MLLT11 Regulates Endometrial Stroma Cell Adhesion, Proliferation and Survival in Ectopic Lesions of Women with Advanced Endometriosis.

MLLT11 is a gene implicated in cell differentiation and the development and progression of human cancers, but whose role in the pathogenesis of endometriosis is still unknown. Using quantitative RT-PCR and immunohistochemistry, we analyzed 37 women with and 33 women without endometriosis for differences in MLLT11 expression. We found that MLLT11 is reduced in the ectopic stroma cells of women with advanced stage endometriosis compared to women without endometriosis. MLLT11 knockdown in control stroma cells resulted in the downregulation of their proliferation accompanied by G1 cell arrest and an increase in the expression of p21 and p27. Furthermore, the knockdown of MLLT11 was associated with increased apoptosis resistance to camptothecin associated with changes in BCL2/BAX signaling. Finally, MLLT11 siRNA knockdown in the control primary stroma cells led to an increase in cell adhesion associated with the transcriptional activation of ACTA2 and TGFB2. We found that the cellular phenotype of MLLT11 knockdown cells resembled the phenotype of the primary endometriosis stroma cells of the lesion, where the levels of MLLT11 are significantly reduced compared to the eutopic stroma cells of women without the disease. Overall, our results indicate that MLLT11 may be a new clinically relevant player in the pathogenesis of endometriosis.

Atezolizumab plus nab-paclitaxel for unresectable, locally advanced or metastatic breast cancer: real-world results from a single academic center in Austria.

PURPOSE: IMpassion130 led to the approval of atezolizumab plus nab-paclitaxel as first-line treatment for patients with unresectable locally advanced or metastatic triple-negative, PD-L1 immune-cell positive breast cancer (BC) by the European Medicines Agency (EMA). The objective of the present study was to investigate the implementation, safety and efficacy of this combination in the initial phase after approval. METHODS: A retrospective data analysis including all BC patients who received atezolizumab and nab-paclitaxel between 1.1.2019 and 31.10.2020 at the Department of Obstetrics and Gynecology and the Department of Medicine 1, respectively, at the Medical University of Vienna, Austria, was performed. Progression-free survival (PFS) and overall survival (OS) were estimated with the Kaplan-Maier product-limit method. Owing to the retrospective nature of this study, all statistics must be considered exploratory. RESULTS: In total 20 patients were included in the study. Median follow-up was 7.1 months (IQR 5.2-9.1). Median PFS was 3.0 months (SE = .24; 95% CI [2.5; 3.5]). Median OS was 8.94 months (SE = 2.34, 95%CI [4.35; 13.53]). No new safety signals were observed. CONCLUSION: The present study showed a considerably shorter PFS (3.0 vs. 7.5 months) and OS (8.94 vs. 25.0 months) than IMpassion130 putatively owing to the use of atezolizumab in later treatment lines, more aggressive tumors and a study population with higher morbidity compared to the pivotal trial.

Conventional versus reverse sequence of neoadjuvant epirubicin/cyclophosphamide and docetaxel: sequencing results from ABCSG-34.

BACKGROUND: Preoperative chemotherapy containing anthracyclines and taxanes is well established in early-stage breast cancer. Previous studies have suggested that the chemotherapy sequence may matter but definitive evidence is missing. ABCSG trial 34 evaluated the activity of the MUC1 vaccine tecemotide when added to neoadjuvant treatment; the study provided the opportunity for the second randomisation to compare two different anthracycline/taxane sequences. METHODS: HER2-negative early-stage breast cancer patients were recruited to this randomised multicentre Phase 2 study. Patients in the chemotherapy cohort (n = 311) were additionally randomised to a conventional or reversed sequence of epirubicin/cyclophosphamide and docetaxel. Residual cancer burden (RCB) with/without tecemotide was defined as primary study endpoint; RCB in the two chemotherapy groups was a key secondary endpoint. RESULTS: No significant differences in terms of RCB 0/I (40.1% vs. 37.2%; P = 0.61) or pathologic complete response (pCR) rates (24.3% vs. 25%, P = 0.89) were observed between conventional or reverse chemotherapy sequence. No new safety signals were reported, and upfront docetaxel did not result in decreased rates of treatment delay or discontinuation. CONCLUSION: Upfront docetaxel did not improve chemotherapy activity or tolerability; these results suggest that upfront neoadjuvant treatment with anthracyclines remains a valid option.

