[Is it possible to detect surface antigen CD133 on patient-derived glioblastoma continuous cell cultures using fluorescent aptamers?] / Vozmozhna li detektsiya poverkhnostnogo antigena CD133 na perevivaemykh kul'turakh kletok glioblastomy patsientov s pomoshch'yu fluorestsentnykh aptamerov?

Theranostics combines diagnostics and therapeutic exposure. Regarding glioblastomas, theranostics solves the problem of detecting and destroying tumor stem cells resistant to irradiation and chemotherapy and causing tumor recurrence. Transmembrane surface antigen CD133 is considered as a potential marker of tumor stem cells. OBJECTIVE: To detect CD133 in patient-derived glioblastoma continuous cell cultures using fluorescence microscopy and modified aptamers (molecular recognition elements) anti-CD133. MATERIAL AND METHODS: To detect CD133, we used mousey fluorescence monoclonal antibodies anti-CD133 MA1-219, FAM-modified DNA aptamers anti-CD133 AP-1-M and Cs5. Non-aptamer DNA oligonucleotide NADO was used as a negative control. Detection was performed for three samples of patient-derived glioblastoma continuous cell cultures coded as 1548, 1721 and 1793. RESULTS: MA1-219 antibodies brightly stained cell culture 1548, to a lesser extent - 1721. There was diffuse staining of cell culture 1793. Cs5-FAM aptamer stained cells in a similar way, but much weaker. AP-1-M-FAM aptamer interacted with cells even weaker and diffusely stained only cell culture 1793. Non-aptamer NADO did not stain cell culture 1548 and very weakly diffusely stained cell culture 1793. CONCLUSION: For both molecular recognition elements (MA1-219 antibody and Cs5 aptamer), 3 cell culture samples can be arranged in the following order possibly reflecting CD133 status decrease: strong signal for cell culture 1548, much weaker for 1721, even weaker for 1793. Only cell culture 1548 can be considered CD133 positive with combination of Cs5+ and NADO signals. Cell culture 1793 is CD133 false positive with combination of Cs5+ and NADO+ signals.

[Thrombosis of the inferior vena cava and right portions of the heart in a relapsing retroperitoneal malignant tumour]. / Tromboz nizhnei poloi veny i pravykh otdelov serdtsa pri retsidivnoi zabriushinnoi zlokachestvennoi opukholi.

Presented herein is a clinical case report regarding successful treatment of a female patient with a relapsing malignant retroperitoneal tumour complicated by disseminated thrombosis of the inferior vena cava, right atrium and right ventricle of the heart. A relapse of the malignant uterine tumour had developed 3 years after the primary operation and was represented by a large-size mass ingrowing into the infrarenal segment of the inferior vena cava, aorta, as well as jejunum. Additional examination revealed the presence of a tumorous thrombus extending from the primary tumorous node in the lumen of the inferior vena cava to the right atrium and ventricle. The procedures performed consisted in removal of the tumour with resection of the inferior vena cava, aorta, and jejunum, followed by thrombectomy from the right portions of the heart under extracorporeal circulation. The postoperative period turned out uneventful, with no complications observed. The woman was discharged on POD 15. Twelve-month postoperative follow up revealed neither relapse nor progression of the disease. Currently, the patient continues undergoing specific treatment (second-line chemotherapy). Also discussed in the article are current challenges concerning both the classification of tumour thrombosis of the inferior vena cava in retroperitoneal sarcomas, and the choice of optimal strategy and policy of treatment.

Trifluridine/tipiracil plus bevacizumab in patients with untreated metastatic colorectal cancer ineligible for intensive therapy: the randomized TASCO1 study.

