Valerijus Ostapenko

Immunohistochemistry profiles of breast ductal carcinoma: factor analysis of digital image analysis data

BackgroundMolecular studies of breast cancer revealed biological heterogeneity of the disease and opened new perspectives for personalized therapy. While multiple gene expression-based systems have been developed, current clinical practice is largely based upon conventional clinical and pathologic criteria. This gap may be filled by development of combined multi-IHC indices to characterize biological and clinical behaviour of the tumours. Digital image analysis (DA) with multivariate statistics of the data opens new opportunities in this field.MethodsTissue microarrays of 109 patients with breast ductal carcinoma were stained for a set of 10 IHC markers (ER, PR, HER2, Ki67, AR, BCL2, HIF-1α, SATB1, p53, and p16). Aperio imaging platform with the Genie, Nuclear and Membrane algorithms were used for the DA. Factor analysis of the DA data was performed in the whole group and hormone receptor (HR) positive subgroup of the patients (n = 85).ResultsMajor factor potentially reflecting aggressive disease behaviour (i-Grade) was extracted, characterized by opposite loadings of ER/PR/AR/BCL2 and Ki67/HIF-1α. The i-Grade factor scores revealed bimodal distribution and were strongly associated with higher Nottingham histological grade (G) and more aggressive intrinsic subtypes. In HR-positive tumours, the aggressiveness of the tumour was best defined by positive Ki67 and negative ER loadings. High Ki67/ER factor scores were strongly associated with the higher G and Luminal B types, but also were detected in a set of G1 and Luminal A cases, potentially indicating high risk patients in these categories. Inverse relation between HER2 and PR expression was found in the HR-positive tumours pointing at differential information conveyed by the ER and PR expression. SATB1 along with HIF-1α reflected the second major factor of variation in our patients; in the HR-positive group they were inversely associated with the HR and BCL2 expression and represented the major factor of variation. Finally, we confirmed high expression levels of p16 in Triple-negative tumours.ConclusionFactor analysis of multiple IHC biomarkers measured by automated DA is an efficient exploratory tool clarifying complex interdependencies in the breast ductal carcinoma IHC profiles and informative value of single IHC markers. Integrated IHC indices may provide additional risk stratifications for the currently used grading systems and prove to be useful in clinical outcome studies.Virtual SlidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1512077125668949

Membrane connectivity estimated by digital image analysis of HER2 immunohistochemistry is concordant with visual scoring and fluorescence in situ hybridization results: algorithm evaluation on breast cancer tissue microarrays

IntroductionThe human epidermal growth factor receptor 2 (HER2) is an established biomarker for management of patients with breast cancer. While conventional testing of HER2 protein expression is based on semi-quantitative visual scoring of the immunohistochemistry (IHC) result, efforts to reduce inter-observer variation and to produce continuous estimates of the IHC data are potentiated by digital image analysis technologies.MethodsHER2 IHC was performed on the tissue microarrays (TMAs) of 195 patients with an early ductal carcinoma of the breast. Digital images of the IHC slides were obtained by Aperio ScanScope GL Slide Scanner. Membrane connectivity algorithm (HER2-CONNECT™, Visiopharm) was used for digital image analysis (DA). A pathologist evaluated the images on the screen twice (visual evaluations: VE1 and VE2). HER2 fluorescence in situ hybridization (FISH) was performed on the corresponding sections of the TMAs. The agreement between the IHC HER2 scores, obtained by VE1, VE2, and DA was tested for individual TMA spots and patients maximum TMA spot values (VE1max, VE2max, DAmax). The latter were compared with the FISH data. Correlation of the continuous variable of the membrane connectivity estimate with the FISH data was tested.ResultsThe pathologist intra-observer agreement (VE1 and VE2) on HER2 IHC score was almost perfect: kappa 0.91 (by spot) and 0.88 (by patient). The agreement between visual evaluation and digital image analysis was almost perfect at the spot level (kappa 0.86 and 0.87, with VE1 and VE2 respectively) and at the patient level (kappa 0.80 and 0.86, with VE1max and VE2max, respectively). The DA was more accurate than VE in detection of FISH-positive patients by recruiting 3 or 2 additional FISH-positive patients to the IHC score 2+ category from the IHC 0/1+ category by VE1max or VE2max, respectively. The DA continuous variable of the membrane connectivity correlated with the FISH data (HER2 and CEP17 copy numbers, and HER2/CEP17 ratio).ConclusionHER2 IHC digital image analysis based on membrane connectivity estimate was in almost perfect agreement with the visual evaluation of the pathologist and more accurate in detection of HER2 FISH-positive patients. Most immediate benefit of integrating the DA algorithm into the routine pathology HER2 testing may be obtained by alerting/reassuring pathologists of potentially misinterpreted IHC 0/1+ versus 2+ cases.Virtual SlidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1973465132560208.

