Prognostic relevance of the HER2 status of circulating tumor cells in metastatic breast cancer patients screened for participation in the DETECT study program.

BACKGROUND: Circulating tumor cells (CTCs) have been reported to predict clinical outcome in metastatic breast cancer (MBC). Biology of CTCs may differ from that of the primary tumor and HER2-positive CTCs are found in some patients with HER2-negative tumors. PATIENTS AND METHODS: Patients with HER2-negative MBC were screened for participation in DETECT III and IV trials before the initiation of a new line of therapy. Blood samples were analyzed using CELLSEARCH. CTCs were labeled with an anti-HER2 antibody and classified according to staining intensity (negative, weak, moderate, or strong staining). RESULTS: Screening blood samples were analyzed in 1933 patients with HER2-negative MBC. As many as 1217 out of the 1933 screened patients (63.0%) had ≥1 CTC per 7.5 ml blood; ≥5 CTCs were detected in 735 patients (38.0%; range 1-35 078 CTCs, median 8 CTCs). HER2 status of CTCs was assessed in 1159 CTC-positive patients; ≥1 CTC with strong HER2 staining was found in 174 (15.0%) patients. The proportion of CTCs with strong HER2 staining among all CTCs of an individual patient ranged between 0.06% and 100% (mean 15.8%). Patients with estrogen receptor (ER)- and progesterone receptor (PR)-positive tumors were more likely to harbor ≥1 CTC with strong HER2 staining. CTC status was significantly associated with overall survival (OS). Detection of ≥1 CTC with strong HER2 staining was associated with shorter OS [9.7 (7.1-12.3) versus 16.5 (14.9-18.1) months in patients with CTCs with negative-to-moderate HER2 staining only, P = 0.013]. In multivariate analysis, age, ER status, PR status, Eastern Cooperative Oncology Group performance status, therapy line, and CTC status independently predicted OS. CONCLUSION: CTC detection in patients with HER2-negative disease is a strong prognostic factor. Presence of ≥1 CTC with strong HER2 staining was associated with shorter OS, supporting a biological role of HER2 expression on CTCs.

Prognostic factors for and pattern of lymph-node involvement in patients with operable cervical cancer.

PURPOSE: Lymph node metastases significantly worsen the prognosis in cervical carcinoma. Risk factors-pathological and patient related-could select patients at high risk for lymph node involvement. METHODS: This retrospective analysis was performed by analyzing data from patients with cervical carcinoma treated between 2000 and 2017 at the Department of Obstetrics and Gynecology of the University Hospital Ulm. RESULTS: In total, 261 patients with cervical carcinoma (International Federation of Gynecology and Obstetrics (FIGO) stage IA-IIB) and lymphadenectomy with at least 10 removed lymph nodes were available for analysis. Overall, 86 (33.0%) patients had lymph node metastases; 73 patients had pelvic lymph node metastases only and 13 patients had both pelvic and paraaortic lymph node metastases. Lymph node metastases were found most often in the region of the external iliac artery and obturator fossa, with 57.0% and 54.7% of all 86 node-positive patients, respectively. Univariable analyses showed that presence of lymph node metastases was significantly associated with both preoperative FIGO stage (p = 0.001) and final pathological tumor stage (p < 0.001), status of resection margin (p = 0.002), lymphovascular space invasion (LVSI), (p < 0.001) and vascular space invasion, (p < 0.001). In a multivariable logistic regression model with presence of lymph node metastases (yes/no) as binary response variable, only LVSI (p < 0.001) and body mass index (BMI), (p = 0.035) remained as significant independent predictors of lymph node involvement. Subgroup analyses showed that LVSI was a significant predictive factor for lymph node involvement in patients with a preoperatively assessed FIGO stage < IIB (p < 0.001), but not for patients with a preoperatively assessed FIGO stage ≥ IIB (p = 0.122). CONCLUSIONS: The risk factor LVSI should play an important role in deciding whether an individualized therapy concept is based on escalating or deescalating treatment. In future, the sentinel concept could reduce morbidity and at the same time provide an important prognostic assessment for a subset of cervical cancer patients.

Pelvic exenteration as ultimate ratio for gynecologic cancers: single-center analyses of 37 cases.

