Antonino Maiorana

Comparison of HER-2 and hormone receptor expression in primary breast cancers and asynchronous paired metastases: impact on patient management.

INTRODUCTION The assessment of hormone receptors (HRs) and human epidermal growth factor receptor (HER)-2 is necessary to select patients who are candidates for hormonal and anti-HER-2 therapy. The evaluation of these parameters is generally carried out in primary tumors and it is not clear if reassessment in metastatic lesions might have an impact on patient management. The primary aim of this analysis was to compare HER-2 and HR status in primary tumors versus metastatic sites in breast cancer patients. PATIENTS AND METHODS Seventy-five patients with available samples from primary tumors and paired metastases were included. HER-2 status was evaluated by immunohistochemistry (IHC) and/or fluorescence in situ hybridization (FISH); HR status was assessed by IHC. RESULTS Nineteen percent of primary tumors were HER-2 positive; 77% were HR positive. Sites of biopsied or resected metastases were: locoregional soft tissues (n = 30), liver (n = 20), central nervous system (n = 5), bone (n = 5), pleura (n = 4), distant soft tissues (n = 3), abdomen (stomach, colon, peritoneum) (n = 3), bronchus (n = 3), and bone marrow (n = 2). For paired metastases, the HER-2 status was unchanged in 84% of cases; two patients changed from positive to negative, while 10 patients converted from negative to positive (agreement, 84%; kappa = 0.5681). A change in HR status was observed in 16 cases (21%): nine cases from positive to negative and seven cases from negative to positive (agreement, 78.7%; kappa = 0.4158). CONCLUSIONS Further studies are necessary to better define the level of discordance in HER-2 or HR status between primary tumors and paired metastases. However, a biopsy of metastatic disease can be recommended, if feasible with minimal invasiveness, because treatment options might change for a significant proportion of patients.

A prognostic model based on nodal status and Ki-67 predicts the risk of recurrence and death in breast cancer patients with residual disease after preoperative chemotherapy

BACKGROUND Preoperative chemotherapy (PCT) allows for in vivo testing of treatment effects on tumor and its microenvironment. Aim of this analysis was to evaluate the effect of PCT on tumor biomarker expression and to evaluate the prognostic role of treatment-induced variation of these biomarkers (molecular response). METHODS Two hundred and twenty-one stage II-III breast cancer patients were included. The following parameters were evaluated at baseline and on surgical specimens after PCT: estrogen receptor (ER), progesterone receptor (PgR), human epidermal growth factor receptor 2 (HER2), Ki-67, p53, human epidermal growth factor receptor (EGFR), vascular endothelial growth factor receptor 2 (VEGFR2), and apoptosis. RESULTS A pathological complete response was observed in 8.8% of the patients. PCT induced a significant reduction in the expression of ER, PgR, Ki-67, and apoptosis. As by multivariable model, Ki-67 > or = 15% and nodal positivity after preoperative chemotherapy (PCT) were significant predictors of worse disease-free survival [hazard ratio (HR) 3.79, P < 0.0001 and HR 2.31, P = 0.037, respectively]. Ki-67 > or = 15% after PCT was also a significant predictor of overall survival (HR 3.75, P = 0.013). On the basis of these two parameters, patients were classified into three groups: (i) low risk (negative nodes and Ki-67 <15%), (ii) intermediate risk (nodal positivity or Ki-67 > or = 15%), and (iii) high risk (nodal positivity and Ki-67 > or = 15%). As compared with the low-risk group, the HRs for recurrence were 3.1 and 9.3 for the intermediate- and high-risk group, respectively (P = 0.0001); the HRs for death were 2.4 and 6.5 for the intermediate- and high-risk group, respectively (P = 0.042). CONCLUSIONS Ki-67 and nodal status have been used to generate a simple and easily reproducible prognostic model, able to discriminate patients with worse prognosis among the heterogeneous group of women with residual disease after PCT.

