Search

Background: Circadian clocks involve 15 specific genes, which control cell cycle, apoptosis, DNA repair and metabolism. Circadian disruption is associated with both an increased risk of breast cancer, and poor survival in patients (pts) with metastatic cancer (IARC#98; Innominato et al. Cancer Res 2009). Objective: to provide first evidence of clinical relevance of clock genes expression for the therapeutic sensitivity of primary breast cancer. Method: Affymetrix Hgu133plus2 microarrays data were derived from 189 primary breast cancers. All the pts had non metastatic disease and were registered in multicenter Phase II trial of neoadjuvant chemotherapy with epirubicin-cyclophosphamide (4 courses) then docetaxel ± herceptin (4 courses). In this study, negative hormonal receptor status and limited tumor size predicted for pathological complete response rate (pCR) (Pierga JY, et al Breast Cancer Res Treat. 2010).The expression data of 15 core clock genes, within the whole tumor transcriptome, were related both to tumor mitotic index (MI) and estrogen receptors (ESR1), and to chemotherapy-induced pCR. Statistical validation involved T-test statistics, with corrections for multiple testing. Results: In the tumor biopsy obtained before treatment onset, both Per2 and Cry2 expressions correlated positively with ESR1 (r≥0.43; P Clock gene expression and pCR Conclusion: Low expressions of three core genes in the negative loop of the molecular circadian clock strongly predicted for the induction of a pathologic complete response of primary breast cancer with neoadjuvant chemotherapy. While circadian clock disruption seems to constitute a poor prognostic factor in cancer patients, our data suggest that the downregulation of core circadian genes in tumors significantly enhances susceptibility to chemotherapy. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P2-09-26.

KIEKKAS P., THEODORAKOPOULOU G., SPYRATOS F. & BALTOPOULOS G.I. (2010) Psychological distress and delusional memories after critical care: a literature review. International Nursing Review57, 288–296 Background: A considerable number of intensive care unit (ICU) survivors report delusional memories, which refer to dreams, nightmares, paranoid delusions and hallucinations experienced in the ICU. These memories often have a strong vividness, long duration and high emotional impact. Aim: The aim of this review was to investigate and synthesize published literature about psychological distress associated with delusional memories of adult ICU survivors. Methods: Using key terms, a search was conducted in major health care electronic databases [Cumulative Index for Nursing and Allied Health Literature (CINAHL), PubMed, Web of Science and PsycInfo] focusing on articles published between 1990 and 2009 in English-language journals. Findings: Ten articles met the inclusion criteria. Recall of delusional memories at various intervals after ICU discharge was associated with post-traumatic stress disorder (PTSD)-related symptoms in many studies, while associations with other aspects of psychological distress, mainly feelings of fear, anxiety and depression, were also reported. Recent studies did not seem to confirm the protective role of factual memories. Conclusions: The findings support the association between delusional memories and PTSD-related symptoms, but further research is needed to confirm their association with other psychological disorders. Development of a safety sense in the ICU can protect patients against the emotional impact of both delusional and stressful factual ICU memories. Appropriate follow-up of high-risk patients could improve their long-term psychological recovery.

The estrogen receptor α (ERα) status of breast tumors is used to identify patients who may respond to endocrine agents such as tamoxifen. However, ERα status alone is not perfectly predictive, and there is a pressing need for more reliable markers of endocrine responsiveness. In this aim, we used a two-step strategy. We first screened genes of interest by a pangenomic 44 K oligonucleotide microarray in a series of ten ERα-positive tumors from five tamoxifen-treated postmenopausal patients who relapsed (distant metastasis) and five tamoxifen-treated postmenopausal patients who did not relapse, matched with respect to age, Scarff–Bloom–Richardson grade, lymph node status, and macroscopic tumor size. Genes of interest (n=24) were then investigated in an independent well-characterized series of ERα-positive unilateral invasive primary breast tumors from postmenopausal women who received tamoxifen alone as adjuvant hormone therapy after primary surgery. We identified four genes (HRPAP20, TIMELESS, PTPLB, and MGC29814) for which high mRNA levels were significantly associated with shorter relapse-free survival (log-rank test). We also showed that hormone-regulated proliferation-associated 20 kDa protein (HRPAP20) and TIMELESS are 17β-estradiol-regulated in vitro and are ectopically expressed in OH-Tam-resistant cell lines. In conclusion, these findings point to HRPAP20 and TIMELESS as promising markers of tamoxifen resistance in women with ERα-positive breast tumors.

Quantitative reverse transcription-polymerase chain reaction (RT-PCR) used to detect minor changes in specific mRNA concentrations may be associated with poor reproducibility. Stringent quality control is therefore essential at each step of the protocol, including the PCR procedure. We performed inter-laboratory quality control of quantitative PCR between two independent laboratories, using in-house RT-PCR assays on a series of hormone-related target genes in a retrospective consecutive series of 79 breast tumors. Total RNA was reverse transcribed in a single center. Calibration curves were performed for five target genes (estrogen receptor (ER)α, ERβ, progesterone receptor (PR), CYP19 (aromatase) and Ki 67) and for two reference genes (human acidic ribosomal phosphoprotein PO (RPLPO) and TATA box-binding protein (TBP)). Amplification efficiencies of the calibrator were determined for each run and used to calculate mRNA expression. Correlation coefficients were evaluated for each target and each reference gene. A good correlation was observed for all target and reference genes in both centers using their own protocols and kits (P < 0.0001). The correlation coefficients ranged from 0.90 to 0.98 for the various target genes in the two centers. A good correlation was observed between the level of expression of the ERα and the PR transcripts (P < 0.001). A weak inverse correlation was observed in both centers between ERα and ERβ levels, but only when TBP was the reference gene. No other correlation was observed with other parameters. Real-time PCR assays allow convenient quantification of target mRNA transcripts and quantification of target-derived nucleic acids in clinical specimens. This study addresses the importance of inter-laboratory quality controls for the use of a panel of real-time PCR assays devoted to clinical samples and protocols and to ensure their appropriate accuracy. This can also facilitate exchanges and multicenter comparison of data.