BACKGROUND: Pelvic exenterations are a last resort procedure for advanced gynecologic malignancies with elevated risks in terms of patients' morbidity. METHODS: This single-center analysis reports surgical details, outcome and survival of all patients treated with exenteration for non-ovarian gynecologic malignancies at our university hospital during a 13-year time period. We collected data regarding patients and tumor characteristics, surgical procedures, peri- and postoperative management, transfusions, complications, and analyzed the impact on survival outcomes. RESULTS: We identified 37 patients between 2005 and 2013 with primary or relapsed cervical cancer (59.5%), vulvar cancer (24.3%) or endometrial cancer (16.2%). Median age was 60 years and most patients (73%) had squamous cell carcinomas. Median progression-free survival was 26.2 months and median overall survival was 49.9 months. The 5-year survival rates were 34.4% for progression-free survival and 46.4% for overall survival. There were no significant differences in progression-free survival and overall survival with regard to disease entity. Patients with tumor at the resection margins (R1) had a nearly significantly worse progression-free survival (median: 28.5 vs. 7.3 months, HR 2.59, 95% CI 0.98-6.88, p = 0.056) and a significantly worse overall survival (median: not reached vs. 10.9 months, HR 4.04, 95% CI 1.40-11.64, p = 0.010) compared to patients with complete tumor resection (R0). In addition, patients without lymphovascular space invasion had a significantly better progression-free survival (p = 0.017) and overall survival (p = 0.034) then patients with lymphovascular space invasion. We observed complications in 14 patients (37.8%), 10 of those were classified as Clavien-Dindo 3 or 4. There was a trend to worse progression-free survival in patients that suffered complications (p = 0.052). Median total amount of transfused blood products was 4 (range 0-20). CONCLUSION: Pelvic exenteration is a procedure that provides substantial progression-free survival and overall survival improvement and-in selected patients-can even achieve cure in otherwise hopeless clinical situations. Patients need to be offered earnest counseling for sufficient informed consent with realistic expectations what to expect.

Hypotension due to spinal anesthesia influences fetal circulation in primary caesarean sections.

PURPOSE: Hypotension due to spinal anesthesia is a well-known side effect in pregnant women receiving caesarean section. Little is known about its impact on fetal blood circulation. METHODS: 40 women with uncomplicated singleton term pregnancies prepared for caesarean section were prospectively evaluated by Doppler sonography before and immediately after spinal anesthesia. RESULTS: In 90% of the women, blood pressure significantly decreased after spinal anesthesia and 42.5% of the patients suffered from severe hypotension. We found a significant negative correlation between maternal blood pressure change and the resistant index (RI) of the umbilical artery (rs = - 0.376, p = 0.017) and a significant positive correlation between maternal blood pressure and fetal middle cerebral artery. CONCLUSION: Healthy fetuses seem to compensate well in situations with decreased uteroplacental blood flow due to maternal hypotension measured by means of RI changes in the fetal umbilical and middle cerebral artery. This raises the question if growth-restricted and/or preterm fetuses are able to compensate similarly or if general anesthesia would be a method of choice.

The HER2 phenotype of circulating tumor cells in HER2-positive early breast cancer: A translational research project of a prospective randomized phase III trial.

BACKGROUND: HER2 is one of the predominant therapeutic targets in breast cancer. The metastatic selection process may lead to discrepancies between the HER2 status of the primary tumor and circulating tumor cells (CTCs). This study analyzed the HER2 status of CTCs in patients with HER2-positive primary breast cancer at the time of diagnosis. Aim of the study was to assess potential discordance of HER2 status between primary tumor and CTCs, as this may have important implications for the use of HER2-targeted therapy. METHODS: The number and HER2 status of CTCs out of 30ml peripheral blood were assessed in 642 patients using the CellSearch System (Janssen Diagnostics, USA). The cutoff for CTC positivity was the presence of at least 1 CTC, and the cutoff for HER2 positivity of CTCs was the presence of at least 1 CTC with a strong HER2 staining. RESULTS: 258 (40.2%) of the 642 patients were positive for CTCs (median 2; range 1-1,689). 149 (57.8%) of these 258 patients had at least 1 CTC with strong HER2 staining. The presence of HER2-positive CTCs was not associated with tumor size (p = 0.335), histopathological grading (p = 0.976), hormone receptor status (ER: p = 0.626, PR: p = 0.263) or axillary lymph node involvement (p = 0.430). Overall, 83 (32.2%) of the CTC-positive patients exclusively had CTCs with strong HER2 staining, whereas 31 (12.0%) had only CTCs with negative HER2 staining. Within-sample variation in the HER2 status of CTCs was found in 86 (57.8%) of the 149 patients with more than 1 CTC. CONCLUSION: This study demonstrated that discordance between the HER2 expression of CTCs and that of the primary tumor frequently occurs in early breast cancer. Future follow-up evaluation will assess whether this discrepancy may contribute to trastuzumab resistance.