Xenogeneic therapeutic cancer vaccines as breakers of immune tolerance for clinical application: To use or not to use?

Accumulation of firm evidence that clinically apparent cancer develops only when malignant cells manage to escape immunosurveillance led to the introduction of tumor immunotherapy strategies aiming to reprogramm the cancer-dysbalanced antitumor immunity and restore its capacity to control tumor growth. There are several immunotherapeutical strategies, among which specific active immunotherapy or therapeutic cancer vaccination is one of the most promising. It targets dendritic cells (DCs) which have a unique ability of inducing naive and central memory T cell-mediated immune response in the most efficient manner. DCs can be therapeutically targeted either in vivo/in situ or by ex vivo manipulations followed by their re-injection back into the same patient. The majority of current DC targeting strategies are based on autologous or allogeneic tumor-associated antigens (TAAs) which possess various degrees of inherent tolerogenic potential. Therefore still limited efficacy of various tumor immunotherapy approaches may be attributed, among various other mechanisms, to the insufficient immunogenicity of self-protein-derived TAAs. Based on such an idea, the use of homologous xenogeneic antigens, derived from different species was suggested to overcome the natural immune tolerance to self TAAs. Xenoantigens are supposed to differ sufficiently from self antigens to a degree that renders them immunogenic, but at the same time preserves an optimal homology range with self proteins still allowing xenoantigens to induce cross-reactive T cells. Here we discuss the concept of xenogeneic vaccination, describe the cons and pros of autologous/allogeneic versus xenogeneic therapeutic cancer vaccines, present the results of various pre-clinical and several clinical studies and highlight the future perspectives of integrating xenovaccination into rapidly developing tumor immunotherapy regimens.

Quality of life of Lithuanian women with early stage breast cancer

BackgroundIn the last decades, there have been no studies carried out in Lithuania on the quality of life of breast cancer patients. The aim of the present study was to evaluate changes in the quality of life of Lithuanian women with the early stage of breast cancer nine months after surgery and its dependence on surgical strategy, adjuvant chemotherapy and the social and demographic status of the patients.MethodsSeventy-seven patients with early stage breast cancer filled in the FACT-An questionnaire twice: one week and nine months after the surgery. The main age of the patients was 53.1 ± 10.6 years. We distinguished the mastectomy group and breast conserving treatment (BCT) group with/without chemotherapy. The groups were identical in their social and demographic status (age, education, occupation and marital status). Changes in the quality of life in these groups were compared nine months after surgery.ResultsNine months after surgery, the overall quality of life was found worse in both mastectomy and BCT groups. Changes were induced by the worsening of the emotional and social well-being. The quality of life became worse in the mastectomy plus chemotherapy sample. No changes were detected in the mastectomy group without chemotherapy. In addition, the multivariate analysis showed that the marital status was quite a significant determinant of the functional well-being.ConclusionNine months after surgery, the study revealed a worsening of the overall quality of life in both groups of patients – those who had undergone mastectomy and BCT. The quality of life became considerably worse in the mastectomy plus chemotherapy group. Marital status was found to exert the most considerable influence on the womens quality of life in comparison with other social and demographic factors.