Contexte. - La Federation nationale des centres de lutte contre le cancer (FNCLCC) et les Centres regionaux de lutte contre le cancer (CRLCC), en collaboration avec des partenaires des secteurs publics (CHU, CHG), prives et certaines societes savantes ont entrepris, depuis 1993, d'elaborer des recommandations pour la pratique clinique en cancerologie: les « Standards, Options et Recommandations » (SOR). L'objectif de l'operation SOR est d'ameliorer la qualite et l'efficience des soins aux patients atteints de cancer en fournissant aux praticiens une aide a la decision facilement utilisable. La methodologie d'elaboration des SOR repose sur une revue et une analyse critique des donnees de la litterature scientifique par un groupe pluridisciplinaire d'experts, permettant de definir, sur la base du niveau de preuve scientifique et du jugement argumente des experts, des Standards, des Options et des Recommandations. Avant publication, les SOR sont revus par des experts independants. Objectifs. - Definir, sur la base d'une revue de la litterature et de l'accord d'experts, des Standards, Options et Recommandations pour la radiotherapie des patientes atteintes de cancers de sein infiltrants non metastatiques. Methodes. - Un groupe pluridisciplinaire mis en place par la FNCLCC a revu les donnees scientifiques disponibles concernant les cancers du sein infiltrants non metastatiques. Apres selection des articles, synthese des resultats et redaction des SOR, le document a ete soumis pour relecture et approbation a 148 relecteurs independants. Resultats. - Ce document presente le chapitre radiotherapie de la mise a jour 2001 du SOR paru initialement en 1996. Les modifications des Standards, Options et Recommandations apportees dans cette actualisation sont liees a de nouvelles publications. Les principales recommandations sont: il l'irradiation du sein apres chirurgie conservatrice diminue significativement le risque de recidive locale (niveau de preuve A) et l'irradiation de la paroi apres mastectomie diminue d'autant plus le risque qu'il existe des facteurs de recidive locale (niveau de preuve A). (2) Lorqu'une chirurgie conservatrice du sein est effectuee, une radiotherapie mammaire doit toujours etre delivree, a la dose minimale de 50 Gy en 25 fractions (standard, niveau de preuve A). (3) Chez les femmes de moins de 50 ans, un complement d'irradiation doit toujours etre delivre dans le lit tumoral, y compris lorsque les berges sont saines (standard, niveau de preuve B). (4) L'irradiation de la chaine mammaire interne est indiquee lorque la tumeur est interne ou centrale, en l'absence d'envahissement ganglionnaire axillaire (accord d'experts) et dans tous les cas d'envahissement ganglionnaire axillaire (standard, niveau de preuve B1). (5) L'irradiation des ganglions sus- et sous-claviculaires est indiquee en presence d'envahissement ganglionnaire axillaire (standard, niveau de preuve B1).

Resume A cote des recepteurs des ˦strogenes et de la progesterone, de nombreux parametres biologiques tissulaires sont proposes dans le cancer du sein comme indicateurs de pronostic ou de reponse therapeutique. Il existe cependant a l'heure actuelle une dissociation entre l'evaluation parfois incomplete de ces facteurs et leur mise a disposition sur le marche, entrainant des couts non controles, voire des consequences nefastes pour les malades. La validation des parametres biologiques doit se concevoir en trois etapes: validation technique (evaluation des diverses approches methodologiques, standardisation des approches choisies et etablissement d'un controle de qualite), validation clinique dans le cadre de protocoles therapeutiques definis et selection des facteurs les plus performants. Ces etapes ne peuvent etre realisees a l'echelon individuel, du fait de l'importance de l'equipement technique et du recrutement necessaires. Le Groupe de biopathologie tissulaire et moleculaire s'est constitue en reseau de 30 laboratoires experts, universitaires, hospitaliers et de centres de lutte contre le cancer. Son but est de contribuer a la mise en place au niveau national de cette strategie d'evaluation, pour permettre la selection de nouveaux facteurs, reellement informatifs et non redondants, et susceptibles d'etre utilises en pratique clinique courante dans le respect des bonnes pratiques de laboratoire.

Les cliniciens souhaitent disposer de biomarqueurs selectionnant les patientes porteuses d’un cancer du sein RE+ qui pourraient ne pas recevoir de chimiotherapie. Le degre de proliferation cellulaire est un element essentiel dans ce choix : Ki-67 est connu depuis de nombreuses annees et est facilement realise par les anatomopathologistes, mais il n’est pas considere comme un standard dans la pratique quotidienne.

A considerable number of intensive care unit (ICU) survivors report delusional memories, which refer to dreams, nightmares, paranoid delusions and hallucinations experienced in the ICU. These memories often have a strong vividness, long duration and high emotional impact.The aim of this review was to investigate and synthesize published literature about psychological distress associated with delusional memories of adult ICU survivors.Using key terms, a search was conducted in major health care electronic databases [Cumulative Index for Nursing and Allied Health Literature (CINAHL), PubMed, Web of Science and PsycInfo] focusing on articles published between 1990 and 2009 in English-language journals.Ten articles met the inclusion criteria. Recall of delusional memories at various intervals after ICU discharge was associated with post-traumatic stress disorder (PTSD)-related symptoms in many studies, while associations with other aspects of psychological distress, mainly feelings of fear, anxiety and depression, were also reported. Recent studies did not seem to confirm the protective role of factual memories.The findings support the association between delusional memories and PTSD-related symptoms, but further research is needed to confirm their association with other psychological disorders. Development of a safety sense in the ICU can protect patients against the emotional impact of both delusional and stressful factual ICU memories. Appropriate follow-up of high-risk patients could improve their long-term psychological recovery.

The estrogen receptor alpha (ERα) plays a critical role in the pathogenesis and clinical behavior of breast cancer. To obtain further insights into the molecular basis of estrogen-dependent forms of this malignancy, we used real-time quantitative reverse transcription (RT)-PCR to compare the mRNA expression of 560 selected genes in ERα-positive and ERα-negative breast tumors. Fifty-one (9.1%) of the 560 genes were significantly upregulated in ERα-positive breast tumors compared with ERα-negative breast tumors. In addition to well-known ERα-induced genes (PGR, TFF1/PS2, BCL2, ERBB4, CCND1, etc.) and genes recently identified by cDNA microarray-based approaches (GATA3, TFF3, MYB, STC2, HPN/HEPSIN, FOXA1, XBP1, SLC39A6/LIV-1, etc.), an appreciable number of novel genes were identified, many of, which were weakly expressed. This validates the use of large-scale real-time RT-PCR as a method complementary to cDNA microarrays for molecular tumor profiling. Most of the new genes identified here encoded secreted proteins (SEMA3B and CLU), growth factors (BDNF, FGF2 and EGF), growth factor receptors (IL6ST, PTPRT, RET, VEGFR1 and FGFR2) or metabolic enzymes (CYP2B6, CA12, ACADSB, NAT1, LRBA, SLC7A2 and SULT2B1). Importantly, we also identified a large number of genes encoding proteins with either pro-apoptotic (PUMA, NOXA and TATP73) or anti-apoptotic properties (BCL2, DNTP73 and TRAILR3). Surprisingly, only a small proportion of the 51 genes identified in breast tumor biopsy specimens were confirmed to be ERα-regulated and/or E2-regulated in vitro (cultured cell lines). Therefore, this study identified a limited number of genes and signaling pathways, which better delineate the role of ERα in breast cancer. Some of the genes identified here could be useful for diagnosis or for predicting endocrine responsiveness, and could form the basis for novel therapeutic strategies.