Aggressive B-cell lymphomas in patients with myelofibrosis receiving JAK1/2 inhibitor therapy.

Inhibition of Janus-kinase 1/2 (JAK1/2) is a mainstay to treat myeloproliferative neoplasms (MPN). Sporadic observations reported the co-incidence of B-cell non-Hodgkin lymphomas during treatment of MPN with JAK1/2 inhibitors. We assessed 626 patients with MPN, including 69 with myelofibrosis receiving JAK1/2 inhibitors for lymphoma development. B-cell lymphomas evolved in 4 (5.8%) of 69 patients receiving JAK1/2 inhibition compared with 2 (0.36%) of 557 with conventional treatment (16-fold increased risk). A similar 15-fold increase was observed in an independent cohort of 929 patients with MPN. Considering primary myelofibrosis only (N = 216), 3 lymphomas were observed in 31 inhibitor-treated patients (9.7%) vs 1 (0.54%) of 185 control patients. Lymphomas were of aggressive B-cell type, extranodal, or leukemic with high MYC expression in the absence of JAK2 V617F or other MPN-associated mutations. Median time from initiation of inhibitor therapy to lymphoma diagnosis was 25 months. Clonal immunoglobulin gene rearrangements were already detected in the bone marrow during myelofibrosis in 16.3% of patients. Lymphomas occurring during JAK1/2 inhibitor treatment were preceded by a preexisting B-cell clone in all 3 patients tested. Sequencing verified clonal identity in 2 patients. The effects of JAK1/2 inhibition were mirrored in Stat1-/- mice: 16 of 24 mice developed a spontaneous myeloid hyperplasia with the concomitant presence of aberrant B cells. Transplantations of bone marrow from diseased mice unmasked the outgrowth of a malignant B-cell clone evolving into aggressive B-cell leukemia-lymphoma. We conclude that JAK/STAT1 pathway inhibition in myelofibrosis is associated with an elevated frequency of aggressive B-cell lymphomas. Detection of a preexisting B-cell clone may identify individuals at risk.

Diffusion-weighted imaging (DWI) with apparent diffusion coefficient (ADC) mapping as a quantitative imaging biomarker for prediction of immunohistochemical receptor status, proliferation rate, and molecular subtypes of breast cancer.

BACKGROUND: Diffusion-weighted imaging (DWI) with apparent diffusion coefficient (ADC) mapping is one of the most useful additional MRI parameters to improve diagnostic accuracy and is now often used in a multiparameric imaging setting for breast tumor detection and characterization. PURPOSE: To evaluate whether different ADC metrics can also be used for prediction of receptor status, proliferation rate, and molecular subtype in invasive breast cancer. STUDY TYPE: Retrospective. SUBJECTS: In all, 107 patients with invasive breast cancer met the inclusion criteria (mean age 57 years, range 32-87) and underwent multiparametric breast MRI. FIELD STRENGTH/SEQUENCE: 3 T, readout-segmented echo planar imaging (rsEPI) with IR fat suppression, dynamic contrast-enhanced (DCE) T1 -weighted imaging, T2 -weighted turbo-spin echo (TSE) with fatsat. ASSESSMENT: Two readers independently drew a region of interest on ADC maps on the whole tumor (WTu), and on its darkest part (DpTu). Minimum, mean, and maximum ADC values of both WTu and DpTu were compared for receptor status, proliferation rate, and molecular subtypes. STATISTICAL TESTS: Wilcoxon rank sum, Mann-Whitney U-tests for associations between radiologic features and histopathology; histogram and q-q plots, Shapiro-Wilk's test to assess normality, concordance correlation coefficient for precision and accuracy; receiver operating characteristics curve analysis. RESULTS: Estrogen receptor (ER) and progesterone receptor (PR) status had significantly different ADC values for both readers. Maximum WTu (P = 0.0004 and 0.0005) and mean WTu (P = 0.0101 and 0.0136) were significantly lower for ER-positive tumors, while PR-positive tumors had significantly lower maximum WTu values (P = 0.0089 and 0.0047). Maximum WTu ADC was the only metric that was significantly different for molecular subtypes for both readers (P = 0.0100 and 0.0132) and enabled differentiation of luminal tumors from nonluminal (P = 0.0068 and 0.0069) with an area under the curve of 0.685 for both readers. DATA CONCLUSION: Maximum WTu ADC values may be used to differentiate luminal from other molecular subtypes of breast cancer. LEVEL OF EVIDENCE: 3 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2019;50:836-846.

