-losing patients with non-small cell lung cancer (NSCLC) during anti-neoplastic treatment. METHODS: Patients with NSCLC (n = 62) were observed during first-line systemic anti-neoplastic treatment. Body weight and dietary intake were assessed on the first and second cycle, and after completing three cycles of treatment. Longitudinal changes were analysed in three groups: weight stable, weight losers and mixed weight. RESULTS: Nutrient intake did not change during treatment in weight stable, although weight losers significantly increased the relative protein intake. Weight stable maintained the food pattern during treatment apart from a decreased consumption of oral nutritional support (ONS). At baseline, weight losers were characterised by pretreatment weight loss, high consumption of ONS, as well as low consumption of grains and animal products. During treatment, weight losers increased the consumption of protein, fatty foods and ONS but decreased the consumption of sweets and alcohol. CONCLUSIONS: Large heterogeneity in nutrient and food intake was observed in NSCLC patients during anti-neoplastic treatment. Weight losers and weight stable had a similar nutrient intake although protein intake increased in weight losers. Grains and animal products were lower and ONS higher in weight losers compared to weight stable during treatment. Weight losers further increased the consumption of ONS and fatty foods, while the consumption of sweets and alcohol decreased during treatment.

Local recurrence rate in a national Danish patient cohort after curative treatment for rectal cancer.

AIM: Several trials have shown that preoperative (chemo)radiotherapy (CRT) reduces local recurrence rates (LRRs) in rectal cancer (RC). The use of CRT varies greatly between countries. It is unknown whether the restrictive use of CRT in Denmark results in a higher LRR relative to other countries. The aim was to evaluate the LRR in a national Danish consecutive cohort of patients with RC. METHODS: All data from patients with RC in Denmark in 2009-2010 who were operated on with curative intent were retrieved from the Danish Colorectal Cancer Group database. Patients with metastases at the time of diagnosis, patients with synchronous colon cancer, and patients, in whom only local surgical procedures were performed, were excluded. In total, 1633 patients met the inclusion criteria. Clinical follow-up was at least five years with a cut-off date of 31 December 2015. RESULTS: Clinical follow-up was 5.4 years (median) with an interquartile range of 4.5-6.1 years. Of all included patients, 479 (29%) were treated with preoperative long-course CRT. Local recurrence was found in 68 patients, resulting in an LRR of 4.2%, and 182 (11%) patients developed distant metastases. Five-year overall survival was 74% (95% CI: 71.64-75.91). CONCLUSIONS: Five-year follow-up of curatively treated patients with RC in Denmark revealed a low LRR. This figure is identical to those reported in other Nordic countries, despite Denmark's considerably stricter guidelines for CRT. The obtained results justify the currently adopted restrictive use of preoperative CRT in Denmark.

The established and future biomarkers of malignant pleural mesothelioma.

Malignant pleural mesothelioma (MPM) is an asbestos-related cancer with a median survival of 12months. The MPM incidence is 1-6/100,000 and is increasing as a result of historic asbestos exposure in industrialized countries and continued use of asbestos in developing countries. Lack of accurate biomarkers makes diagnosis, prognostication and treatment prediction of MPM challenging. The aim of this review is to identify the front line of MPM biomarkers with current or potential clinical impact. Literature search using the PubMed and PLoS One databases, the related-articles function of PubMed and the reference lists of associated publications until April 26th 2015 revealed a plethora of candidate biomarkers. The current gold standard of MPM diagnosis is a combination of two positive and two negative immunohistochemical markers in the epithelioid and biphasic type, but sarcomatous type do not have specific markers, making diagnosis more difficult. Mesothelin in serum and pleural fluid may serve as adjuvant diagnostic with high specificity but low sensitivity. Circulating proteomic and microRNA signatures, fibulin-3, tumor cell gene-ratio test, transcriptomic, lncRNA, glycopeptides, pleural fluid FISH assay, hyaluronate/N-ERC mesothelin and deformability cytometry may be important future markers. Putative predictive markers for pemetrexed-platinum are tumor TS and TYMS, for vinorelbine the ERCC1, beta-tubuline class III and BRCA1. Mutations of the BAP1 gene are potential markers of MPM susceptibility. In conclusion, the current status of MPM biomarkers is not satisfactory but encouraging as more sensitive and specific non-invasive markers are emerging. However, prospective validation is needed before clinical application.