Toxicity Assessment of a Phase III Study Evaluating FEC-Doc and FEC-Doc Combined with Gemcitabine as an Adjuvant Treatment for High-Risk Early Breast Cancer: the SUCCESS-A Trial.

Introduction: This paper aims to evaluate the toxicity profile of additive gemcitabine to adjuvant taxane-based chemotherapy in breast cancer patients. Methods: Patients enrolled in this open-label randomized controlled Phase III study were treated with 3 cycles of epirubicin-fluorouracil-cyclophosphamide (FEC) chemotherapy followed by 3 cycles of docetaxel with those receiving 3 cycles of FEC followed by 3 cycles of gemcitabine-docetaxel (FEC-DG). 3690 patients were evaluated according to National Cancer Institute (NCI) toxicity criteria (CTCAE). The study medications were assessed by the occurrence of grade 3-4 adverse events, dose reductions, postponements of treatment cycles and granulocyte colony-stimulating factor (G-CSF) support. Results: No differences in neutropenia or febrile neutropenia were demonstrated. However, thrombocytopenia was significantly increased with FEC-DG treatment (2.0 vs. 0.5 %, p < 0.001), as was leukopenia (64.1 vs. 58.5 %, p < 0.001). With FEC-DG significantly more G-CSF support in cycles 4 to 6 (FEC-DG: 57.8 %, FEC-D: 36.3 %, p < 0.001) was provided. Transaminase elevation was significantly more common with FEC-DG (SGPT: 6.3 %, SGOT: 2 %), whereas neuropathy (1.2 %), arthralgia (1.6 %) and bone pain (2.6 %) were more common using FEC-D. Dose reductions > 20 % (4 vs. 2.4 %) and postponement of treatment cycles (0.9 vs. 0.4 %) were significantly more frequent in the FEC-DG arm. Eight deaths occurred during treatment in the FEC-DG arm and four in the FEC-D arm. Conclusion: The addition of gemcitabine increased hematological toxicity and was associated with more dose reductions and postponements of treatment cycles.

Randomised phase III trial of FEC120 vs EC-docetaxel in patients with high-risk node-positive primary breast cancer: final survival analysis of the ADEBAR study.

BACKGROUND: Taxane-containing adjuvant chemotherapy has been established as standard treatment in node-positive breast cancer. This study compared efficacy and tolerability of epirubicin (E)/cyclophosphamide (C) followed by docetaxel (Doc) with a dose-dense 5-fluorouracil (F)+E+ C regimen. METHODS: The ADEBAR study was a randomised phase III trial for women with primary invasive breast cancer and ⩾4 metastatic axillary lymph nodes (n=1364). Treatment consisted of four 21-day cycles of E plus C, followed by four 21-day cycles of Doc (EC-Doc), or six 28-day cycles of E plus F plus C (FEC120). RESULTS: Disease-free survival (DFS) was similar in the two treatment arms as shown by multivariate Cox regression adjusted for other prognostic factors (EC-Doc vs FEC120, hazard ratio (HR): 1.087; 95% confidence interval (CI): 0.878-1.346, P=0.444). In addition, there was no significant difference in overall survival (OS) between the two groups (HR: 0.974; 95% CI: 0.750-1.264, P=0.841). Haematologic toxicity was more common in FEC120 recipients; non-haematologic toxicities occurred more frequently in the EC-Doc arm. The serious adverse event rate was significantly higher in the FEC120 group (29.7% vs 22.5%). CONCLUSIONS: EC-Doc provides a feasible and effective alternative therapy option to FEC120 with a different safety profile in this high-risk breast cancer cohort.

Heterozygous PALB2 c.1592delT mutation channels DNA double-strand break repair into error-prone pathways in breast cancer patients.