resume Le controle de qualite europeen de l'EORTC qui s'appuie sur l'analyse de preparations cytosoliques ne cor role pas tous le aspects des dosages biochimiques des recepteurs de l'œstradiol (ER) et de la progesterone (PR). L'analyse aes distribuion, obtenues en routine est, de ce fait, egalement indispensable pour assurer la stabilite des mesures dans le temps, independamment du laboratoire et de la technique employee. Cette etude porte sur 83 907 dosages de recepteurs effectues au cours des 17 dern e es annee, par 17 laboratoires affilies au Groupe francais de biopathologie tissulaire et moleculaire. Les distributions en fonction de l'age montrent une faible variabilite interlaboratoire. Une bonne stabilite dans le temps est demontree pour la technique de liaison d'un radiol gand pou ER et PR, et pour la technique immunoenzymatique (EIA) pour PR. En revanche, on observe pour ER-EIA une augmentation sensible des medane; entre 1987 et 1989.

Recent advances in human, bacterial and viral genome projects and the development of quantitative real-time reverse transcription-polymerase chain reaction methods offer the possibility of analysing a large number of gene transcripts. These molecular developments represent an important advancein the field of genetics, cancer, virology, bacteriology and hematology. A limiting step remains the isolation of high quality mRNA purified from biological samples. This review describes the different methods used to isolate mRNA from biological samples and to verify RNA integrity and gives precise details about RNA storage conditions.

The "Standards, Options and Recommendations" (SOR) project, started in 1993, is a collaboration between the Federation of french cancer centers (FNCLCC), the 20 french cancer centers, and specialists from french public universities, general hospitals and private clinics. The main objective is the development of clinical practice guidelines to improve the quality of health care and the outcome of cancer patients. The methodology is based on a literature review and critical appraisal by a multidisciplinary group of experts, with feedback from specialists in cancer care delivery.To develop clinical practice guidelines for non metastatic breast cancer patients according to the definitions of the Standards, Options and Recommendations project.Data were identified by searching Medline, web sites, and using the personal reference lists of members of the expert groups. Once the guidelines were defined, the document was submitted for review to 148 independent reviewers.This article presents the chapter radiotherapy resulting from the 2001 update of the version first published in 1996. The modified 2001 version of the standards, options and recommendations takes into account new information published. The main recommendations are: (1) Breast irradiation after conservative surgery significantly decrease the risk of local recurrence (level of evidence A) and the decrease in the risk of local recidive after chest wall irradiation is greater as the number of risk factors for local recurrence increases (level of evidence A). (2) After conservative surgery, a whole breast irradiation should be performed at a minimum dose of 50 Gy in 25 fractions (standard, level of evidence A). (3) A boost in the tumour bed should be performed in women under 50 years, even if the surgical margins are free (standard, level of evidence B). (4) Internal mammary chain irradiation is indicated for internal or central tumours in the absence of axillary lymph node involvement (expert agreement) and in the presence of lymph node involvement (standard, level of evidence B1). (5) Sub- and supra-claviculr lymph node irradiation is indicated in patients with axillary node involvement (standard, level of evidence B1).

Both DNA ploidy and S-phase ploidy are promising prognostic factors for node-negative breast cancer patients. Based largely on the analysis of one large study, much of the reported problems with these factors have been caused by some unappreciated complexities in categorizing DNA ploidy into low- and high-risk groups and the lack of some necessary adjustments to eliminate unwanted correlations between DNA S-phase and ploidy. When both DNA ploidy and S-phase are compensated properly, they become independent prognostic markers, forming a powerful prognostic model.

We assessed the reliability of hormonal receptors (HR) by means of immunohistochemisty (IHC) on cell blocks obtained from diagnostic fine-needle cytopunctures in a group of 142 primary breast carcinoma. The results were compared to biochemical assessment (EIA) on their corresponding tissue samples (118 surgical specimens and 24 core needle biopsies). Percentage of stained nuclei and a score incorporating the proportion and the intensity of positive nuclei were evaluated. A two-group classification (cutoff 10% of stained nuclei) was used to define HR status. Highly positive tumors (= 50% of stained nuclei) were also individualized. Regarding HR status, concordance rate between immunostaining and biochemical assessment was 86.6% for ER and 76.8% for PR. Major discrepancies were found in 6.3% and 15.5% of cases for ER and PR, respectively. A good correlation was also observed between quantitative values obtained by the two methods (r = 0.69 for ER and 0.60 for PR). Discrepancies were mainly related to weak positive staining, values close to the respective cutoffs and when biochemical evaluation was performed on core needle biopsies. We conclude that IHC on cell blocks prepared from fine-needle cytopuncture specimens of breast carcinomas is useful as a routine procedure for hormonal receptor determination especially when planning neoadjuvant treatment.

Before replacing enzyme immunoassay of estrogen and progesterone receptors by immunohistochemistry, results of both methods were compared on 437 samples obtained from breast cancer patients (342 primary breast carcinomas, 16 local recurrences, 49 biopsies, and 30 tumor specimens obtained after neoadjuvant treatment). Immunohistochemistry (IHC) results were first assessed semiquantitatively on the basis of the estimated proportion of positive tumor cells, and then quantitatively using the "quick score." Semiquantitative IHC hormone receptors results (positive > or = 10%) correlated well with enzyme immunoassay status (positive >15 fmol/mg protein) in 358 surgical samples (342 primary tumors and 16 recurrences), with overall concordance rates of 89.9% and 82.1%, respectively. Among the 100 discordant cases, a large intraductal carcinoma component was observed in 7 of 36 cases for estrogen receptor (ER) and 15 of 64 for progesterone receptor (PR). Thirty-five discordant cases also were observed near the cut-off values. Hormone receptor levels by enzyme immunoassay correlated strongly with the quantitative IHC "quick score." Whatever the method, hormone receptor status was associated with histologic grade (SBR) and tumor size, whereas age correlated strongly with ER positivity. Similar results were obtained for biopsy specimens and posttreatment samples. This comparison improved the reliability of the IHC technique, which is currently routinely used for ER and PR determination in the authors' institution.