BACKGROUND: We designed an open-label, noncomparative phase II study to assess the safety and efficacy of first-line treatment with trifluridine/tipiracil plus bevacizumab (TT-B) and capecitabine plus bevacizumab (C-B) in untreated patients with unresectable metastatic colorectal cancer (mCRC) who were not candidates for combination with cytotoxic chemotherapies. PATIENTS AND METHODS: From 29 April 2016 to 29 March 2017, 153 patients were randomly assigned (1:1) to either TT-B (N = 77) or C-B (N = 76). The primary end point was progression-free survival (PFS). The primary PFS analysis was performed after 100 events (radiological progression or death) were observed. Secondary end points included overall survival (OS), quality of life (QoL; QLQ-C30 and QLQ-CR29 questionnaires), and safety. RESULTS: Median (range) duration of treatment was 7.8 (6.0-9.7) months and 6.2 (4.1-9.1) months in the TT-B and C-B groups, respectively. Median (range) PFS was 9.2 (7.6-11.6) and 7.8 (5.5-10.1) months, respectively. Median (range) OS was 18 (15.2 to NA) and 16.2 (12.5 to NA) months, respectively. QoL questionnaires showed no relevant changes over time for either treatment. Therapies were well tolerated. Patients receiving TT-B had more grade ≥3 neutropenia (47% versus 5% with C-B). Patients receiving C-B had more grade ≥3 hand-foot syndrome (12% versus 0% with TT-B) and grade ≥3 diarrhea (8% versus 1% with TT-B), consistent with the known safety profiles of these agents. CONCLUSION: TT-B treatment showed promising clinical activity in untreated patients with unresectable mCRC ineligible for intensive therapy, with an acceptable safety profile and no clinically relevant changes in QoL. CLINICAL TRIAL INFORMATION: NCT02743221 (ClinicalTrials.gov).

Stereotactic image-guided neoadjuvant ablative single-dose radiation, then lumpectomy, for early breast cancer: the SIGNAL prospective single-arm trial of single-dose radiation therapy.

Background and Purpose: Adjuvant whole-breast irradiation after breast-conserving surgery, typically delivered over several weeks, is the traditional standard of care for low-risk breast cancer. More recently, hypofractionated, partial-breast irradiation has increasingly become established. Neoadjuvant single-fraction radiotherapy (rt) is an uncommon approach wherein the unresected lesion is irradiated preoperatively in a single fraction. We developed the signal (Stereotactic Image-Guided Neoadjuvant Ablative Radiation Then Lumpectomy) trial, a prospective single-arm trial to test our hypothesis that, for low-risk carcinoma of the breast, the preoperative single-fraction approach would be feasible and safe. Methods: Patients presenting with early-stage (T < 3 cm), estrogen-positive, clinically node-negative invasive carcinoma of the breast with tumours at least 2 cm away from skin and chest wall were enrolled. All patients received prone breast magnetic resonance imaging (mri) and prone computed tomography simulation. Treatable patients received a single 21 Gy fraction of external-beam rt (as volumetric-modulated arc therapy) to the primary lesion in the breast, followed by definitive surgery 1 week later. The primary endpoints at 3 weeks, 6 months, and 1 year were toxicity and cosmesis (that is, safety) and feasibility (defined as the proportion of mri-appropriate patients receiving rt). Results: Of 52 patients accrued, 27 were successfully treated. The initial dosimetric constraints resulted in a feasibility failure, because only 57% of eligible patients were successfully treated. Revised dosimetric constraints were developed, after which 100% of patients meeting mri criteria were treated according to protocol. At 3 weeks, 6 months, and 1 year after the operation, toxicity, patient- and physician-rated cosmesis, and quality of life were not significantly different from baseline. Conclusions: The signal trial presents a feasible method of implementing single-dose preoperative rt in early-stage breast cancer. This pilot study did not identify any significant toxicity and demonstrated excellent cosmetic and quality-of-life outcomes. Future randomized multi-arm studies are required to corroborate these findings.

[Multiple sclerosis vaccine: possibilities of the use of dendritic cells immunomodulatory potential].

For the first time we carried out a clinical assessment of the safety, tolerability and clinical efficacy course of repeated administration of experimental modified autologous vaccine interleykin (IL-10) dendritic cells in two patients with secondary-progressive multiple sclerosis patient and one with relapsing-remitting multiple sclerosis. In the course of treatment, we carried out clinical and immunological monitoring. It was found out that intradermal dose of 3 x 106 cells applied to spinal area 6-12. times did not cause any serious side effects. After the treatment with dendritic cells, the following results were observed: 1) a significant positive clinical effect in patients with secondary-progressive multiple sclerosis exacerbations; 2) moderate positive clinical effect in patients with relapsing-remitting multiple sclerosis, in a state of remission; 3) a complete absence of any clinical results in patients with secondary-progressive multiple sclerosis without exacerbations. The immune response was characterized by a significant absolute and relative increase of serum T-regulatory cells. Discovered distinct anti-inflammatory properties of dendritic cell therapy allow us to consider it as a promising area of personalized treatment based on an individual vaccination against multiple sclerosis.