AHR/CYP1A1 interplay triggers lymphatic barrier breaching in breast cancer spheroids by inducing 12(S)-HETE synthesis.

A causal link between overexpression of aryl hydrocarbon receptor (AHR) and its target cytochrome P450 1A1 (CYP1A1) and metastatic outgrowth of various cancer entities has been established. Nevertheless, the mechanism how AHR/CYP1A1 support metastasis formation is still little understood. In vitro we discovered a potential mechanism facilitating tumour dissemination based on the production of 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE). Utilising a three-dimensional lymph endothelial cell (LEC) monolayer & MDA-MB231 breast cancer cell spheroid co-culture model in combination with knock-down approach allowed elucidation of the molecular/biochemical basis of AHR/CYP1A1-induced tumour breaching through the LEC barrier. Enzyme immunoassay evidenced the potential of recombinant CYP1A1 to synthesise 12(S)-HETE in vitro and qPCR and Western blotting measured gene and protein expression in specific experimental settings. In detail, AHR induced CYP1A1 expression and 12(S)-HETE secretion in tumour spheroids, which caused LEC junction retraction thereby forming large discontinuities allowing transmigration of the tumour. This was enforced by the activating AHR ligand 6-formylindolo (3,3-b)carbazole (FICZ), or inhibited by the AHR antagonist 3,3'-diindolylmethane (DIM) as well as by siRNA against AHR and CYP1A1. AHR and NF-κB were negatively cross talking and therefore, the inhibition of AHR (but not CYP1A1) induced RELA, RELB, NFKB1, NFKB2 and the NF-κB target MMP1, which itself promotes tumour intravasation by a mechanism that is different from 12(S)-HETE. Conversely, the inhibition of NFKB2 induced AHR, CYP1A1 and 12(S)-HETE synthesis. The approved clinical drugs guanfacine and vinpocetine, which inhibit CYP1A1 and NF-κB, respectively, significantly inhibited LEC barrier breaching in vitro indicating an option to reduce metastatic dissemination.

Colon cancer cell-derived 12(S)-HETE induces the retraction of cancer-associated fibroblast via MLC2, RHO/ROCK and Ca2+ signalling.

Retraction of mesenchymal stromal cells supports the invasion of colorectal cancer cells (CRC) into the adjacent compartment. CRC-secreted 12(S)-HETE enhances the retraction of cancer-associated fibroblasts (CAFs) and therefore, 12(S)-HETE may enforce invasivity of CRC. Understanding the mechanisms of metastatic CRC is crucial for successful intervention. Therefore, we studied pro-invasive contributions of stromal cells in physiologically relevant three-dimensional in vitro assays consisting of CRC spheroids, CAFs, extracellular matrix and endothelial cells, as well as in reductionist models. In order to elucidate how CAFs support CRC invasion, tumour spheroid-induced CAF retraction and free intracellular Ca2+ levels were measured and pharmacological- or siRNA-based inhibition of selected signalling cascades was performed. CRC spheroids caused the retraction of CAFs, generating entry gates in the adjacent surrogate stroma. The responsible trigger factor 12(S)-HETE provoked a signal, which was transduced by PLC, IP3, free intracellular Ca2+, Ca2+-calmodulin-kinase-II, RHO/ROCK and MYLK which led to the activation of myosin light chain 2, and subsequent CAF mobility. RHO activity was observed downstream as well as upstream of Ca2+ release. Thus, Ca2+ signalling served as central signal amplifier. Treatment with the FDA-approved drugs carbamazepine, cinnarizine, nifedipine and bepridil HCl, which reportedly interfere with cellular calcium availability, inhibited CAF-retraction. The elucidation of signalling pathways and identification of approved inhibitory drugs warrant development of intervention strategies targeting tumour-stroma interaction.