The lack of a standardized methodology for quantifying DNA ploidy and S-phase fraction (SPF) by flow cytometry is hindering routine use of these markers in breast cancer management. In a retrospective clinical multicenter study, we validated a standardized flow cytometry protocol. We tested 633 frozen T(1)T(2), N(0)N(1), M(0) breast tumors obtained in four institutions. Cell preparation was standardized, and precise rules for data interpretation were followed. Three SPF classes were defined on the basis of tertiles after adjustment for ploidy. DNA aneuploidy was observed in 61.0% of cases. No significant difference was observed among centers. Aneuploidy and high SPF were associated with large tumor size, node involvement, high histological grade, and hormone receptor negativity. In the overall population (median follow-up, 69 months), patients with medium and high SPF values had shorter disease-free survival (DFS) than those with low SPF values (P0.0001). Ploidy had no significant influence. By Cox analysis, SPF, pN, and estrogen receptor status were independent predictors of DFS (P = 0.0002, P = 0.001, and P = 0.05). In node-negative patients, SPF was the only predictor of DFS (P = 0.01), whereas in node-positive patients, the risk of relapse increased with both high SPF (P = 0.003) and estrogen receptor negativity (P = 0.004). Low SPF values distinguished grade II tumors with a particularly good outcome. Our results strongly support the use of SPF in multicenter studies and clinical trials and suggest that node-negative patients with slowly proliferating tumors do not require systemic adjuvant therapy.

The aim of this study was to evaluate c-erbB-2 overexpression by means of a quantitative biochemical technique in 488 primary breast cancer patients with long-term follow-up (median, 10 years) and its relation to other biochemical prognostic factors (uPA, p53, and epidermal growth factor receptor) and adjuvant therapy. High levels of c-erbB-2 (500 IU/mg protein) were associated with estrogen receptor (ER) and progesterone receptor negativity, high histoprognostic SBR grade and high levels of uPA and p53. Univariate analyses showed shorter metastasis-free survival (MFS) and overall survival (OS) in patients whose tumors overexpressed c-erbB-2 in the overall population, in subgroups defined by ER and uPA status, and in patients with positive pathological nodal status, SBR grade II, progesterone receptor, and p53-negative tumors. Patients with ER-positive, c-erbB-2-positive tumors had a shorter MFS and OS than those patients with c-erbB-2-negative tumors. No difference was observed between adjuvant-treated and untreated patients (chemotherapy and/or hormone therapy) in the c-erbB-2-negative subgroup. There was a trend toward a longer short-term MFS in c-erbB-2-positive patients treated with chemotherapy, whereas an opposite effect was observed with hormone therapy. Cox multivariate analyses showed that high levels of c-erbB-2 negatively influenced MFS in the overall population as well as in node-positive patients and in tamoxifen-treated patients, along with pN and uPA. Results for OS were comparable with those obtained for MFS. These results suggest that c-erbB-2 overexpression in breast cancer may be a better predictor of the response to tamoxifen than is ER status alone.

Among the proteases involved in the tumor invasion process, components of the plasminogen activator system (plasminogen activator type-urokinase uPA, its membrane receptor uPAR and its two inhibitors PAI-1 and PAI-2) appear to define high risk patients in primary breast cancer. As individual analysis of each component of the plasminogen activator system does not reflect the complex interactions between the different components, we studied the prognostic impact of a dissemination risk index combining the four variables. We found that this index was the most powerful prognostic factor, particularly in node-negative patients.

We used non-linear kernel discriminant analysis (KDA) to predict the outcome of 134 axillary node-negative primary breast cancer patients not treated with adjuvant therapy in a non censored database.Posterior probabilities of relapse at 5 years were estimated using probabilistic neural networks (PNN) and a cross-validation (leave-one-out) technique to avoid overfitting the data. A stepwise method was used to construct the models to define the best combination of risk factors among eleven prognostic factors: age, menopausal status, Scarff-Bloom-Richardson grade, clinical tumor size, pathological tumor size, estrogen and progesterone receptor status, urokinase-type plasminogen activator, p53 protein level, c-erbB-2 protein and epidermal growth factor receptor. The different variables were tested individually and in combination to determine their prognostic power using a ROC indicator, which measures the separation between the probability distributions of the output neuron activations under the null hypothesis (no recurrence at 5 years) and under the alternative hypothesis (recurrence at 5 years).The best predictive one-dimensional model was obtained with uPA (ROC indicator = 0.75). A two-factor model including uPA and clinical tumor size (T) gave the best discrimination between recurrence and non recurrence at 5 years (ROC indicator = 0.84). Additional variables did not improve the accuracy of the prediction. The uPA-T model generated a map useful in predicting the posterior probability of cancer recurrence in a given patient. This representation allows the entire database to be easily visualized and each patient can be compared with the entire database.This is a powerful approach to analyze the impact of prognostic factors and it could find clinical applications in breast cancer.