ARTERIAL CHEMOINFUSION IN PATIENTS WITH LOCALLY ADVANCED AND METASTATIC PANCREAS CANCER.

The chemoinfusions (310) were carried out in celiac trunk in 167 patients with non-removed pancreas cancer at the period from 2000 to 2015. Locally advanced timorous process (stage III, n=79) was revealed in 79 patients and liver metastases (stage IV, n=88) were noted in 88 cases. The celiac axis infusion by Gemcitabine (1000 mg/m²) was applied for patients and GEMOX (Gemcitabine+Oxaliplatin 75 mg/m²) has been using since 2012. Symptomatic improvement such as decrease of pain, growth of body weight was noted in majority of patients. An average lifetime, median and one-year survival consisted of 7,6 months, 5,8 months and 10%. The patients (133) were treated by 1­2 cycles and after that by course of total body chemotherapeutics. There weren't any serious complications. Toxic manifestations of chemotherapy weren't higher than I­II degree and they were arrested by corrective therapy in 92 patients (55%). The celiac axis infusion is safe in patients with locally advanced and inoperable pancreas cancer. Symptomatic improvement showed the most patients. The objective response to the treatment had 20% patients and performance of repeated cycles led to increase of their survival.

[Experience with the use of gefitinib in patients with inoperable non-squamous cell lung cancer with activating EGFR mutations].

Discovery of activating EGFR mutations led to dramatic modification of treatment schemes for nonsquamous lung cancer. 70 patients with activating EGFR mutations were treated by gefitinib being either a part of prospective phase II trial (n = 25) or, subsequently, subjected to routine clinical management (n = 45). Objective response rate approached to 32.7%. Median time to disease progression was 14 months, and median overall survival was 26.1 months. Subgroup analysis revealed statistically longer time to disease progression (p < 0,0001) and overall survival (p = 0,001) in latter vs. former group, despite the lower rate of objective response (22% vs 48%). Possible explanations include more relaxed standards for routine gefitinib use, i.e. inclusion of the patients with non-measurable tumor lumps, continuation of gefitinib uptake upon slow disease progression, and increasing availability and quality of radiosurgery for brain metastases.

A follow-up cytogenetic study of workers highly exposed inside the Chernobyl sarcophagus.

Data are presented on some of the engineers and scientists who regularly entered highly radioactive areas of the sarcophagus chamber constructed around the ruins of the Chernobyl reactor. Previous investigations on six of them by unstable chromosomal aberrations, quartz fibre electrometers and, in some cases, tooth electron spin resonance have all indicated high doses accumulated over several years of work inside the sarcophagus. Here, the authors present the data on eleven of the men who agreed to be monitored cytogenetically although two have since died aged 45 and 66 y. The present data were combined with the previous to examine the time-courses of these individuals' changes in their aberration frequencies. As expected, dicentric aberrations showed a clear drop down to 2-3 per 100 cells since the men ceased working inside the sarcophagus. In contrast, the translocation yields remained at a high level showing no tendency to decrease and so proved reliable for retrospective biodosimetry. These data are particularly useful in demonstrating the value of FISH long after high but protracted and fractionated exposure.

Monte Carlo calculation of dose distributions in oligometastatic patients planned for spine stereotactic ablative radiotherapy.

Dosimetric consequences of plans optimized using the analytical anisotropic algorithm (AAA) implemented in the Varian Eclipse treatment planning system for spine stereotactic body radiotherapy were evaluated by re-calculating with BEAMnrc/DOSXYZnrc Monte Carlo. Six patients with spinal vertebral metastases were planned using volumetric modulated arc therapy. The planning goal was to cover at least 80% of the planning target volume with a prescribed dose of 35 Gy in five fractions. Tissue heterogeneity-corrected AAA dose distributions for the planning target volume and spinal canal planning organ-at-risk volume were compared against those obtained from Monte Carlo. The results showed that the AAA overestimated planning target volume coverage with the prescribed dose by up to 13.5% (mean 8.3% +/- 3.2%) when compared to Monte Carlo simulations. Maximum dose to spinal canal planning organ-at-risk volume calculated with Monte Carlo was consistently smaller than calculated with the treatment planning system and remained under spinal cord dose tolerance. Differences in dose distribution appear to be related to the dosimetric effects of accounting for body composition in Monte Carlo simulations. In contrast, the treatment planning system assumes that all tissues are water-equivalent in their composition and only differ in their electron density.