Hereditary heterozygous mutations in a variety of DNA double-strand break (DSB) repair genes have been associated with increased breast cancer risk. In the Finnish population, PALB2 (partner and localizer of BRCA2) represents a major susceptibility gene for female breast cancer, and so far, only one mutation has been described, c.1592delT, which leads to a sixfold increased disease risk. PALB2 is thought to participate in homologous recombination (HR). However, the effect of the Finnish founder mutation on DSB repair has not been investigated. In the current study, we used a panel of lymphoblastoid cell lines (LCLs) derived from seven heterozygous female PALB2 c.1592delT mutation carriers with variable health status and six wild-type matched controls. The results of our DSB repair analysis showed that the PALB2 mutation causes specific changes in pathway usage, namely increases in error-prone single-strand annealing (SSA) and microhomology-mediated end-joining (MMEJ) compared with wild-type LCLs. These data indicated haploinsufficiency regarding the suppression of error-prone DSB repair in PALB2 mutation carriers. To the contrary, neither reduced HR activities, nor impaired RAD51 filament assembly, nor sensitization to PARP inhibition were consistently observed. Expression of truncated mutant versus wild-type PALB2 verified a causal role of PALB2 c.1592delT in the shift to error-prone repair. Discrimination between healthy and malignancy-presenting PALB2 mutation carriers revealed a pathway shift particularly in the breast cancer patients, suggesting interaction of PALB2 c.1592delT with additional genomic lesions. Interestingly, the studied PALB2 mutation was associated with 53BP1 accumulation in the healthy mutation carriers but not the patients, and 53BP1 was limiting for error-prone MMEJ in patients but not in healthy carriers. Our study identified a rise in error-prone DSB repair as a potential threat to genomic integrity in heterozygous PALB2 mutation carriers. The used phenotypic marker system has the capacity to capture dysfunction caused by polygenic mechanisms and therefore offers new strategies of cancer risk prediction.

Evaluation of two different analytical methods for circulating tumor cell detection in peripheral blood of patients with primary breast cancer.

BACKGROUND: Evidence is accumulating that circulating tumor cells (CTC) out of peripheral blood can serve as prognostic marker not only in metastatic but also in early breast cancer (BC). Various methods are available to detect CTC. Comparisons between the different techniques, however, are rare. MATERIAL AND METHODS: We evaluate two different methods for CTC enrichment and detection in primary BC patients: the FDA-approved CellSearch System (CSS; Veridex, Warren, USA) and a manual immunocytochemistry (MICC). The cut-off value for positivity was ≥1 CTC. RESULTS: The two different nonoverlapping patient cohorts evaluated with one or the other method were well balanced regarding common clinical parameters. Before adjuvant CHT 21.1% (416 out of 1972) and 20.6% (247 out of 1198) of the patients were CTC-positive, while after CHT 22.5% (359 out of 1598) and 16.6% (177 out of 1066) of the patients were CTC-positive using CSS or MICC, respectively. CTC positivity rate before CHT was thus similar and not significantly different (P = 0.749), while CTC positivity rate immediately after CHT was significantly lower using MICC compared to CSS (P < 0.001). CONCLUSION: Using CSS or MICC for CTC detection, we found comparable prevalence of CTC before but not after adjuvant CHT.

Does Non-Placement of a Drain in Breast Surgery Increase the Rate of Complications and Revisions?

Purpose: Although surgical therapy for breast cancer has become less radical, intrasurgical placement of drains and the use of compression bandages is still standard practice. However, evidence for the clinical benefit of wound drains is controversial, and use of drains is associated with increased pain and longer hospital stays. This raises the question whether, given the latest surgical techniques, wound drainage is still medically necessary. Material and Method: A retrospective analysis was done of patients with breast cancer treated surgically between January 2009 and April 2012 in the Breast Centre Hohenlohe (n = 573). Complication rates and revision following surgery with and without placement of wound drains were compared for patients who had breast-conserving surgery (n = 425) and patients who underwent mastectomy (n = 148). Results: The baseline characteristics (age, number of resected lymph nodes, numbers of patients who had sentinel lymph node resection, tumour characteristics, receptor status and affected side) were comparable for the investigated patient groups. The overall rate of complications was 4 %. There was no significant difference with regard to complication rates after surgery with and without placement of wound drains between the group of patients with breast-conserving surgery and the group of patients with mastectomy (p = 0.68 and p = 0.54, respectively). Conclusion: Our data indicate that non-placement of a wound drain does not influence complication or revision rates after breast-conserving surgery or mastectomy.