Ki67/SATB1 ratio is an independent prognostic factor of overall survival in patients with early hormone receptor-positive invasive ductal breast carcinoma

Biological diversity of breast cancer presents challenges for personalized therapy and necessitates multiparametric approaches to understand and manage the disease. Multiple protein biomarkers tested by immunohistochemistry (IHC), followed by digital image analysis and multivariate statistics of the data, have been shown to be effective in exploring latent profiles of tumor tissue immunophenotype. In this study, based on tissue microarrays of 107 patients with hormone receptor (HR) positive invasive ductal breast carcinoma, we investigated the prognostic value of the integrated immunophenotype to predict overall survival (OS) of the patients. A set of 10 IHC markers (ER, PR, HER2, Ki67, AR, BCL2, HIF-1α, SATB1, p53, and p16) was used. The main factor of the variance was characterized by opposite loadings of ER/PR/AR/BCL2 and Ki67/HIF-1α; it was associated with histological grade but did not predict OS. The second factor was driven by SATB1 expression along with moderate positive HIF-1α and weak negative Ki67 loadings. Importantly, this factor did not correlate with any clinicopathologic parameters, but was an independent predictor of better OS. Ki67 and SATB1 did not reach statistical significance as single predictors; however, high Ki67/SATB1 ratio was an independent predictor of worse OS. In addition, our data indicate potential double prognostic meaning of HIF-1α expression in breast cancer and necessitate focused studies, taking into account the immunophenotype interactions and tissue heterogeneity aspects.

CD8highCD57+ T-cell population as an independent predictor of response to chemoradiation therapy in extensive-stage small cell lung cancer

OBJECTIVES Tangible clinical benefit is achieved in only a relatively small proportion of extensive-stage small cell lung cancer (SCLC) patients receiving current treatment strategies. Therefore, a more personalized use of current and novel treatment approaches is of critical importance. Individualized therapy relies on the identification of specific biomarkers predictive of response to a particular type of cancer treatment. Immune-related parameters emerge as powerful biomarkers among a variety of predictors of clinical response to various types of cancer treatment. PATIENTS AND METHODS Using multicolor flow cytometry, we evaluated a predictive value of CD8(high)CD57(+) T-cell population and its immunosuppressive (FOXP3(+), NKG2A(+)) and cytotoxic (Perforin(+)) subsets in the peripheral blood of extensive-stage SCLC patients (n=82) treated with either chemotherapy-alone (n=24), or chemoradiation therapy (n=42), or receiving best supportive care (n=16). RESULTS The low level (<20%) of CD8(high)CD57(+) T cells within the peripheral blood CD8(+) T-cell population and the low level (<3%) of the immunosuppressive FOXP3-positive subset within the CD8(high)CD57(+) T-cell population were independent predictors of a better response to treatment with chemoradiation therapy, but not with chemotherapy alone or best supportive care. Importantly there was no significant survival difference between SCLC patients who were: (i) treated with chemoradiation, but had an unfavourable immune profile (≥20% of CD8(high)CD57(+) T cells and ≥3% of its FOXP3-positive subset), (ii) treated with chemotherapy alone, or (iii) received best supportive care. CONCLUSIONS We show that only a combination of chemotherapy with radiation therapy offered a considerable survival benefit that was confined to a subset of extensive-stage SCLC patients with a favourable predictive immune profile in the peripheral blood.