Stereotactic body radiotherapy: volumetric modulated arc therapy versus 3D non-coplanar conformal radiotherapy for the treatment of early stage lung cancer.

Stereotactic body radiotherapy (SBRT) is a treatment option for patients with early stage lung cancer. Treatment duration can be >30 minutes per fraction with non-coplanar 3D-conformal radiotherapy (3D-CRT). Whilst this is generally well tolerated, faster delivery techniques are desirable. Volumetric modulated arc therapy (VMAT) allows for fast delivery of radiation treatment. The purpose of this planning study was to compare SBRT with 3D-CRT and VMAT, with VMAT plans generated using both single arc and 3 non-coplanar partial arcs. Ten patients who previously underwent SBRT (48 Gy in 4 fractions) with 3D-CRT were selected. VMAT plans were generated to treat the PTV while limiting doses to organs at risk. Cumulative dose volume histogram (DVH) parameters were compared between the 3 techniques using the Wilcoxon matched pairs test. Treatment delivery time was also assessed. Both VMAT techniques covered target volumes more conformally than 3D-CRT with a mean V48/VPTV of 1.21 for 3D-CRT, 1.03 for 3 arc plans and 1.01 for single arc plans (p = 0.005). Dose constraints to organs at risk were met using all three techniques. Mean lung doses were 2.93 Gy for 3D-CRT, 2.87 Gy for single arc and 2.73 Gy for the 3 arc technique (3-arc vs. 3D-CRT: p = 0.009). Lung V20 for 3D-CRT, 1 arc and 3 arcs were 3.24%, 2.89% and 2.73%, respectively (3 arc vs. 3D-CRT: p = 0.028). Mean time to deliver a single fraction was 13 minutes for 3D-CRT, 9.2 minutes for 3 arcs and 5.5 minutes for 1 arc. VMAT resulted in improved conformality compared to 3D-CRT. The 3 arc technique appears to have the lowest dose to lung although the magnitude is unlikely to be clinically significant. The main advantage of VMAT over 3D-CRT is faster treatment delivery time. Shortened treatment times are anticipated to improve tolerability of this treatment and reduce the chance of error due to intra-fraction motion.

Radiobiological effects of altering dose rate in filter-free photon beams.

To validate that altering radiotherapy dose rate through either changing pulse repetition frequency or instantaneous dose rate does not have an effect on cell survival, two human carcinoma and a hamster lung cell line were irradiated with various beam settings. Varian TrueBeam linac with a flattening filter free mode of operation was used for all experiments. The results obtained indicate that either method of changing dose rate has no effect on cell survival in the three cell lines studied. Filtered and filter free modes were also compared in treatments with protracted dose delivery which significantly increases overall treatment time. Cell survival indicated no difference between filter and filter free beam delivery in any of the protraction schemes. An increase in survival was seen in both modes upon protracting dose delivery to 15, 30 or 60 min rather than delivering acutely. Further, analysis of induced DNA double-strand breaks via the γH2AX assay showed no difference between filtered and unfiltered beams. The following study suggests that increasing dose rate is an acceptable manner of decreasing radiotherapy treatment time that does not have any detrimental effects on in vitro cell eradication.

[Pharmacoeconomic analysis of gefitinib therapy in patients with non-cell cell lung cancer].

We performed a treatment efficacy analysis for non-small cell lung cancer (NSCLC) patients' population with EGFR mutation aimed at optimization of pharmacoeconomic factors. The employment of gefitinib leads to an increase in patients' life expectancy for a median of 1.05 years. The average cost-effectiveness of this therapy is 934.8 thousand rubles per additional year (903.9-1100.5 thousand rubles for each year). If gefitinib therapy is given only to patients with proved EGFR mutation it can decrease the average expenses by 211.6-251.8 thousand rubles per patient in comparison to undiagnosed patients's population receiving gefitinib without a decrease in clinical effect. Comparison of selective gefitinib administration with isolated chemotherapy (CT) yields an incremental cost-effectiveness ratio of 960.7 to 1010.0 thousand rubles per additional year. Therefore, the strategy of EGFR gene mutations testing in patients with inoperable NSCLC with consequent gefitinib therapy administration in patients positive for mutation lead to an increase in life expectancy and is characterized by acceptable cost-effectiveness.