Neuroendocrine cells in pancreatic duct adenocarcinoma: an immunohistochemical study.

Pancreatic ductal adenocarcinomas can display disseminated neuroendocrine (NE) cells. Controversies exist as to their relative incidence, histogenesis, hormone production, and the prognostic implications of their presence. These issues were elucidated by means of a broad immunohistochemical (IHC) investigation of the resected specimens from 47 patients. Chromogranin A (CgA) was chosen as the major NE marker. In addition, the sensitivity of the conventional IHC procedure was increased by means of the TSA (Tyramide Signal Amplification) technique. In tumours with CgA immunoreactive (IR) cells, detected by the conventional or the TSA methods, these NE cells were further IHC analyzed, using antisera raised against a broad spectrum of neurohormonal peptides, serotonin, and IGF-1. The IHC observations were correlated with clinical and histopathological data, the nuclear IR for the Ki67 antigen (proliferation) of the neoplastic cells, and their IR against the p53 protein. Distinct CgA IR cells were found in 5 out of 47 (11%) tumours when studied by the conventional method, and in 9 out of 47 (19%) when examined by the TSA technique. Corresponding figures, if tumours with only questionable IR against CgA were also included, were 14 (30%) and 23 (50%), respectively. Out of the 9 cases with unequivocal CgA IR, only 3 displayed an IR to an additional hormone or growth factor; this hormone turned out to be somatostatin (only minimal foci). Insulin and glucagon cells also appeared exceptionally. The NE differentiation was found to be unrelated to proliferation, p53 protein expression, and to the survival of the patients. It occurred mainly (7 out of 9) in poorly differentiated adenocarcinomas. Thus, the plain NE immunoprofile of the CgA IR cells, together with the increased IR observed when the TSA technique was used, indicates that the NE cells in these adenocarcinomas are only poorly differentiated. When the CgA IR cells exceptionally become highly differentiated, they can express islet hormones. Using strict structural and IHC criteria, a NE differentiation occurs in less than 20 % of cases; its clinico-pathological significance seems to be non relevant.

Immunoreactivity of gastric ECL and A-like cells in fasted and fed rats and mice.

The oxyntic mucosa of rat and mouse stomach harbors histamine-producing ECL cells and ghrelin-producing A-like cells. The ECL cells are known to be active when the circulating gastrin levels are elevated in response to food intake. The A-like cells are the main source of circulating ghrelin. In response to starvation, the circulating ghrelin is elevated as a hunger signal. The aim of the present work was to study the correlation between the immunoreactivities and cellular activities of the ECL cells and A-like cells. Rats were either fed or fasted for 48 h and mice for 24 h. Immunohistochemical examination with antiserum against chromogranin A-derived fragment pancreastatin revealed both the ECL cells and the A-like cells without a difference between fasted and fed animals. Histamine was limited to the ECL cells with no significant difference between fasted and fed animals. Histidine decarboxylase (HDC) immunoreactivity occurred predominately in the ECL cells of the fed, but not fasted, animals in which the HDC enzymatic activity in the oxyntic mucosa was higher than in fasted animals. Ghrelin immunoreactivity was increased in terms of intensity, but not cell density in fasted animals. Thus, the immunoreactivities of ECL cells and A-like cells might be affected by starvation.

IGF-I in human breast cancer: low differentiation stage is associated with decreased IGF-I content.