The "Standards, Options and Recommendations" (SOR) project, started in 1993, is a collaboration between the Federation of the French Cancer Centres (FNCLCC), the 20 French Cancer Centres and specialists from French Public Universities, General Hospitals and Private Clinics. The main objective is the development of clinical practice guidelines to improve the quality of health care and outcome for cancer patients. The methodology is based on literature systematic review and critical appraisal by a multidisciplinary group of experts, with feedback from specialists in cancer care delivery.To develop clinical practice guidelines according to the definitions of Standards, Options and Recommendations for endocrine therapy in patients with non metastatic breast cancer.Data have been identified by literature search using Medline, Embase, Cancerlit and Cochrane databases - until july 1999 - and the personal reference lists of the expert group. Once the guidelines were defined, the document was submitted for review to 125 independent reviewers.The main recommendations for the endocrine therapy of patients with non metastatic breast cancer are: 1) Endocrine therapy modalities depend on menopausal status or age of women: ovarian suppression for premenopausal women, antiestrogen drug therapy for postmenopausal women (standard). 2) Tamoxifen (20 mg/d - 5 years) is beneficial to women with positive estrogen receptor tumor (standard, level of evidence A). There is no indication of tamoxifen treatment for women with negative estrogen receptor tumor (standard, level of evidence A). 3) For postmenopausal women with positive estrogen receptor tumor, tamoxifen is the standard adjuvant treatment (level of evidence A). For postmenopausal women with negative estrogen receptor, adjuvant chemotherapy has to be considered (option, level of evidence A). No adjuvant treatment has to be considered for women with poor health condition (option). 4) For premenopausal women with estrogen receptor tumor, results of clinical trials of chemotherapy versus endocrine therapy, suggest a benefit for endocrine therapy. However, there is no sufficient evidence to consider endocrine therapy alone as a standard adjuvant treatment. 5) For premenopausal women, chemotherapy + ovarian suppression or chemotherapy + tamoxifen are not better than chemotherapy alone (level of evidence A). 6) For postmenopausal women, administration of chemotherapy plus adjuvant tamoxifen versus the same tamoxifen alone, is of additional benefit in reducing recurrences but not in prolonging overall survival (standard, level of evidence A). 7) Balance of known benefits (delay to recurrence and death) and risks (side-effects of therapy) for adjuvant chemoendocrine therapy has to be taken into consideration before decision making. Chemoendocrine therapy can be indicated for women at high risk of developing metastatic disease (recommendation, experts agreement).

As part of a clinical research project, proliferative parameters were studied in primary breast cancer: standardization and technical validation of thymidine kinase (TK), thymidylate synthase (TS) and protein tyrosine kinase (PTK) are described. A total of 633 frozen tumor specimens, available in four institutions, was analyzed in three flow cytometry laboratories for DNA content and percentage of S-phase cells (%S) measurement. 1) The standardization step consisted in developing a common protocol for sample preparation; then, common cell suspensions were analyzed in order to perform an inter-laboratory control. Objective guidelines were elaborated to interpret DNA histograms in breast carcinoma. 2) DNA-aneuploidy was observed in 61% of cases of the retrospective series. Compared with DNA-aneuploid tumors, mean %S was significantly lower in case of DNA-diploidy (respectively: 6.4% and 2.2%, p0.001). When compared between the four institutions, %S distributions did not differ significantly. 3) %S is strongly correlated with TK, TS and PTK and high percentages were also observed in high grade tumors or tumor without hormone receptors. These results show that a standardization in using flow cytometers and DNA software allows multicenter studies.

The aim of this study was to achieve a better definition of the respective indications of flow cytometry (FCM) and image cytometry (ICM) in clinical practice by comparing their efficiency in a series of 104 primary breast carcinomas that were sampled by means of fine-needle cytopuncture and analyzed by both methods. The comparison involved the DNA content and the estimation of S-phase fraction (SPF). For this purpose, a manual rectangular model was used for ICM, and Modfit software was used for FCM. With respect to DNA ploidy, the concordance rate between the two methods was 87% (DNA diploid vs. DNA nondiploid) and 81% when subclasses of DNA ploidy were used (hypodiploid, diploid, hyperdiploid, tetraploid, or multiploid). True discordance was observed in 19 cases (18%). Eleven of these discordances were the result of an underestimation of multiploidy by FCM. In the 91 cases evaluable by ICM, the SPF median value was 3.5 for DNA-diploid tumors and 8 for DNA-nondiploid tumors. In the 67 cases evaluable by FCM, the SPF median value was 2.31 for DNA-diploid tumors and 5.25 for DNA-nondiploid tumors. The concordance between ICM and FCM in the 46 uniploid tumors was 0.90. Our results led us to obtain samples for the two techniques systematically, to perform FCM routinely because it is more rapid and adequate in a large number of cases, to confirm by ICM any DNA-diploid or DNA near-diploid peak revealed by FCM, and to use ICM when FCM is inadequate, i.e., in case of low cellularity (fewer than 5000 cells for analysis of the cell cycle), suspected tetraploidy, samples not exclusively composed of malignant cells, the identification of several morphologically different malignant populations, or populations with large coefficients of variation and high background.

To evaluate nuclear grading on fine needle cytopunctures of breast carcinoma, which is of special interest when neoadjuvant chemotherapy is planned.In a prospective study, we compared cytologic grading, based on nuclear parameters (pleomorphism and mitosis), to modified Scarff-Bloom-Richardson histologic grading in 105 primary operable breast carcinomas. The results of these two nuclear grading systems were compared to Feulgen image analysis data from the corresponding cytologic samples.The concordance rate between the two grading systems was 76%. Concordance between cytologic and histologic grading was observed more frequently in purely invasive carcinomas (85%) than in cases combining invasive and in situ components (56%). A highly significant relationship was observed between the two grading systems and indices of proliferative activity (S-phase fraction, proliferation index, 5c exceeding rate and endoreduplication rate), particularly in concordant grading. Furthermore, nuclear area correlated with the results of the two grading systems.Cytologic nuclear grading appeared to be a reliable tool for a large proportion of breast tumors. Despite difficulties related to tumor heterogeneity, which could be detected by careful cytologic examination, it is a useful alternative to histologic grading.

Recent studies have suggested that human choriogonadotropin (hCG), in addition to its function in regulating steroidogenesis, may also play a role as a growth factor. Immunocytochemistry using two different monoclonal antibodies (LHR29 and LHR1055) raised against the human luteinizing hormone/human chorionic gonadotropin (LH/hCG) receptor allowed us to detect this receptor in breast cancer cell lines (T47D, MCF7, and ZR75) in individual cancer biopsies and in benign breast lesions. The receptor was also present in epithelial cells of normal human and sow breast. In the latter, its concentration increased after ovulation. The presence of LH/hCG receptor mRNA was confirmed by reverse transcription-PCR using primers extending over exons 2-4, 5-11, and 9-11. The proportion of LH/hCG-receptor positive cells and the intensity of the immunolabeling varied in individual biopsies, but there was no obvious correlation with the histological type of the cancer. These results are compatible with previous studies suggesting that during pregnancy, hCG is involved in the differentiation of breast glandular epithelium and that this hormone may play an inhibitory role in mammary carcinogenesis and in the growth of breast tumors.