Candidatus Dirofilaria hongkongensis as Causative Agent of Human Ocular Filariosis after Travel to India.

We report a human case of ocular Dirofilaria infection in a traveler returning to Austria from India. Analysis of mitochondrial sequences identified the worm as Candidatus Dirofilaria hongkongensis, a close relative of Dirofilaria repens, which was only recently described in Hong Kong and proposed as a new species.

Expansion of BCR/ABL1+ cells requires PAK2 but not PAK1.

The p21-activated kinases (PAKs) are key nodes in oncogenic signalling pathways controlling growth, survival, and motility of cancer cells. Their activity is increased in many human cancers and is associated with poor prognosis. To date, PAK deregulation has mainly been studied in solid tumours, where PAK1 and PAK4 are the main isoforms deregulated. We show that PAK1 and PAK2 are the critical isoforms in a BCR/ABL1+ haematopoietic malignancy. In suspension, leukaemic cells deficient for PAK1 and PAK2 undergo apoptosis, while the loss of either protein is well tolerated. Transfer of medium conditioned by shPAK2- but not shPAK1-expressing leukaemic cells interferes with endothelial cell growth. We found that leukaemic cells produce exosomes containing PAK2. Transfer of isolated exosomes supports endothelial cell proliferation. In parallel, we found that leukaemic cells explicitly require PAK2 to grow towards an extracellular matrix. PAK2-deficient cells fail to form colonies in methylcellulose and to induce lymphomas in vivo. PAK2 might therefore be the critical isoform in leukaemic cells by controlling tumour growth in a dual manner: vascularization via exosome-mediated transfer to endothelial cells and remodelling of the extracellular matrix. This finding suggests that the PAK2 isoform represents a promising target for the treatment of haematological diseases.

Investigating the prediction value of multiparametric magnetic resonance imaging at 3 T in response to neoadjuvant chemotherapy in breast cancer.

OBJECTIVE: To explore the predictive value of parameters derived from diffusion-weighted imaging (DWI) and contrast-enhanced (CE)-MRI at different time-points during neoadjuvant chemotherapy (NACT) in breast cancer. METHODS: Institutional review board approval and written, informed consent from 42 breast cancer patients were obtained. The patients were investigated before and at three different time-points during neoadjuvant chemotherapy (NACT) using tumour diameter and volume from CE-MRI and ADC values obtained from drawn 2D and segmented 3D regions of interest. Prediction of pathologic complete response (pCR) was evaluated using the area under the curve (AUC) of receiver operating characteristic analysis. RESULTS: There was no significant difference between pathologic complete response and non-pCR in baseline size measures (p > 0.39). Diameter change was significantly different in pCR (p < 0.02) before the mid-therapy point. The best predictor was lesion diameter change observed before mid-therapy (AUC = 0.93). Segmented volume was not able to differentiate between pCR and non-pCR at any time-point. The ADC values from 3D-ROI were not significantly different from 2D data (p = 0.06). The best AUC (0.79) for pCR prediction using DWI was median ADC measured before mid-therapy of NACT. CONCLUSIONS: The results of this study should be considered in NACT monitoring planning, especially in MRI protocol designing and time point selection. KEY POINTS: • Mid-therapy diameter changes are the best predictors of pCR in neoadjuvant chemotherapy. • Volumetric measures are not strictly superior in therapy monitoring to lesion diameter. • Size measures perform as a better predictor than ADC values.

Diffusion-weighted imaging of breast lesions: Region-of-interest placement and different ADC parameters influence apparent diffusion coefficient values.