OBJECTIVE: Few investigations on the potential role of IGF-I in human breast cancer have used morphological criteria, and the data presented on the localisation of IGF-I are controversial. Moreover, little information exists on a potential correlation between local IGF-I and the grade of malignancy or prognostic factors. Therefore, we investigated the immunohistochemical localisation of IGF-I in specimens of human breast cancer tumours of the ductal type, graded as G1/G2 (well-/moderately differentiated, n=115) and G3 (poorly differentiated, n=28). METHODS: IGF-I immunoreactivity was quantified using a scaling from no (-) to numerous (+++) IGF-I-immunoreactive cells. From 29 of the G1/G2 and 17 of the G3 tumours IGF-I was also measured by RIA. Cytosolic oestrogen receptor (ER) and progesterone receptor (PR) levels, and S-phase fraction were established and related to the number of IGF-I-immunoreactive cells. RESULTS: IGF-I immunoreactivity occurred predominantly in ductal epithelial cells. Of G3 tumours, 57% exhibited IGF-I immunoreactivity as compared with 84% of G1/G2 tumours. Correspondingly, the amount of IGF-I measured by RIA was significantly lower in G3 tumours (6.9+/-0.9 ng/g wet weight) than in G1/G2 tumours (10.5+/-1.1 ng/g wet weight) (P=0.031). G1/G2 tumours exhibited a higher percentage of IGF-I-immunoreactive cells (16% -, 23% +, 41% ++, 20% +++) than G3 tumours (43% -, 37% +, 12% ++, 8% +++). When comparing the - with the +++ G1/G2 tumours, the frequency of IGF-I-immunoreactive cells was related significantly to the ER (P<0.016) and the PR (P<0.008) levels. In G1/G2 and G3 tumours, the ER and PR levels increased with the amount of IGF immunoreactivity while the S-phase fraction increased with decreasing IGF-I content. In 25% of the specimens, IGF-I immunoreactivity occurred in stromal cells, but there was no obvious difference between the different types of tumours. The survival of the G1/G2 tumour patients increased with increasing numbers of IGF-I-immunoreactive cells. CONCLUSIONS: It is concluded that IGF-I is associated with the more-differentiated type of epithelial cells and that increasing dedifferentiation goes along with decreased IGF-I content. Thus, the presence of IGF-I immunoreactivity in breast cancer epithelial cells indicates a lower degree of malignancy than the lack of IGF-I.

Absence of somatostatin receptor expression in vivo is correlated to di- or tetraploid 1p36-deleted neuroblastomas.

BACKGROUND: Poor prognosis in childhood neuroblastoma is associated with deletions of chromosome region 1p36 and di/tetraploid DNA content. PROCEDURE: Forty-six patients with histopathologically proven neuroblastoma were investigated for in vivo expression of somatostatin receptors (SR) by 111In-pentetreotide scintigraphy. All tumors were analyzed for cytometric DNA content and chromosome 1p36 integrity. RESULTS: SR expression was detected in 28 tumors (61%) and correlated with young age, localized clinical stage, and favorable outcome. Fourteen tumors showed deletion at chromosome 1p36, thirteen of which did not show SR expression (P< 0.001). A triploid DNA content was correlated with the presence of SR (23 of 25, P< 0.001). No tumor with deletion of chromosome 1p36 and di/tetra DNA content showed SR expression (chi2 = 29.88, d.o.f. = 2, P < 0.001). CONCLUSIONS: We conclude that SR expression is related to genetic features of prognostic significance. This may be assessed with a minimally invasive scintigraphic method.

Pattern of distribution and prognostic value of angiogenesis in pancreatic duct carcinoma: a semiquantitative immunohistochemical study of 45 patients.

In specimens obtained from resected pancreata, the intratumoral microvessel density (IMD), the proliferation rate of the neoplastic parenchymal cells, and their p53 protein expression were assessed. The sources of errors were great in the measurements of the IMD. This statement can be illustrated by the finding that when the IMD was calculated by manual counting in five areas of intense neovascularization (hot spot regions), using x200 and x400 magnifications, the numbers of microvessels per square millimeter were 65+/-23 and 106+/-8, respectively, which reflects a significant difference. Two patterns of microvessel distribution could be identified: one with hot spots only in the stroma (n = 19) and one in which the hot spots were located in areas of neoplastic parenchyma (including its stroma) (n = 26). The IMD was significantly greater in the latter group. There was no general correlation of neoplastic disease with the IMD. However, when a scoring system was used to assess the angiogenesis, hot spots in areas of neoplastic parenchyma were associated with a greater proliferation rate of the tumor cells, and with a short length of survival of the patients from their neoplastic disease.

Parathyroid hormone-related protein in patients with primary breast cancer and eucalcemia.