[Phase II clinical trial of autologous dendritic cell vaccine with immunologic adjuvant in cutaneous melanoma patients].

This paper describes the clinical results and immunologic changes in cutaneous melanoma patients receiving active specific immunotherapy with autologous dendritic cell vaccine (DCV) in combination with cyclophosphamide used as immunologic adjuvant. Twenty eight patients with morphologically verified stage III-IV cutaneous melanoma receiving therapy in N. N. Petrov Research Institute of Oncology between 2008 and 2011 were included in the study. All patients signed an informed consent form. Nineteen patients (67,9%) received DCV in therapeutic setting, 9 (32,1%) received it in adjuvant setting. DCV therapy was well tolerated. No serious adverse events were registered. Frequent adverse events included Grade 1-2 unspecific symptoms (fever, fatigue, flu-like symptoms) observed in 22% patients after 3,5% of vaccinations. In therapeutic settings the use DCV lead to clinical effect (PR+SD) in 36,6% of patients. PR was observed in 5% of (95% CI 0-15%) patients, SD in 31,6% (95% CI 13-56%). Duration of the objective responses was 168-965+days. Addition of immunologic adjuvant (cyclophosphamide 300 mg/m2 IV 2 hours) 3 days before vaccination increased its efficacy. In this patients group (n=12) the therapy lead to clinical benefit in 42% (95% CI 17-69%) of cases, median time to progression was 91 (95% CI 55-126) days. This regimen was selected for adjuvant therapy. In the adjuvant therapy group (n=9) the median time to progression was 112 (95% CI 58-166) days. Immunologic monitoring showed correlation ofT- and B-cell immune response with DCV clinical efficacy (p<0,05), no correlation with delayed hypersensivity reaction was observed (p>0,1). DCV is well tolerated and shows immunological and clinical response in stage III-IV skin melanoma patients.

SU-E-T-01: Applications of 6MV FFF Photon Beams in Optimizing Radiobiological Response for Respiratory-Gated Liver SBRT.

PURPOSE: To establish a radiobiological basis for gated stereotactic body radiotherapy of primary and metastatic liver cancers using volumetric arc radiotherapy in a flattening filter free (FFF) mode. METHODS: Human cervical carcinoma, SiHa, non-small cell lung carcinoma, H460, and Chinese hamster V79 cells were irradiated in a water bath with 6MV photons from a Varian TrueBeam linear accelerator. To establish dose-response and its sensitivity to dose rate following acute irradiation, doses of 2, 4, 6, 8 and 10 Gy were delivered in FFF mode at 400 and 1200 MU/min. To investigate whether removal of the flattening filter affects cell response, doses of 5 and 10 Gy were delivered to SiHa and H460 cells in FFF and filtered modes at 400 MU/min. Finally, to assess the effect of protracting dose delivery by gating, a dose of 10 Gy was delivered to SiHa and H460 cells acutely and also over 15, 30 and 60 min. RESULTS: Dose-response over doses examined was independent of dose rate in FFF mode. Differences in cell survival following irradiation in FFF and filtered modes were not significant. However a significant increase in survival for both H460 and SiHa cells was observed for 15 min split-dose irradiation compared to acute irradiation but further increase in irradiation time to 60 min did not affect cell survival. CONCLUSIONS: Dose rate and presence of a flattening filter showed no effect on cell survival, however, survival was significantly affected when dose delivery time was protracted to that typical of conformal field therapy. Volumetric arc based gated SBRT may be beneficial for tumor cell kill, though the gating window and duty cycle have to be balanced against the effect of dose delivery protraction. Research Support (Varian Medical Systems).

SU-E-T-642: Patient Stratification Based on Improvements in Conformity Indices and Normal Tissue Sparing with IMRT in Arteriovenous Malformation Treatments.