OBJECTIVES: To investigate the influence of region-of-interest (ROI) placement and different apparent diffusion coefficient (ADC) parameters on ADC values, diagnostic performance, reproducibility and measurement time in breast tumours. METHODS: In this IRB-approved, retrospective study, 149 histopathologically proven breast tumours (109 malignant, 40 benign) in 147 women (mean age 53.2) were investigated. Three radiologists independently measured minimum, mean and maximum ADC, each using three ROI placement approaches:1 - small 2D-ROI, 2 - large 2D-ROI and 3 - 3D-ROI covering the whole lesion. One reader performed all measurements twice. Median ADC values, diagnostic performance, reproducibility, and measurement time were calculated and compared between all combinations of ROI placement approaches and ADC parameters. RESULTS: Median ADC values differed significantly between the ROI placement approaches (p < .001). Minimum ADC showed the best diagnostic performance (AUC .928-.956), followed by mean ADC obtained from 2D ROIs (.926-.94). Minimum and mean ADC showed high intra- (ICC .85-.94) and inter-reader reproducibility (ICC .74-.94). Median measurement time was significantly shorter for the 2D ROIs (p < .001). CONCLUSIONS: ROI placement significantly influences ADC values measured in breast tumours. Minimum and mean ADC acquired from 2D-ROIs are useful for the differentiation of benign and malignant breast lesions, and are highly reproducible, with rapid measurement. KEY POINTS: • Region of interest placement significantly influences apparent diffusion coefficient of breast tumours. • Minimum and mean apparent diffusion coefficient perform best and are reproducible. • 2D regions of interest perform best and provide rapid measurement times.

Standardized evaluation of tumor-infiltrating lymphocytes in breast cancer: results of the ring studies of the international immuno-oncology biomarker working group.

Multiple independent studies have shown that tumor-infiltrating lymphocytes (TIL) are prognostic in breast cancer with potential relevance for response to immune-checkpoint inhibitor therapy. Although many groups are currently evaluating TIL, there is no standardized system for diagnostic applications. This study reports the results of two ring studies investigating TIL conducted by the International Working Group on Immuno-oncology Biomarkers. The study aim was to determine the intraclass correlation coefficient (ICC) for evaluation of TIL by different pathologists. A total of 120 slides were evaluated by a large group of pathologists with a web-based system in ring study 1 and a more advanced software-system in ring study 2 that included an integrated feedback with standardized reference images. The predefined aim for successful ring studies 1 and 2 was an ICC above 0.7 (lower limit of 95% confidence interval (CI)). In ring study 1 the prespecified endpoint was not reached (ICC: 0.70; 95% CI: 0.62-0.78). On the basis of an analysis of sources of variation, we developed a more advanced digital image evaluation system for ring study 2, which improved the ICC to 0.89 (95% CI: 0.85-0.92). The Fleiss' kappa value for <60 vs ≥60% TIL improved from 0.45 (ring study 1) to 0.63 in RS2 and the mean concordance improved from 88 to 92%. This large international standardization project shows that reproducible evaluation of TIL is feasible in breast cancer. This opens the way for standardized reporting of tumor immunological parameters in clinical studies and diagnostic practice. The software-guided image evaluation approach used in ring study 2 may be of value as a tool for evaluation of TIL in clinical trials and diagnostic practice. The experience gained from this approach might be applicable to the standardization of other diagnostic parameters in histopathology.

Bilateral diffusion-weighted MR imaging of breast tumors with submillimeter resolution using readout-segmented echo-planar imaging at 7 T.