Clinical use of flow cytometric (FCM) DNA analysis requires effective quality controls. Thirty-two laboratories with various degrees of FCM experience participated in the first phase of a quality control program organized by the Association Française de Cytométrie. All received diskettes containing ten list-mode files and ten histogram files that were derived from FCM analysis of various unfixed tumor specimens. A total of 610 responses on DNA ploidy and cell cycle were obtained with three different DNA analysis softwares: CellFit used by (44% of responses), MultiCycle (44%), and ModFit (12%). After statistical analysis, 31% of the responses were excluded from the final analysis for precise reasons. The groups were too small to carry out a valid analysis of the slight differences in the percentage of cells in the DNA synthesis phase (S%) between CellFit and MultiCycle. To estimate the influence of gating on the final cell-cycle results, five of the histogram files were derived from corresponding list-mode files, but the participating laboratories were unaware of this. A good correlation (r = 0.98) was obtained for S% values in the five paired files. The fact that 31% of the responses had to be excluded clearly reflects inadequate training in the use of these analysis softwares and, in some cases, a failure to grasp the biological meaning of the results. In contrast, the laboratories fulfilling consensus recommendations obtained remarkably homogeneous results, showing that standardization is feasible.

All French laboratories that routinely assay estradiol (ER) and progesterone (PR) receptors participate in the European EORTC quality control program based on twice-yearly analysis of 5 cytosolic preparations. This system has considerably reduced inter-laboratory variations, but does not cover all aspects of these assays. Analysis of receptor value distributions is also crucial to ensure that receptor measurements remain stable with time, independently of the laboratory and assay method. This study involved 83907 receptor assays carried out in the last 17 years by 17 laboratories belonging to the French Study Group on Tissue and Molecular Biopathology. The assays were based on radioligand binding (RLA) or immunoenzymology (EIA). For each laboratory, the medians and positivity rates were analysed according to two totally objective criteria, the patient's age and the year of assay, and according to histological grade and histological type of the tumor in order to verify the correlations classically described. Age-related distributions varied little between laboratories, compared with data published by 7 European EORTC laboratories [1]. The results remained relatively stable with time in the RLA method for ER and PR, and in the EIA method for PR. Median ER-EIA data showed a marked increase between 1987 and 1989, mainly due to changes in the quality of Abbott reagents during this period. Otherwise, this analysis confirms previous pathophysiological observations.

The urokinase type plasminogen activator (u-PA) and the plasminogen activator inhibitor-1 (PAI-1) are among the best second-generation prognostic tissue factors in breast cancer. However, different extraction procedures and assay kits are used in different laboratories. A total of 79 breast tumour tissues stored in liquid nitrogen were analysed in this study. We compared u-PA and PAI-1 levels determined with the American Diagnostics (AD) kit after various extraction procedures. The median cytosolic extraction yield in the presence of 0.4 mol/l KCl, calculated relative to extraction in the presence of 10 ml/l Triton X100 when adapted to standard laboratory working hours (incubation for 2 h instead of 12 h) was 74.4% for u-PA and 85.8% for PAI-1. In addition, the correlations were acceptable. Cytosolic extracts prepared with KCl could permit optimal u-PA and PAI-1 assays while also enabling hormone receptors to be determined with the same specimens. Further studies with clinical data are now necessary to determine the prognostic relevance of this extraction procedure.

It is now clearly established that proteolytic enzymes, and in particular plasminogen activator (uPA), play an important role in breaking down the extracellular matrix, which is considered to be a step in metastasis formation. Plasminogen activators are controlled at various levels. Two inhibitors, PAI-1 and PAI-2, have been identified, the latter being more specific for uPA. In attempts to determine their prognostic value, it is essential to investigate the relative importance of these parameters and their interactions. We used an immunoenzymatic method to assay uPA, PAI-1 and PAI-2 antigens in cytosols prepared from 314 primary breast tumors. The patients were followed up for a minimum of six years and all relevant clinical and laboratory findings had been recorded. Univariate analysis confirmed the poor outcome of patients whose tumors contained large amounts of uPA and PAI-1. In addition, low levels of PAI-2 correlated with shorter disease-free survival in the overall population (P = 0.02), post-menopausal women (P = 0.02) and women without lymph node involvement (P = 0.02). Multivariate analysis using the "Main Effects" Cox model identified node involvement, macroscopic tumor size and PAI-2 as significant variables. The "interactive" Cox model, taking into account interactions between uPA and its two inhibitors, identified a first subgroup with a very poor prognosis associating either high levels of PAI-1 with low levels of PAI-2 in the overall population as well as following stratification for axillary node negative disease, or high levels of uPA with low levels of PAI-2 in the group of menopausal women. We conclude that PAI-1 provides the same prognostic informations as uPA, and does not appear to play its role as an inhibitor. In contrast, PAI-2 increased the prognostic value of both uPA, particularly in post-menopausal women, as well as PAI-1 in a subgroup of axillary node negative patients.

The use of prognostic factors to help select breast cancer patients for adjuvant therapy is of considerable concern to the oncology community. This need for selection of prognostically less favorable cases is stimulating investigators to identify new and more powerful prognostic factors. Unfortunately however, this identification process is becoming more confusing because of a lack of guidelines for investigators to use to study new factors and for reviewers and readers to use to evaluate papers on this topic. In this paper, we will describe across our experience the main problems encountered in the study of biological prognostic studies. Considering evaluation criteria to be developed in the future, it appears that only multicentric and multidisciplinary structures are able to define decisional trees based on technically and clinically validated parameters in particular patients subgroups. Such a structure exists at the european level ("Receptor Study Group" of the EORTC) and a similar structure has now been created in France to answer these questions.

Pilot retrospective studies have pointed to the prognostic value of thymidine kinase (TK) in breast cancer. We studied the Prolifigen TK-REA assay (Sangtec Medical, Sweden), usually applied to serum, for TK analysis in breast cancer cytosols. Reproducibility was good, provided that small volume pipetting was performed carefully. The TK assay was not influenced by the short-term storage of cytosols in liquid nitrogen or at -80 degrees C. However, some steps appeared critical for good laboratory practice. The TK level was affected by thawing the cytosols more than twice. Tumour storage in liquid nitrogen should be preferred over storage at -80 degrees C. The components of the homogenisation buffer, especially sodium molybdate and KCl can have a marked influence on results. Finally, linearity problems arose with some cytosols. Thus, although assay of TK in cytosols is apparently simple, care must be taken in practice. The TK-REA kit should be standardised before widespread use in breast cancer.