'Y'Scenedesmus (Chlorophyta, Chlorophyceae): the internal transcribed spacer 2 rRNA secondary structure re-revisited.

Including RNA secondary structures improves accuracy and robustness in reconstruction of phylogenetic trees. It is possible to simultaneously infer alignments and phylogenies on the primary sequence and the secondary structure information. For the internal transcribed spacer 2 (ITS2), a phylogenetic RNA transcript marker, two different structure conformations (I or Y shape for helix I) were published for Scenedesmaceae, and a third appeared in the ITS2 database. We contrast the effects on phylogenetic tree reconstruction of different structure sets for a small scenedesmacean subset, using neighbour-joining, maximum parsimony and, for the first time, maximum likelihood, on sequence-structure alignments. Generally our study supports inclusion of secondary structure information. However, we found that any of the three structure conformations is equally fit for phylogenetic studies, but prefer the I shape for helix I. Moreover, our results enable us to give general recommendations on how to build a phylogenetic tree using ITS2 sequence-structure alignments, including different methods to obtain the secondary structures. Thus, we hope to provide a valuable contribution not only for scenedesmacean ITS2 phylogeny, but also for other approaches using RNA transcript markers.

[Verrucous squamous cell carcinoma complicating hypertrophic lichen planus. Three case reports and review of the literature]. / Verruköses spinozelluläres Karzinom auf Lichen ruber hypertrophicus. Drei Fallberichte mit Literaturüberblick.

Lichen planus is a chronic mucocutaneous T-cell-mediated disease, whose cause is still unknown. The first case of lichen planus that transformed into squamous cell carcinoma was reported in 1903. We present three patients in whom squamous cell carcinomas were identified in chronic lichen planus. The world literature includes at least 91 cases, including our three cases. In an epidemiological study, no significant risk of transformation of cutaneous lichen planus into squamous cell carcinomas was found. In contrast, there is a significantly higher risk of malignant transformation in mucosal lichen planus, so that the WHO had graded mucosal lichen planus as a premalignant condition.

Deployment of the encapsulation-dehydration protocol to cryopreserve microalgae held at the Sammlung von Algenkulturen, Universitat Gottingen, Germany.

Encapsulation-dehydration was applied to cryopreserve 14 diverse algal strains, representing eukaryotic terrestrial microalgae; of these 12 survived to form cell colonies after recovery from cryostorage. Surviving algae had varying degrees of tolerance to osmotic dehydration and desiccation in this vitrification-based cryoprotective strategy. The extent of algal regrowth was affected by the mode of desiccation (silica gel or air-flow), the duration of evaporative desiccation and exposure to light during early recovery phase. This paper: (i) demonstrates the versatility of the encapsulation/dehydration method to cryopreserve diverse microalgae; (ii) confirms the successful transfer of this cryostorage technology to the Culture Collection of Algae at Gottingen University (SAG); and (iii) recommends encapsulation/dehydration as a feasible alternative to controlled rate cooling for preserving algae held in international culture collections.

Cryopreservation and conservation of microalgae: the development of a Pan-European scientific and biotechnological resource (the COBRA project).

Microalgae are one of the most biologically important elements of worldwide ecology and could be the source of diverse new products and medicines. COBRA (The COnservation of a vital european scientific and Biotechnological Resource: microAlgae and cyanobacteria) is the acronym for a European Union, RTD Infrastructures project (Contract No. QLRI-CT-2001-01645). This project is in the process of developing a European Biological Resource Centre based on existing algal culture collections. The COBRA project's central aim is to apply cryopreservation methodologies to microalgae and cyanobacteria, organisms that, to date, have proved difficult to conserve using cryogenic methods. In addition, molecular and biochemical stability tests have been developed to ensure that the equivalent strains of microorganisms supplied by the culture collections give high quality and consistent performance. Fundamental and applied knowledge of stress physiology form an essential component of the project and this is being employed to assist the optimisation of methods for preserving a wide range of algal diversity. COBRA's "Resource Centre" utilises Information Technologies (IT) and Knowledge Management practices to assist project coordination, management and information dissemination and facilitate the generation of new knowledge pertaining to algal conservation. This review of the COBRA project will give a summary of current methodologies for cryopreservation of microalgae and procedures adopted within the COBRA project to enhance preservation techniques for this diverse group of organisms.