PURPOSE: To evaluate the image quality, robustness, and diagnostic performance of submillimeter in-plane resolution diffusion-weighted ( DW diffusion-weighted ) magnetic resonance (MR) imaging at 7 T in the assessment of breast tumors. MATERIALS AND METHODS: Institutional review board approval and written informed consent of five volunteers and 33 patients with 33 breast lesions (31 with histopathologic confirmation, two with confirmation at follow-up) were obtained. Image quality optimization and comparisons of readout-segmented echo-planar imaging ( rs-EPI readout-segmented echo-planar imaging ) and single-shot echo-planar imaging ( ss-EPI single-shot echo-planar imaging ) with or without parallel imaging were performed in volunteers. In all patients, bilateral DW diffusion-weighted imaging was performed in 3 minutes 35 seconds by using combined rs-EPI readout-segmented echo-planar imaging and parallel imaging with 0.9 × 0.9 mm in-plane resolution with a 7-T whole-body MR imager. Image quality, lesion conspicuity, and image properties (ie, signal-to-noise ratio, contrast-to-noise ratio) were assessed. Regions of interest were drawn in the largest lesion in each patient (23 malignant lesions, 10 benign lesions) by two independent readers. Apparent diffusion coefficient ( ADC apparent diffusion coefficient ) values were used to differentiate between benign and malignant breast tumors. RESULTS: DW diffusion-weighted imaging with combined parallel imaging and rs-EPI readout-segmented echo-planar imaging reduced artifacts (ie, blurring and geometric distortions) by a calculated factor of seven when compared with DW diffusion-weighted imaging with ss-EPI single-shot echo-planar imaging , and it improved image quality from a score of 1 of 10 to a score of 8 of 10. The rs-EPI readout-segmented echo-planar imaging sequence with a b value of 0 sec/mm(2) yielded high-spatial-resolution T2-weighted MR images. An ADC apparent diffusion coefficient threshold of 1.275 × 10(-3) mm(2)/sec enabled differentiation between benign and malignant breast lesions, with sensitivity and specificity of 96% and 100%, respectively, for both independent readers. CONCLUSION: At 7 T, one DW diffusion-weighted imaging examination of less than 4 minutes yielded high-quality ADC apparent diffusion coefficient maps and high-spatial-resolution T2-weighted MR images that were used to assess tumor and breast morphology. ADC apparent diffusion coefficient quantification alone enabled excellent differentiation of benign and malignant breast lesions.

Multiparametric MR Imaging with High-Resolution Dynamic Contrast-enhanced and Diffusion-weighted Imaging at 7 T Improves the Assessment of Breast Tumors: A Feasibility Study.

PURPOSE: To ascertain whether multiparametric magnetic resonance (MR) imaging of the breast in combination with dynamic contrast material-enhanced (DCE) imaging and diffusion-weighted imaging (DWI) at 7 T is feasible and improves diagnostic accuracy. MATERIALS AND METHODS: From December 2011 to December 2013, 40 patients with suspicious breast lesions were included in this institutional review board-approved prospective study. Before bilateral multiparametric MR imaging of the breast at 7 T, all patients gave written informed consent. Lesions were classified according to Breast Imaging Reporting and Data System (BI-RADS) and assessed for apparent diffusion coefficient (ADC) values by two readers independently. For combined analysis of DCE MR imaging and DWI, the BI-RADS-adapted reading algorithm, which adapted ADC thresholds to the BI-RADS assessment category, was used. Diagnostic values of multiparametric, DCE MR imaging, and DWI were calculated. Receiver operating characteristic curve analysis was performed. Image quality and interreader agreement were assessed. Histopathologic results were used as the highest standard. RESULTS: There were 29 malignant and 17 benign lesions (range, 6-95 mm; mean, 23.3 mm). Multiparametric MR imaging yielded a sensitivity of 100% (29 of 29 lesions), a specificity of 88.2% (16 of 18 lesions), and an area under the curve of 0.941, which was greater than for DCE MR imaging (P = .003), which had a sensitivity of 100% (29 of 29 lesions), a specificity of 53.2% (nine of 17 lesions), and an area under the curve of 0.765. DWI had a sensitivity of 93.1% (27 of 29 lesions), a specificity of 88.2% (15 of 17 lesions), and an area under the curve of 0.907. Multiparametric MR imaging at 7 T of the breast eliminated all false-negative findings and reduced false-positive findings, from eight false-positive findings with DCE MR imaging to two false-positive findings. Thus, if used clinically, 7-T multiparametric MR imaging may have potentially obviated unnecessary breast biopsies in six of eight lesions (P = .031). Multiparametric MR imaging demonstrated either excellent or good image quality and interreader agreement (κ = 0.89-1.00). CONCLUSION: The clinical use of 7-T multiparametric MR imaging is feasible, provides good or excellent image quality, and has the potential to improve diagnostic accuracy.

Histopathological characterization of ulcerated breast cancer and comparison to their non-ulcerated counterparts.