PURPOSE: Stereotactic radiosurgery (SRS) is a treatment of choice for Arteriovenous Malformations (AVMs) in anatomical locations associated with a risk of surgical complication. The aim of this study was: 1) to determine which AVM patients benefit from IMRT treatment over 3D-Conformal Treatment (3DCRT) based on PTV size, shape and normal tissue (NT) dose constraints imposed upon IMRT plans; 2) to assess optimized dose distributions against prospectively collected data for symptomatic radiation injury following SRS. METHODS: 31 patients previously treated for AVMs with 3DCRT were replanned with static gantry IMRT. PTV volumes, defined as AVM+1mm margin, ranged from <1cc to >25cc. IMRT plans were generated for BrainLab microMLC using the iPlan treatment planning system. First, PTV constraints were used. The conformity of the dose prescribed to the PTV margin was compared between the two plan types for all patients. Second, NT constraints were introduced into each IMRT plan at 7Gy and 12Gy. Finally, these dose constraints were manipulated to achieve maximum NT sparing while maintaining the desired dose coverage of the PTV. RESULTS: Examination of the conformity index for an increase of >0.03 showed a separation of patients into those who did and did not benefit from IMRT. Some patients who did not have a significant benefit from conformity alone showed a reduction in max PTV dose by as much as 9%. Moreover, with NT dose constraints, the volume of NT irradiated decreased by a considerable amount for a subset of patients by 10-21.8% and 9-16.9% for the 7Gy and 12Gy constraints, respectively. CONCLUSIONS: This work indicates the potential for significant dose reduction to NT surrounding AVMs and provides rules to determine which patients are likely to benefit from IMRT. By utilizing IMRT plans, with and without dose constraints, NT sparing can be improved and risk of symptomatic injury reduced.

Factors predictive of obliteration after arteriovenous malformation radiosurgery.

OBJECTIVE: To investigate predictive factors of complete obliteration following treatment with linac-based stereotactic radiosurgery for intracerebral arteriovenous malformations. METHODS: Archived plans for 48 patients treated at the British Columbia Cancer Agency and who underwent post-treatment digital subtraction angiography to assess obliteration were studied. Actuarial estimates of obliteration were calculated using the Kaplan-Meier method. Univariate and multivariate Cox proportional hazards models were used for analysis of incidence of obliteration. Log-rank test was used to search for parameters associated with obliteration. RESULTS: Complete nidus obliteration was achieved in 38/48 patients (79.2%). Actuarial rate of obliteration was 75.9% at 4 years (95% confidence interval 63.1%-88.6%). On univariate analysis, prescribed dose to the margin (p=0.002) and dose to isocentre (p=0.022) showed statistical significance. No parameters were significant in a multivariate model. According to the log-rank test, prescribed dose to the margin of >20 Gy (p=0.004) and dose to the isocentre of >25 Gy (p=0.004) were associated with obliteration. CONCLUSION: Reported series in the literature suggest a number of different factors are predictive of complete obliteration of arteriovenous malformations following radiosurgery. However, differing definitions of volume and complete obliteration makes direct comparison between series difficult. This study demonstrates that complete obliteration of the nidus following linear accelerator-based stereotactic radiosurgery for arteriovenous malformations appears to be most closely related to the prescribed marginal dose. In particular, a marginal dose of >20 Gy is strongly associated with obtaining complete obliteration of the nidus.

[Investigation of hepatoprotector Metrop GP influence on tumor growth or antitumor action of doxorubicin].

An experimental model of solid tumor of Ehrlich failed to reveal any influence of the hepatoprotector Metrop GP on tumor growth or affect antitumor action of doxorubicin.

[Expression of Ki-67, p53, bcl-2, estrogen receptors alpha in patients with clear cell renal carcinoma and epidermal growth factor receptor mutation].

Three hundred and thirty-six cases of clear-cell renal carcinoma (CCRC) were examined for epidermal growth factor receptor (EGFR) mutation: exon 19 deletion and L858R mutation in exon 21 of the EGFR gene. The expression of Ki-67, bcl-2, p53, and estrogen receptors alpha was studied in CCRC with EGFR mutation. There were 4 cases of CCRC with EGFR exon 19 deletion. The frequency of EGFR gene mutations was 1.2%. L858R missense mutations in exon 21 of the EGFR gene were absent. In CCRC, EGFR gene mutation (exon 19 deletion) was detected in 3 men and 1 woman with an age range of 50 to 60 years and Fuhrman differentiation grade 2 or 3. The Ki-67 index varied from 4 to 23%. The expression of bcl-2 and p53 was negative. A moderate estrogen receptor alpha expression was revealed in 1 of 4 cases.