Genetic diversity of algal and fungal partners in four species of Umbilicaria (Lichenized Ascomycetes) along a transect of the Antarctic peninsula.

Lichens from the genus Umbilicaria were collected across a 5,000-km transect through Antarctica and investigated for DNA sequence polymorphism in a region of 480-660 bp of the nuclear internal transcribed spacer region of ribosomal DNA. Sequences from both fungal (16 ascomycetes) and photosynthetic partners (22 chlorophytes from the genus Trebouxia) were determined and compared with homologs from lichens inhabiting more temperate, continental climates. The phylogenetic analyses reveal that Antarctic lichens have colonized their current habitats both through multiple independent colonization events from temperate embarkation zones and through recent long-range dispersal in the Antarctic of successful preexisting colonizers. Furthermore, the results suggest that relichenization-de novo establishment of the fungus-photosynthesizer symbiosis from nonlichenized algal and fungal cells-has occurred during the process of Antarctic lichen dispersal. Independent dispersal of algal and fungal cultures therefore can lead to a successful establishment of the lichen symbiosis even under harsh Antarctic conditions.

The distribution of group I introns in lichen algae suggests that lichenization facilitates intron lateral transfer.

The nuclear-encoded small subunit ribosomal DNA gene of many lichen-forming green algae in the genus Trebouxia contains a group I intron at Escherichia coli genic position 1512. We studied the evolutionary history of the 1512 intron in Trebouxia spp. (Trebouxiophyceae) by analyzing intron and "host" cell phylogenies. The host trees were constructed by comparing internal transcribed spacer regions of rDNA. Maximum-likelihood, maximum-parsimony, and distance analyses suggest that the 1512 intron was present in the common ancestor of the green algal classes Trebouxiophyceae, Chlorophyceae, and Ulvophyceae. The 1512 intron, however, was laterally transferred at least three times among later-diverging Trebouxia spp. that form lichen partnerships. Intron secondary structure analyses are consistent with this result. Our results support the hypothesis that lichenization may facilitate 1512 group I intron lateral transfer through the close cell-to-cell contact that occurs between the lichen algal and fungal symbionts in the developing lichen thallus.

Nuclear-encoded rDNA group I introns: origin and phylogenetic relationships of insertion site lineages in the green algae.

Group I introns are widespread in eukaryotic organelles and nuclear-encoded ribosomal DNAs (rDNAs). The green algae are particularly rich in rDNA group I introns. To better understand the origins and phylogenetic relationships of green algal nuclear-encoded small subunit rDNA group I introns, a secondary structure-based alignment was constructed with available intron sequences and 11 new subgroup ICI and three new subgroup IB3 intron sequences determined from members of the Trebouxiophyceae (common phycobiont components of lichen) and the Ulvophyceae. Phylogenetic analyses using a weighted maximum-parsimony method showed that most group I introns form distinct lineages defined by insertion sites within the SSU rDNA. The comparison of topologies defining the phylogenetic relationships of 12 members of the 1512 group I intron insertion site lineage (position relative to the E. coli SSU rDNA coding region) with that of the host cells (i.e., SSU rDNAs) that contain these introns provided insights into the possible origin, stability, loss, and lateral transfer of ICI group I introns. The phylogenetic data were consistent with a viral origin of the 1512 group I intron in the green algae. This intron appears to have originated, minimally, within the SSU rDNA of the common ancestor of the trebouxiophytes and has subsequently been vertically inherited within this algal lineage with loss of the intron in some taxa. The phylogenetic analyses also suggested that the 1512 intron was laterally transferred among later-diverging trebouxiophytes; these algal taxa may have coexisted in a developing lichen thallus, thus facilitating cell-to-cell contact and the lateral transfer. Comparison of available group I intron sequences from the nuclear-encoded SSU rDNA of phycobiont and mycobiont components of lichens demonstrated that these sequences have independent origins and are not the result of lateral transfer from one component to the other.