Approximately 6-15 % of breast cancer patients are diagnosed with primary ulcerated breast cancer (ULBC). ULBC is known to be associated with short recurrence free and poor overall survival. Therefore, the purpose of this study was to characterize ULBC and compare the histopathological findings with those of non-ulcerative breast cancer (NULBC). A total of 152 ULBCs were evaluated and compared to 304 consecutive non-ulcerated, age-matched breast malignancies. Patients mean age was 65 years (SD = 13.0 ULBC, SD = 14.0 NULBC). ULBC was associated with a higher rate of poorly differentiated tumors (p = < 0.001), as well as larger tumor sizes (p = < 0.001). As expected, the rate of axillary lymph node involvement was higher in ULBC patients (p = <0.001). In addition to that, ULBC was associated with a higher rate of triple negative breast cancer (p = 0.002), and higher Ki67 expression (p = < 0.001). ULBC showed more aggressive histopathological features in comparison to NULBC which may contribute to the generally known poorer prognosis of women with ULBC.

Clinical applications of next-generation sequencing-based ctDNA analyses in breast cancer: defining treatment targets and dynamic changes during disease progression.

The advancements in the detection and characterization of circulating tumor DNA (ctDNA) have revolutionized precision medicine and are likely to transform standard clinical practice. The non-invasive nature of this approach allows for molecular profiling of the entire tumor entity, while also enabling real-time monitoring of the effectiveness of cancer therapies as well as the identification of resistance mechanisms to guide targeted therapy. Although the field of ctDNA studies offers a wide range of applications, including in early disease, in this review we mainly focus on the role of ctDNA in the dynamic molecular characterization of unresectable locally advanced and metastatic BC (mBC). Here, we provide clinical practice guidance for the rapidly evolving field of molecular profiling of mBC, outlining the current landscape of liquid biopsy applications and how to choose the right ctDNA assay. Additionally, we underline the importance of exploring the clinical relevance of novel molecular alterations that potentially represent therapeutic targets in mBC, along with mutations where targeted therapy is already approved. Finally, we present a potential roadmap for integrating ctDNA analysis into clinical practice.

Impact of HER-2-targeted therapy on overall survival in patients with HER-2 positive metastatic breast cancer.

Upon disease progression on trastuzumab-based therapy, patients with HER-2 positive metastatic breast cancer (MBC) may switch to lapatinib or continue on trastuzumab. We aimed to assess the impact of both strategies on overall survival (OS) in all patients treated for HER-2 positive MBC at the Medical University Vienna from 1999 until 2009. A total of 201 patients were identified from a breast cancer data base. Of these 115 (57.2%) received multiple lines of trastuzumab-based therapy, whereas 58 (28.9%) were treated with a single line. A control group of 28 patients (13.9%) had never received trastuzumab as they were treated before 1999, when trastuzumab was registered. OS from diagnosis of metastatic disease was defined as primary study endpoint. Trastuzumab significantly prolonged OS in HER-2 positive MBC (41 versus 13 months; p < 0.001). Administration of multiple lines further improved OS; this, however, did not reach statistical significance (47 versus 28 months; p = 0.069). Positive estrogen receptor (ER) status (HR 1.6; 95% CI 1.13-2.27) was associated with better outcome compared to negative estrogen receptor status (p = 0.02). Addition of lapatinib did not improve OS significantly in patients with prior trastuzumab-based therapy (62 versus 47 months; p = n.s.). Patients receiving lapatinib after diagnosis of BM, however, experienced an improvement of OS (22 versus 5 months; p = 0.022). Trastuzumab improves OS in patients with HER-2 positive MBC with further nonsignificant improvement when administered in multiple lines. Lapatinib did not further improve OS in the entire population; however, lapatinib might improve OS in patients with BM.

Successful delayed interval delivery of a triplet pregnancy using conservative management.