Prognostic relevance of serum β2 microglobulin in patients with follicular lymphoma treated with anthracycline-containing regimens. A GISL study

Background and Objectives Although serum β2 microglobulin (β2 M) is an easy parameter to measure, and over-expressed in a large number of lymphoproliferative diseases, its prognostic value has been largely underestimated. The present study examined the influence of β2M levels on overall survival (OS) of patients with follicular lymphoma (FL). Design and Methods The prognostic role of β2M was evaluated in 236 patients with FL identified from the databases of the Gruppo Italiano per lo Studio dei Linfomi (GISL) and treated with anthracycline-based regimens from 1993 to 2003. Results Elevated serum β2M levels were found in 82 patients (35%). According to multivariate logistic regression analysis, elevated β2M levels were associated with elevated lactate dehydrogenase (LDH) (p=0.021), age (p=0.029), and number of involved nodal areas (p<0.001). The percentage of elevated β2M levels increased progressively with increasing FLIPI scores (17%, 38%, and 63% in the low-, intermediate-, and high-risk groups, respectively). Five-year OS was 61% (95% CI, 47–73%) and 89% (95% CI, 82–93%) for patients with elevated vs normal β2M levels respectively (p<0.001). Cox regression analysis showed that β2M level had an independent and stable prognostic value (HR=3.0; 95%CI, 1.6–5.7). In a multivariate analysis the impact of β2M level on survival was independent of FLIPI score, with a HR of 2.94 (95% CI, 1.54–5.62). Interpretation and Conclusions Our results demonstrate that in patients treated in the pre-rituximabera, β2M level was an independent prognostic marker in addition to FLIPI score. We thus suggest that β2M be routinely assessed and tested in future prognostic studies of FL patients treated with combination chemotherapy and anti-CD20 agents.

Decreasing incidence of gastric MALT lymphomas in the era of anti-Helicobacter pylori interventions: results from a population-based study on extranodal marginal zone lymphomas

BACKGROUND Few studies have been carried out to date that have addressed the epidemiology of extranodal marginal zone lymphomas (EN-MZLs). PATIENTS AND METHODS We carried out a population-based study to investigate incidence rates (IRs) and time trends of EN-MZL diagnosed in the province of Modena (Italy) from 1997 to 2007. RESULTS One hundred and sixty-five cases were identified from the Modena Cancer Registry that corresponded to an age-standardized IR of 2.3 cases per 100 000. A bimodal distribution of age was shown with the group of young patients mostly represented by males with cutaneous lymphoma. No time trends were observed for the IR; the incidence of gastric mucosa-associated lymphoid tissue (g-MALT) lymphomas (N = 51) markedly declined during the study period, dropping from 1.4 in 1997 to 0.2 in 2002 and then remaining stable until 2007; the calculated annual percent change for g-MALT was -17.0% (95% confidence interval -26.6% to -6.2%). We also observed a significant decrease in the rate of g-MALT associated with Helicobacter pylori (HP) infection from 61% to 17% of patients diagnosed before and after 2002 (P = 0.007; P for trend = 0.016). CONCLUSION This population-based study provides new insights into recent changes in the epidemiology of EN-MZL, mainly represented by the sharp reduced incidence of HP-positive g-MALT lymphomas.

Double-Blind, Placebo-Controlled, Multicenter, Randomized, Phase IIB Neoadjuvant Study of Letrozole-Lapatinib in Postmenopausal Hormone Receptor–Positive, Human Epidermal Growth Factor Receptor 2–Negative, Operable Breast Cancer

PURPOSE This is a randomized, double-blind, placebo-controlled study aimed to evaluate the clinical and biologic effects of letrozole plus lapatinib or placebo as neoadjuvant therapy in hormone receptor (HR) -positive/human epidermal growth factor receptor 2 (HER2) -negative operable breast cancer. METHODS Ninety-two postmenopausal women with stage II to IIIA primary breast cancer were randomly assigned to preoperative therapy consisting of 6 months of letrozole 2.5 mg orally daily plus lapatinib 1,500 mg orally daily or placebo. Surgery was performed within 2 weeks from the last study medication. Clinical response was assessed by ultrasonography. Pre- and post-treatment samples were evaluated for selected biomarkers. Fresh-frozen tissue samples were collected for genomic analyses. RESULTS Numerically similar clinical response rates (partial + complete response) were observed (70% for letrozole-lapatinib and 63% for letrozole-placebo). Toxicities were generally mild and manageable. A significant decrease in Ki-67 and pAKT expression from baseline to surgery was observed in both arms. Overall, 34 patients (37%) had a mutation in PIK3CA exon 9 or 20. In the letrozole-lapatinib arm, the probability of achieving a clinical response was significantly higher in the presence of PIK3CA mutation (objective response rate, 93% v 63% in PIK3CA wild type; P = .040). CONCLUSION The combination of letrozole-lapatinib in early breast cancer was feasible, with expected and manageable toxicities. In unselected estrogen receptor-positive/HER2-negative patients, letrozole-lapatinib and letrozole-placebo resulted in a similar overall clinical response rate and similar effect on Ki-67 and pAKT. Our secondary end point findings of a significant correlation between PIK3CA mutation and response to letrozole-lapatinib in HR-positive/HER2-negative early breast cancer must now be independently confirmed.