Among the great many prognostic factors currently available in breast cancer, three classes of tissue biological parameters appear to be the most reliable in the establishment of a clinical decision flowchart, when they will have been technically and clinically validated: parameters of hormone dependence, tumour aggressiveness and invasion and parameters of proliferation. This article discusses the difficulties encountered in the evaluation of some of these parameters (hormone receptors by various methodological approaches, proteases, enzymes involved in cell proliferation), with emphasis on standardisation of techniques, development of quality controls, clinical validation and objective information of the medical and scientific communities.

Parathyroid hormone-related protein (PTHrP) is associated with the syndrome of humoral hypercalcemia of malignancy. A high incidence of positive staining for PTHrP is observed in breast cancer and positivity is more frequent in patients who develop bone metastases. We assessed the presence of PTHrP mRNA by using the polymerase chain reaction in 38 normocalcemic breast cancer patients with long-term follow-up (minimum, 5 years) selected for the presence or absence of later bone metastasis development. In all the patients except one, the PTHrP gene was expressed in the breast tumor. The level of amplified PTHrP complementary DNA was inversely related to age (P0.02) and positively related to the proportion of invaded nodes (P0.02) but was not related to the other usual prognostic factors. The level of PTHrP mRNA was not different between the group of patients without recurrence or metastases (n = 11) and the group of patients who later developed metastases in soft tissues (n = 10). By contrast, patients who subsequently developed bone metastases (n = 17) showed higher PTHrP gene expression than patients in the other two groups (P0.001). This study suggests that strong PTHrP gene expression in breast tumors is associated with the onset of bone metastases.

An enzyme-linked immunosorbent assay using monoclonal and polyclonal antibodies against recombinant pS2 was devised. It was used to measure pS2 concentration in the cytosol of 339 breast cancer, 15 fibroadenomas, 16 cases of benign breast disease, and 6 normal breast tissues. The mean value of pS2 concentration was higher in cancer, but the protein could be detected readily in benign tumors and even in normal breast. The concentration of pS2 was significantly lower in postmenopausal women and tumors of differentiation Grade 3. The pS2 concentration was correlated strongly with the presence of estrogen receptors (ER) and progesterone receptors (PR). No correlation was observed with the size, histologic type of the tumor, and lymph node status. The prognostic value of pS2 appeared relatively limited. It was clear cut only for a relatively small group of patients (approximately 15%), who had low concentrations of pS2 (less than or equal to 0.32 ng/mg of protein). These patients had a shorter disease-free interval and overall survival time. The most striking correlation was observed with the outcome of adjuvant hormone therapy. pS2 concentration was shown to be the most potent prognostic factor, preceding even ER.

Background: The aim of this study was to investigate and quantify the contribution of transcriptomic markers, in addition to strong predictors such as oestrogen receptor status, to the prediction of pathological complete response (pCR) in locally advanced breast cancer.Patients: The RNA profiles were analyzed using U133 plus 2.0 Affymetrix. We included 189 patients out of 340 patients entered in a neoadjuvant chemotherapy trial for large operable and locally advanced breast cancer. After four cycles of epirubicin–cyclophosphamide, patients were randomly allocated to four cycles of docetaxel with or without celecoxib for patients with HER2-negative tumors, and docetaxel with or without trastuzumab for patients with HER2-positive tumors, respectively. Proportions of pCR in each group were equal to 0.12, 0.16, 0.15 and 0.24 respectively. Patients who received trastuzumab (N=36) were discard from our example, in order to deal with similar proportions of pCR.Methods: The whole sample was divided into a training set (N=81) and a validation set (N=72). Using the training set, two predictive models were built using multivariate logistic regression models. In the first model (M1), usual clinical and biological significant markers were included. In the second model (M2), in addition to the significant parameters of M1, significant transcriptomic variables were included. Diagnostics of both predictive models were assessed on the validation set through sensitivity and specificity estimates. Simulations were performed to investigate stability of model M2.Results: In M1, oestrogen receptor status and tumor size were found to have a strong predictive role in the prediction of pCR. In addition to these classical markers, genes belonging to biological pathways involved in proliferation and microtubule stabilization appeared to have a strong role in the prediction of pCR (model M2). Validation of M1 on the validation set provided 70% of sensitivity and 86% of specificity. Validation of M2 on the validation set yielded to a better sensitivity of 80% and a specificity of 81%. Using simulations, we showed that several different predictive models M2 yielded to similar performances on the validation set. Conclusion: Our study showed that transcriptomic markers provided significant information in addition to usual biological markers for the prediction of pCR. In addition, predictive model with both usual and transcriptomic markers may lead in an improvement of the classification performances. However, as illustrated by simulations, predictive models with both classical and transcriptomic markers are not exclusive. The contribution of transcriptomic data for the prediction of pCR is straightforward, but finding a stable predictive model remains a great challenge.Supported by PHRC AOM/2OO2/02117, Pfizer inc., Roche, sanofi-aventis.ISRCTN10059974 Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 2035.

Background: We have previously demonstrated that circulating tumor cells (CTC) detection by the CellSearch system is strongly associated with metastatic outcome in non-metastatic breast cancer (BC) treated by neoadjuvant chemotherapy (NACT) [Pierga, Clin Cancer Res 2008]. Transcriptomic analysis of primary tumors may uncover molecular phenotypes associated with CTC detection.Methods: Both CTC detection in blood at diagnosis and transcriptomic analysis of the primary tumor have been performed prospectively in 58 non-metastatic BC patients (pts) treated by NACT in a phase II trial (REMAGUS02). We searched for an association between CTC detection and (i) intrinsic molecular subtypes, (ii) stemness signature, (iii) other published signatures, and (iv) expression of molecular markers involved in CTC detection. CTC-associated genes were also studied (v).Results: (i) CTC detection was not statistically associated with an intrinsic molecular subtype: 29% in basal-like (n=5/17pts), 33% in HER2+ (3/9pts), 13% in luminal A (2/13pts), 20% in luminal B (2/10pts) and 29% in normal-like (2/7pts) BC. (ii) No association was found between CTC detection and the stemness signature. (iv) CTC detection was also not related to the “stemness profile”; it was independent of EpCAM, CK8 and CK18 transcriptomic expression. Other results (iii, v) will be disclosed at the meeting.Conclusion: Our study is the first to compare tumor gene expression profiles and blood dissemination of cancer cells in early BC pts. In these pts, CTC detection by the CellSearch system does not depend on intrinsic molecular subtypes, contrary to a published report based on in vitro-grown cells lines [Sieuwerts, J Natl Cancer Inst 2009]. Being independent from molecular prognostic factors, CTC detection is likely to play a critical role in early BC management.Supported by PHRC AOM/2OO2/02117, Pfizer inc., Roche, sanofi-aventis.ISRCTN10059974 Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 3005.