Parathyroid hormone-related protein (PTHrP) is a causative factor of humoral hypercalcemia in breast cancer and other malignancies. We studied circulating PTHrP levels with three different immunoassays directed against different parts of the PTHrP molecule in 48 patients with breast cancer and eucalcemia. The methods used were: (a) a RIA with antibodies directed toward the midregion (63-78); (b) an immunofluorometric assay with two antibodies against 1-34 and 38-67; and (c) an immunoradiometric assay with antibodies against 1-40 and 1-72. Although most patients had PTHrP levels indistinguishable from normal when measured by all three methods, four patients had increased serum levels in the IFMA. PTHrP was detected by immunohistochemistry in tumors from nearly all patients. One patient with elevated PTHrP in plasma measured by IFMA showed intense staining of tumor by immunohistochemistry; the tumor was histologically graded as III (severe) and was the largest of all tumors in this patient group. The IFMA can identify increased serum PTHrP in some patients with breast cancer who are not hypercalcemic. This assay may be especially useful in screening patients for this tumor during a relative early phase of the disease.

Prognosticating tools in primary neuroendocrine (Merkel-cell) carcinomas of the skin: histopathological subdivision, DNA cytometry, cell proliferation analyses (Ki-67-immunoreactivity) and NCAM immunohistochemistry. A clinicopathological study in 25 patients.

Histopathologically, 18 of our patients had classical Merkel-cell carcinomas (MCC); seven had neuroendocrine (NE) carcinomas with features different from MCC, here called "aberrant MCC". These patients showed a progressive neoplastic disease with a fatal outcome in four of them. The cytometric DNA distribution pattern of the tumor cell nuclei of all the aberrant MCCs was found to be of the aneuploid type. By contrast, the neoplastic disease of the majority of patients with classical MCC ran a milder course; a fatal outcome occurred in only one of them. Here, the DNA ploidy pattern was of the euploid (diploid or tetraploid) type in eight cases and of the aneuploid type in another eight. Our recently described "proliferation cell index" (PCI), based on nuclear immunoreactivity (IR) with the proliferation "marker" antigen Ki-67, was significantly lower in those five MCCs of the classical "DNA-diploid" type than in the seven "DNA-aneuploid" ones. These five patients presented a mild neoplastic disease; only one had a local recurrence and none had metastases. Otherwise, neither the PCI values nor the NCAM IR of the MCC cells were found to be of any prognostic significance.

High-molecular weight IGF-2 expression in a haemangiopericytoma associated with hypoglycaemia.

Spontaneous hypoglycaemia is usually caused by an insulin-producing islet-cell tumour of the pancreas. Rarely, it can be caused by non-islet cell tumours. Most of the tumours are of mesenchymal type, large, and slowly growing. One representative is haemangiopericytoma (HAP). The present report describes a case of a large recurrent retroperitoneal HAP associated with severe hypoglycaemia. Blood serum insulin and proinsulin concentrations were low. By means of acid-gel chromatography and dot-blot techniques, an increased amount of a high-molecular-weight IGF-2 peptide was found. By using antigen retrieval procedures, IGF-2-immunoreactive tumour cells were found in specimens of the recent tumour recurrence-but not in the original. When the in situ hybridization technique was used it could be shown that IGF-2 mRNA labelling had already occurred in the original tumour specimen, 11 years before the onset of hypoglycaemic symptoms. These observations confirm the hypothesized hypoglycaemic effects of high-molecular-weight (HMW) IGF-2, but also point to the presence of a prolonged compensation of this effect. A literature review, based on 17 similar cases of haemangiopericytoma with hypoglycaemia, is presented. Our observation and findings in the literature review support the idea that non-islet-cell tumour hypoglycaemia is caused by an overproduction of a HMW IGF-2 peptide. The insulin-like effect is mediated via non-specific binding to the insulin receptors. To anticipate patients at risk of developing this kind of hypoglycaemia, the histopathological investigation should include not only immunohistochemical analyses of the presence of IGF-2 peptide, but also in situ hybridization of the IGF-2 mRNA expression.

Prognostic significance of Ki-67 antigen and p53 protein expression in pancreatic duct carcinoma: a study of the monoclonal antibodies MIB-1 and DO-7 in formalin-fixed paraffin-embedded tumour material.