Impact of season of diagnosis on mortality among breast cancer survivors

Introduction: There is mounting evidence that the time of breast cancer diagnosis and the start of treatment can improve survival rates. The aim of this study was to test the relationship between the season of breast cancer diagnosis and the survival of women patients receiving standard surgery treatment with radiotherapy. Materials and Methods: The nonmetastatic breast cancer patients (n = 991) were followed from the date of diagnosis until death. Cox proportional hazards models were used to calculate multivariate hazard ratios (HRs) for all-cause mortality. HRs and 95% confidence intervals (CIs) were estimated in models adjusted for clinicopathologic and treatment factors. Results: After adjusting for independent prognostic variables, we found that patients diagnosed in summer and autumn had a 40% reduced risk for 0–3-year mortality when compared to those diagnosed in spring. Among women aged <50 years, HRs comparing autumn with spring diagnosis categories were 0.53 (95% CI: 0.31–0.91) for 0–5-years and 0.68 (95% CI: 0.46–0.89) for 5–10-years after diagnosis. Diagnosis in autumn was associated with improving survival in younger patients treated with adjuvant chemotherapy (HR = 0.61, 95% CI: 0.39–0.96, P = 0.003). Conclusions: The diagnosis in summer and autumn was associated with a better overall prognosis. The effect of season of diagnosis on survival rate was most pronounced in the young age patients receiving chemotherapy.

Value ER, PR and HER Status as Independent Prognostic Factor of Local Recurrence after Breast Conserving Surgery.

Aim : Evaluate value ER, PR and HER2 status as independent prognostic factor of local recurrence after breast conserving surgery Methods and materials: The retrospective study of 537 breast cancer patients stage I,II,III which were combined treated in Institute Oncology, Vilnius University in 2005-2006.was made The surgery treatment was used as primarily selection criteria, the data on vital status of all patients was summarized by December, 2008. The median follow-up time was 33 month. Pts age was from 29 years to 76 years. Premenopausal pts ( =10) - 166 pts (30,9%), medium (score 6-9) - 146 pts (27,2%), week (scrore =10) - 141 pts (26,6%), medium (score 6-9) - 125 pts (23,3%), week (scrore Results: Local recurrence rate was calculated according Nelson-Aalen cumulative hazard estimate (analysis time 4 years) and was equal to 8 %.Local recurrence rate according ER-, PR-, HER2 0 vs ER+, PR+, HER2 3+ was equal to 14% vs 4,5%.In 63,5% of all local recurrence ER, PR were negative “0 “,in 40.9% of all local recurrences HER2 was positive (3+) Conclusion: According to our preliminary results ER, PR and HER2 status are independent prognostic factor of local recurrence after breast. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6047.