Purpose: To assess anti tumor activity of sequential epirubicin/cyclophosphamide followed by docetaxel with the randomized addition of celecoxib In Her2-ve patients or trastuzumab in Her2+ve patients versus no additional treatment, in terms of pathological complete response (pCR), defined as absence of residual invasive breast carcinoma and of nodal involvement.Patients and methods: 340 patients with stage II and III breast adenocarcinoma, with tumors ineligible for breast conservative surgery received 8 sequential 3 weekly cycles with epirubicin (75mg/m2)-cyclophosphamide (750mg/m2) for 4 cycles followed by docetaxel (100 mg/m2) for 4 cycles. According to HER2 status (IHC + FISH) they were randomized to receive together with docetaxel, in HER2 negative group (220 patients) celecoxib 800 mg/day during cycles 5-8 or no additional treatment and in HER2 positive group (120 patients) trastuzumab (8mg/kg then 6mg/kg) infused together with docetaxel or no additional treatment. All patients except eight with HER2+ tumor received adjuvant trastuzumab for a cumulative duration of 12 months.Results: In the HER2 negative group, pCR (grade 1 and 2 of Chevallier's classification) was observed in 11.5% and 13% of patients treated without and with neoadjuvant Celecoxib respectively. In the HER2+ group, pCR rate reached 26% in those having received neoadjuvant trastuzumab versus 19% in the others. There was no cardiac toxicity and no toxic death. Triple negative breast cancers experience the highest pCR rate of 30%.Conclusion: It is the first report on the effect of the addition of celecoxib to chemotherapy in neoadjuvant setting in breast cancer, showing that celecoxib does not improve the pCR rate. Addition of trastuzumab does, yet not to the extent reported with a protracted co-administration before surgery. Expression of hormonal receptors appears to be the major prognosticator for pCR. Molecular studies of gene expression profiling should allow improving such prediction.Supported by PHRC AOM/2OO2/02117, Pfizer inc., Roche, sanofi-aventis.ISRCTN10059974 Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5054.

Non-surgical first-line treatments of operable breast cancer raise the problem of pre-therapeutic collection of diagnostic and prognostic data. The advantages and limitations of cytopuncture and microbiopsy are discussed. The authors consider that the safest procedure is cytopuncture as part of diagnostic evaluation, completed by cutting-needle biopsy for information on tissues.

This study concerns the correlation between ER and PR status, menopausal status and the effect of adjuvant hormonotherapy in high risk primary breast cancer patients. We have compared the results obtained in a randomized trial (Institut Paoli-Calmettes, Marseille) with those of a historic series (Centre René Huguenin, Saint-Cloud). The patients presented the same clinical and histological criteria and received identical therapeutic protocols (chemotherapy and/or hormonotherapy). Compared with patients receiving no adjuvant treatment, it appeared that: 1) in post-menopausal patients, a significant detrimental effect of tamoxifen was found in ER-negative patients, while in ER-positive patients hormonal treatment was well correlated with both the presence and level of steroid receptors; 2) in pre-menopausal patients, hormonal therapy (oophorectomy + tamoxifen) appeared to be mediated by a complex mechanism involving more than an ER-positive cell population. In the light of the published results, the present findings underline the importance of reevaluating the indications of hormonotherapy in terms of hormone receptors and menopausal status. They also indicate the importance of biological factors in the evaluation of response to therapy. They can identify, subsets of patients in whom a given therapeutic protocol is detrimental, even though it may be beneficial for the overall population. heterogeneity in response to therapy among patient subsets is one of the most important problems which confronts medical statisticians and clinical investigators.

PTHrP, a newly characterized peptide is responsible for humoral hypercalcemia of malignancy in squamous cancers. It is also expressed by a variety of normal tissue and might have growth factor propriety. It is expressed by lactating breast and is frequently present in breast cancer. Although it is responsible for humoral hypercalcemia in an animal model of breast cancer its role in breast cancer hypercalcemia is still putative. The possibility that it might be one of the autocrine growth factors implicated in breast cancer cell proliferation has been suggested.

1038 Background: Breast cancer with synchronous metastases is considered as a noncurable disease for which systemic treatment remains the cornerstone of management. However several observational studies have reported a survival benefit for those patients (pts) who were offered a complete excision of the primary tumor. In such setting, locoregional treatment (LRT) may also consist of exclusive locoregional radiotherapy (LRR), as favoured in our institution for the past 2 decades. From our database, we have evaluated the impact of this alternative LRT to surgery on survival of breast cancer pts presenting with synchronous metastases. Methods: Among 18,753 cases diagnosed and treated at Rene Huguenin Cancer Center between 1980 and 2004, 598 pts (3.2%) presented with initial simultaneous distant metastasis. Information regarding demographics, tumor characteristics, site(s) of metastases and treatments were prospectively recorded in the database. The impact of delivery of LRT on overall survival (OS) was evalu...

Breast cancer is the most common of all cancers affecting women in France; its frequency increases in countries with a high standard of living. A family history and certain types of mastosis are unquestionable risk factors, although their weight has not yet been well established, but there is no absolute proof that feeding habits (notably fats and alcohol), which have been blamed by some authors, play a role in the genesis of breast cancer. Among the classical prognostic factors, which are necessary for surgical decisions, the size of the tumour, its histological grade and above all the number of axillary lymph nodes involved are the most important. However, a better knowledge of breast cancer biology has yielded factors that seem to be more promising than hormonal receptors, notably the DNA content of tumoral cells and the presence or absence of a protease, procathepsin 52 K, which reflects tumoral aggressiveness. As for the study of oncogens described elsewhere in this monograph, it will provide a better definition of high risk subjects and more precise information on the progress of the cancer.