Formalin-fixed paraffin-embedded material from 57 patients in whom curative resection of pancreatic carcinoma had been attempted was analysed by an immunohistochemical procedure to estimate proliferation and p53 protein expression. Using the monoclonal antibody (MAb) MIB-1, which recognizes a Ki-67 epitope, the proliferating cell index (PCI, percentage of immunoreactive tumour nuclei) and proliferating cell area (PCA, percentage of immunoreactive tumour nuclear area) were calculated using an interactive image analysis system and were compared with semiquantitative scoring of stainability. MAb DO-7, which recognizes both wild- and mutant-type p53 protein, was used to assess p53 expression in the same material. MIB-1 stainings were of high quality in 53 tumours. The median PCI was 29.7% (range 0.5-82.1%) and the median PCA was 10.6% (range 0.0-36.5%). There was a close correlation between PCI and PCA (P < 0.0001). PCI and PCA values were in conformity with the semiquantitative scoring (P < 0.0001). The p53 immunohistochemical stainings were successful in 48 tumours and the protein was expressed in 22 (46%). High PCI values (> 45%, n = 14) correlated with shorter survival time (P < 0.01). PCA (P < 0.05) and the expression of p53 protein (P < 0.001) were independent prognostic variables.

A technique for automatic/interactive assessment of the proliferating fraction of neoplastic cells in solid tumors. A methodological study on the Ki-67 immunoreactive cells in human mammary carcinomas, including a comparison with the results of conventional S-phase fraction assessments by means of DNA cytometry.

Based on a computerized microscopy technique, a method has been devised which allows the practising pathologist to easily and rapidly assess quantitatively the relative number of actively proliferating neoplastic parenchymal cells in a tumor nodule. Our method has been tested on a series of 20 conventionally formalin-fixed and paraffin-embedded female mammary adenocarcinomas, using immunoreactivity with the MIB-1 monoclonal antibody against the cell proliferation antigen Ki-67. The values of the proportion of the MIB-1 immunoreactive cell nuclei were compared with those obtained DNA-cytometrically for the fraction of cells in the S-phase; a good correlation was found, although the MIB-1 values were consistently somewhat higher. A prerequisite for a success of the method was, of course, to achieve standardization of the MIB-1 immunostaining technique. By making simple adjustments of it, it could actually be improved to such an extent that almost the same color calibration and thresholding setup could be used. The measuring technique could be either interactive or automatic. The total number of immunoreactive and non-immunoreactive nuclei, as well as the total nuclear area of both cell types were registered in a computerized device. The data were accumulated sequentially for each measure field. To investigate the reproducibility of the immunostaining, two slides of each case were stained on different occasions. Each slide was measured three times; systemically randomly in the x- and y-axis-directions as well as in the subjectively defined histopathologically "most proliferative" area of the tumor. The values obtained were in good agreement with each other and obviously gave some valuable and objective supplementary pieces of information to that of the conventional clinical and histopathologic assessment of the degree of aggressiveness of a malignant neoplasm.

The value of cytometric DNA analysis as a prognostic tool in neuroendocrine neoplastic diseases.

In several traditionally non-endocrine, common, human, neoplastic diseases, it has become well established during the last few years, that cytometric analyses of the DNA distribution pattern of the nuclei of tumour cells can be an excellent supplement to the conventional prognostic tools, (such as clinical staging and histopathologic malignancy assessments). When analogous studies of the value of DNA analysis by means of flow cytometry and/or image cytometry are made in neuroendocrine (NE) neoplastic diseases, the ensuing results often become rather disappointing. Thus, clear-cut aneuploid DNA histograms can be found in the neoplastic cell nuclei of clinically and histopathologically completely benign NE adenomas (and even hyperplastic nodules). In contrast, highly aggressive NE carcinomas not seldom reveal themselves to be composed of tumour cells with nuclei, displaying an euploid, i.e. normal, DNA pattern. Statements of this kind have been based on the results of comprehensive investigations in several laboratories, analysing such NE tumours as insulomas/insular carcinomas, bronchial/gastrointestinal carcinoids, phaeochromocytomas, paragangliomas, neuroblastomas, adenomas of the anterior pituitary gland, parathyroid adenomas, medullary carcinoma of the thyroid and Merkel-cell tumours of the skin. Thus, the prognostic value of the cytometric DNA ploidy pattern of the nuclei of neoplastic parenchymal cells is definitely lower in NE tumours than in most of the traditionally non-endocrine carcinomas and sarcomas. Data from published and unpublished series of these kinds of NE tumours, and those of prostatic and breast carcinomas with NE differentiation, are given. By means of a new, consecutive double staining technique, it was shown that in idiopathic nesidioblastosis, the hyperinsulinism is caused by beta cells with a nuclear DNA ploidy pattern of euploid type. By the same technique, it can be shown that in the pathogenesis of the hypergastrinaemia-induced ECL-cell carcinoids of the stomach, a switch from an euploid to an aneuploid nuclear DNA distribution pattern occurs in the ECL-cells when they pass from a state of hyperplasia to that of a genuine neoplasia. In neuroblastomas, a triploid (i.e. aneuploid) DNA pattern is part of an algorithm capable of predicting a 96% survival rate, whereas a diploid/tetraploid (i.e. euploid) DNA pattern predicts a 0% survival.