Preterm birth is a significant cause of perinatal morbidity and mortality, especially in multiple pregnancies. Delayed interval delivery can prolong pregnancy for the remaining fetus(es) in an imminent stillbirth or extremely preterm birth of the first fetus, improving the lastborn's outcomes. We present a case of delayed interval delivery of a triplet pregnancy following preterm prelabour rupture of membranes and progressive cervical insufficiency. Following vaginal delivery of the first fetus at 24+1 gestational weeks, the patient received antibiotics and tocolysis. Cerclage was not conducted as the mother had a vaginal infection. A 15-day delivery interval for the second and third fetuses was achieved. The firstborn required mechanical ventilation and inotropic support, while the others only required continuous positive airway pressure. There is no consensus on the best way to perform delayed interval delivery. We achieved a complications-free interval of 15 days with conservative management in a triplet pregnancy.

Differential immunomodulatory effects of epirubicin/cyclophosphamide and docetaxel in breast cancer patients.

BACKGROUND: Epirubicin/cyclophosphamide (EC) and docetaxel (D) are commonly used in a sequential regimen in the neoadjuvant treatment of early, high-risk or locally advanced breast cancer (BC). Novel approaches to increase the response rate combine this treatment with immunotherapies such as PD-1 inhibition. However, the expected stimulatory effect on lymphocytes may depend on the chemotherapy backbone. Therefore, we separately compared the immunomodulatory effects of EC and D in the setting of a randomized clinical trial. METHODS: Tumor and blood samples of 154 patients from the ABCSG-34 trial were available (76 patients received four cycles of EC followed by four cycles of D; 78 patients get the reverse treatment sequence). Tumor-infiltrating lymphocytes, circulating lymphocytes and 14 soluble immune mediators were determined at baseline and at drug change. Furthermore, six BC cell lines were treated with E, C or D and co-cultured with immune cells. RESULTS: Initial treatment with four cycles of EC reduced circulating B and T cells by 94% and 45%, respectively. In contrast, no comparable effects on lymphocytes were observed in patients treated with initial four cycles of D. Most immune mediators decreased under EC whereas D-treatment resulted in elevated levels of CXCL10, urokinase-type plasminogen activator (uPA) and its soluble receptor (suPAR). Accordingly, only the exposure of BC cell lines to D induced similar increases as compared to E. While treatment of BC cells with E was associated with cell shrinkage and apoptosis, D induced cell swelling and accumulation of cells in G2 phase. CONCLUSION: The deleterious effect of EC on lymphocytes indicates strong immunosuppressive properties of this combination therapy. D, in contrast, has no effect on lymphocytes, but triggers the secretion of stimulatory proteins in vivo and in vitro, indicating a supportive effect on the immune system. Underlying differences in the induced cell death might be causal. These divergent immunomodulatory effects of epirubicin/cyclophosphamide and docetaxel should be considered when planning future combinations with immunotherapies in breast cancer.

Emergence of immune-related adverse events correlates with pathological complete response in patients receiving pembrolizumab for early triple-negative breast cancer.

Based upon results of the KEYNOTE-522 trial and following approval by regulatory authorities, the addition of pembrolizumab to chemotherapy is now the standard-of-care for the treatment of early triple-negative breast cancer (eTNBC) (Clinical stage II-III). Pembrolizumab is a programmed cell death protein 1 monoclonal antibody, known to cause immune-related adverse events (irAEs) in a significant subset of patients. Real-world data on incidence, type and treatment strategies of irAEs in the setting of eTNBC treatment are sparse. In this multicenterretrospective analysis, we characterized real-world incidence of irAEs and treatment outcomes such as pathological complete response (pCR) from the combination of pembrolizumab and chemotherapy as neoadjuvant treatment for eTNBC. We found a rate of irAEs of all grades of 63.9% and of 20% for irAEs of grade 3 or higher. In the overall population, a pCR rate of 57.1% was observed. The emergence of irAEs correlated significantly with pCR (72.2% versus 30.8%; p =.03). Discontinuation of neoadjuvant chemotherapy before week 12 correlated significantly with a lower pCR rate. To our knowledge, this is the first study evaluating the real-world efficacy and safety of a neoadjuvant combination of chemotherapy and pembrolizumab in eTNBC, demonstrating a significant correlation between irAEs and pCR. Early discontinuation of neoadjuvant therapy due to AEs resulted in a lower pCR rate.