Prospective Biomarker Analysis of the Randomized CHER-LOB Study Evaluating the Dual Anti-HER2 Treatment With Trastuzumab and Lapatinib Plus Chemotherapy as Neoadjuvant Therapy for HER2-Positive Breast Cancer

BACKGROUND The CHER-LOB randomized phase II study showed that the combination of lapatinib and trastuzumab plus chemotherapy increases the pathologic complete remission (pCR) rate compared with chemotherapy plus either trastuzumab or lapatinib. A biomarker program was prospectively planned to identify potential predictors of sensitivity to different treatments and to evaluate treatment effect on tumor biomarkers. MATERIALS AND METHODS Overall, 121 breast cancer patients positive for human epidermal growth factor 2 (HER2) were randomly assigned to neoadjuvant chemotherapy plus trastuzumab, lapatinib, or both trastuzumab and lapatinib. Pre- and post-treatment samples were centrally evaluated for HER2, p95-HER2, phosphorylated AKT (pAKT), phosphatase and tensin homolog, Ki67, apoptosis, and PIK3CA mutations. Fresh-frozen tissue samples were collected for genomic analyses. RESULTS A mutation in PIK3CA exon 20 or 9 was documented in 20% of cases. Overall, the pCR rates were similar in PIK3CA wild-type and PIK3CA-mutated patients (33.3% vs. 22.7%; p = .323). For patients receiving trastuzumab plus lapatinib, the probability of pCR was higher in PIK3CA wild-type tumors (48.4% vs. 12.5%; p = .06). Ki67, pAKT, and apoptosis measured on the residual disease were significantly reduced from baseline. The degree of Ki67 inhibition was significantly higher in patients receiving the dual anti-HER2 blockade. The integrated analysis of gene expression and copy number data demonstrated that a 50-gene signature specifically predicted the lapatinib-induced pCR. CONCLUSION PIK3CA mutations seem to identify patients who are less likely to benefit from dual anti-HER2 inhibition. p95-HER2 and markers of phosphoinositide 3-kinase pathway deregulation are not confirmed as markers of different sensitivity to trastuzumab or lapatinib. IMPLICATIONS FOR PRACTICE HER2 is currently the only validated marker to select breast cancer patients for anti-HER2 treatment; however, it is becoming evident that HER2-positive breast cancer is a heterogeneous disease. In addition, more and more new anti-HER2 treatments are becoming available. There is a need to identify markers of sensitivity to different treatments to move in the direction of treatment personalization. This study identified PIK3CA mutations as a potential predictive marker of resistance to dual anti-HER2 treatment that should be further studied in breast cancer.

Microenvironment inflammatory infiltrate drives growth speed and outcome of hepatocellular carcinoma: a prospective clinical study

In HCC, tumor microenvironment, heavily influenced by the underlying chronic liver disease, etiology and stage of the tissue damage, affects tumor progression and determines the high heterogeneity of the tumor. Aim of this study was to identify the circulating and tissue components of the microenvironment immune-mediated response affecting the aggressiveness and the ensuing clinical outcome. We analyzed the baseline paired HCC and the surrounding tissue biopsies from a prospective cohort of 132 patients at the first diagnosis of HCC for immunolocalization of PD-1/PD-L1, FoxP3, E-cadherin, CLEC2 and for a panel of 82 microRNA associated with regulation of angiogenesis, cell proliferation, cell signaling, immune control and autophagy. Original microarray data were also explored. Serum samples were analyzed for a panel of 19 cytokines. Data were associated with biochemical data, histopathology and survival. Patients with a more aggressive disease and shorter survival, who we named fast-growing accordingly to the tumor doubling time, at presentation had significantly higher AFP levels, TGF-β1 and Cyphra 21-1 levels. Transcriptomic analysis evidenced a significant downregulation of CLEC2 and upregulation of several metalloproteinases. A marked local upregulation of both PD-1 and PD-L1, a concomitant FoxP3-positive lymphocytic infiltrate, a loss of E-cadherin, gain of epithelial–mesenchymal transition (EMT) phenotype and extreme poor differentiation at histology were also present. Upregulated microRNA in fast-growing HCCs are associated with TGF-β signaling, angiogenesis and inflammation. Our data show that fast HCCs are characterized not only by redundant neo-angiogenesis but also by unique features of distinctively immunosuppressed microenvironment, prominent EMT, and clear-cut activation of TGFβ1 signaling in a general background of long-standing and permanent inflammatory state.