c-myc oncogene amplification and cytometric DNA ploidy pattern as prognostic factors in musculoskeletal neoplasms.

The relationship between c-myc oncogene amplification in neoplastic cells as determined by means of Southern-blot analysis, and their nuclear DNA content as assessed by combined flow and image cytometry, was investigated in fresh tumor specimens from 33 patients with musculoskeletal neoplasms. Amplification, without rearrangement of the c-myc proto-oncogene, was detected in 4 out of 7 bone sarcomas and in 6 out of 26 soft-tissue sarcomas, but in none of 3 benign giant-cell bone tumors. Among the 10 cases with c-myc amplification, 2 were found to be cytometrically DNA diploid, 2 DNA tetraploid, and 6 DNA aneuploid. Conversely, there were 10 tumors displaying extremely aneuploid DNA patterns without c-myc oncogene amplification. Thus, there was no relationship between c-myc amplification and DNA ploidy; neither did the percentage of S-phase cells, as determined by means of image cytometry, correlate significantly with the occurrence of c-myc amplification. A surprising sex-bias was observed; all 6 cases of c-myc-amplified soft-tissue sarcomas occurred in females, whereas none of the 11 males with such sarcomas showed this amplification. When the clinical follow-up data of the patients were scrutinized, it was found that the DNA ploidy pattern of the neoplastic cell nuclei, in combination with the S-phase values, as well as the occurrence of c-myc amplification, yielded prognostic information, being statistically significant 2 years after the diagnosis.

Trk mRNA and low affinity nerve growth factor receptor mRNA expression and triploid DNA content in favorable neuroblastoma tumors.

Nerve growth factor (NGF) is suggested to play a role in spontaneous differentiation or regression of neuroblastoma. Expression of mRNAs for trk protooncogene and NGF low affinity receptor gene (LNGFR) were analyzed in 45 neuroblastomas with Northern Blot. Trk and LNGFR expression correlated with young age, localized tumors or IV-S disease, absence of N-myc amplification, triploid DNA content, spontaneous tumor regression and favorable prognosis. Regardless of other factors, trk and LNGFR mRNAs distinguished three prognostic subsets: one favorable (trk+, LNGFR+, n = 19, 100% survival probability), one intermediate (trk+, LNGFR-, n = 11, 62%) and one poor (trk-, n = 15, 0%). An algorithm based on the combined analysis of trk and LNGFR mRNAs and DNA ploidy divided the material in two groups with 96 and 0% survival probability respectively (p < 0.001), and predicted outcome accurately in 43 of 45 children. It is hypothesized that loss of functional NGF-receptors constitutes an early critical step in the evolution of unfavorable neuroblastomas prone to progression in spite of current therapy.

DNA ploidy in human colorectal adenocarcinomas.

Image cytometric determination of DNA content was performed on 207 colorectal adenocarcinomas surgically resected from 205 patients. Of these 207 tumors, 193 (93.2%) showed an aneuploid DNA distribution pattern, whereas the remaining 14 (6.8%) exhibited a DNA pattern corresponding to that found in proliferating diploid populations. The DNA content of 20 of these colorectal carcinomas was also studied by flow cytometric measurements on deparaffinized, disintegrated material. Flow cytometric measurement detected aneuploidy in only 9 of these 20 (45%) tumors, whereas 17 of the same 20 (85%) carcinomas showed aneuploidy when using image cytometric measurement on imprints. These results suggest that colorectal carcinomas are either aneuploid (vast majority) or proliferating diploid and that image cytometric DNA measurement on histologically controlled material is superior to flow cytometric measurement.