Vyrų krūties vėžio diagnostika ir gydymas

Algirdas Jackevicius, Leonarda Sarakauskienė, Valerijus Ostapenko, Saulius Bružas, Juozas Kurtinaitis, Algimantas Mudėnas Vilniaus universiteto Onkologijos institutas, Santariskių g.1, LT-08660 Vilnius Kauno medicinos universiteto Onkologijos ligoninė, Volungių g. 16, LT-45434 Kaunas El pastas: algirdasj@is.lt Įvadas / tikslas Vyrų krūties vėžys yra reta onkologinė liga. Siame straipsnyje pateikiame 75 ligonių, gydytų dviejose onkologijos ligoninėse, klinikinius duomenis, tiesioginius ir vėlyvuosius gydymo rezultatus. Ligoniai ir metodai Straipsnyje nagrinėjami Vilniaus universiteto Onkologijos instituto klinikoje ir Kauno universiteto Onkologijos ligoninėje1988–2003 metais gydytų 75 vyrų, sirgusių krūties vėžiu, klinikiniai duomenys. Ligonių amžiaus vidurkis – 69,2 metų (jauniausias 41, vyriausias 90 metų). Astuoniems (10,7%) ligoniams diagnozuotas pirmos stadijos vėžys, 35 (46,7%) – antros, 22 (29,3%) – trecios, 10 (13,3%) ligonių – ketvirtos. Pagal histologinius vėžio tipus dažniausiai diagnozuota intraduktalinė karcinoma – 40 ligonių, 8 ligoniams – lobulinė karcinoma, taciau 14 ligonių vėžio histologinis tipas nenustatytas. Dažniausiai, t. y. 53 ligoniams, buvo atlikta modifikuota Maddeno mastektomija, astuoniems – paprastoji mastektomija, 8 ligoniams taikyta spindulinė terapija, 6 ligoniams dėl sunkių gretutinių ligų – simptominis gydymas. 37 ligoniai gydyti kombinuotu būdų: 30 – spinduline terapija, 5 – chemoterapija, 2 – abiem gydymo metodais. Rezultatai Vėlyvieji gydymo rezultatai nebuvo geri: is 75 gyvena 28 liginiai, 47 mirė. Penkerius metus isgyveno 83,3% (95% PI 27,3–97,5) serganciųjų pirmos stadijos krūties vėžiu, 69,6% (95% PI 49,1–83,1) – antros stadijos ir tik 6,2% (95% PI 4,2–24,2) – trecios. Nė vienas ligonis, kuriam diagnozuota ketvirta ligos stadija, neisgyveno penkerių metų. Isvados Lietuvoje vyrų sergamumas krūties vėžiu per pastaruosius metus nepakito. Nemažai vyrų buvo gydyti nuo IIIB stadijos krūties vėžio, kai navikas jau buvo lokaliai isplites ir susiformavo vėžinė opa. Vyrų, sergancių trecios stadijos krūties vėžiu, prognozė yra blogesnė negu moterų. Ligos stadija nulemia ligonių gyvenimo trukme, tai patvirtina statistinis vėlyvųjų gydymo rezultatų skaiciavimas. Reiksminiai žodžiai: vyrų krūties vėžys, diagnostika, gydymas, vėlyvieji rezultatai The diagnostics and treatment of male breast carcinoma Algirdas Jackevicius, Leonarda Sarakauskienė, Valerijus Ostapenko, Saulius Bružas, Juozas Kurtinaitis, Algimantas Mudėnas Vilnius University Institute of Oncology, Santariskių str. 1, LT-08660 Vilnius, Lithuania Kaunas Medical University Oncological Hospital, Volungių str. 16, LT-45434, Kaunas, Lithuania E-mail: algirdasj@is.lt Background / objective Male breast cancer is an uncommon oncological disease. In this paper, we have analysed the results of treatment of 75 patients treated in two oncologycal clinics. Patients and methods We analysed 75 male patients treated in 1988–2003 in the clinics of the Institute of Oncology of Vilnius University and Hospital of Oncology of Kaunas University of Medicine. The mean age of patients was 69.2 years (range, 46–90 years). The staging of disease: stage I 8 (10.7%) patients, stage II 35 (46.7%), stage III 22 (29.3%). Ten (13.3%) patients were treated in stage IV of the disease. The most common method of treatment was radical mastectomy by Madden which was performed in 53 cases. In 8 cases mastectomy simplex was performed. Eight patients received radiotherapy. The patients received this conservative treatment in late stages of the disease. In seven cases the patients were in poor health state, and only palliative treatment was applied. 37 patients received combined treatment: 30 patients were treated with radiotherapy, two patients received radiotherapy and chemotherapy, in 5 cases after mastectomy the patients were treated with chemotherapy. Results The follow-up results were not satisfactory: from 75 patients 28 patients are alive and 47 died. The 5-year survival of the patients according to the stage of disease: 83.3% (95% CI 27.3–97.5) of patients in stage I, 69.6% (95% CI 49.1–83.1) in stage II, stage III 6.2% (95% CI 4.2–24.2). In this period, all patients in stage IV of the disease died. Conclusions The incidence of male breast cancer in Lithuania is low, and over the last years has remained at the same level. Many of our patients had ulceration of tumor and were treated in stage III B of the disease. The prognosis of male breast cancer in stage III of the disease is worse than of female breast cancer of the same stage. The stage of the disease was statistically significant for the survival of patients. Keywords: male breast carcinoma, diagnosis, treatment, follow-up results