Expression of nuclear survivin in normal skin and squamous cell carcinoma: a possible role in tumour invasion.

Background:Survivin is detected in few adult normal cells and it is highly expressed in cancer. Nuclear survivin facilitates cell cycle entry, whereas the mitochondrial pool protects cells from apoptosis. Survivin is overexpressed in keratinocyte stem cells (KSCs) and protects them from apoptosis.Methods:As KSCs are at the origin of squamous cell carcinoma (SCC), we evaluated survivin expression in normal and cancerous skin in vivo by immunohistochemistry and western blotting. HaCaT cells overexpressing survivin and wound-healing assay are used. Analysis of variance and Student’s T-tests are used for statistical analysis.Results:Survivin is localised in both the cytoplasm and nucleus of normal adult and young keratinocytes. Nuclear survivin is detected in one every 10 of 11 basal keratinocytes. When present in suprabasal cells, nuclear survivin is coexpressed with K10 but not with K15 or p75-neurotrophin receptor (p75NTR), a transit amplifying cell marker. Nuclear, but not cytoplasmic, survivin expression markedly increases in actinic keratosis and in SCC in situ, as compared with normal epidermis, and it is highest in poorly differentiated SCC. In SCC tumours, nuclear survivin-positive cells are mainly K10/p75NTR-negative and K15-positive. In poorly differentiated tumours, survivin mostly localises in the deep infiltrating areas. When overexpressed in keratinocytes, survivin increases cell migration.Conclusion:High survivin expression and the subcellular localisation of survivin correlate with keratinocyte differentiation and are associated with undifferentiated and more invasive SCC phenotype.

Unexpected death: Anaphylactic intraoperative death due to Thymoglobulin carbohydrate excipient

Anaphylactic shock is a life-threatening allergic response characterized by severe hypotension, inducing tissue hypoperfusion with possible multi-organ failure and death. We describe the first case of fatal intra-operative anaphylactic shock due to prolonged infusion of Thymoglobulin during Orthotopic Liver Transplantation (OLT), resulting from recruitment of both mastocytes and basophils, activated and degranulated. Post-mortem serological analysis on a preserved, pre-OLT sample of the patients blood revealed specific IgE against carbohydrate cross-reactive determinants (CCDs), such as MUXF3 and nAna c2, proving that anaphylactic reaction was triggered by the Thymoglobulin carbohydrate excipient (sugar alcohol mannitol), rather than anti-thymocyte globulin itself. Our findings are consistent with scientific data reported in the literature, where only one case of non-fatal anaphylaxis to Thymoglobulin has been described, despite the existence of proven cases of anaphylactic reaction to mannitol. This case highlights the need to pay particular attention in future not only to active substances but also to drug excipients, above all during intra-operative drug delivery. In view of the important role played by basophils in this kind of anaphylaxis, the basophil activation test (BAT) could prove useful in preventing anaphylactic death from CCDs.

Predictive and prognostic role of p53 according to tumor phenotype in breast cancer patients treated with preoperative chemotherapy: a single-institution analysis

Introduction and aims The p53 protein is a mediator of the cellular response to DNA damage. The aim of this study was to evaluate the predictive and/or prognostic value of p53 expression in relation to the molecular subtypes of breast cancer in patients treated with preoperative chemotherapy. Patients and methods Patients with stage II-III breast cancer were included in the study. The expression of p53 was evaluated by immunohistochemistry on the diagnostic core biopsy specimen. Patients received 4–6 courses of preoperative chemotherapy. Pathological complete response (pCR) was defined as complete disappearance of invasive tumor in the breast and axillary lymph nodes. Results 154 patients were included in the study and the molecular subtypes of their tumors were classified as follows: triple negative 18.2%, hormone receptor positive 60.4%, and HER2 positive 21.4%. p53 was expressed in 43.5% of the patients. A significant association between p53 expression and breast cancer molecular subtypes, tumor differentiation, and proliferation was observed. pCR was achieved in 8 patients (5.2%). p53 expression, molecular subtype, and nuclear grading were significant predictors of pCR (odds ratio for pCR in patients with p53-expressing tumors 10.03, p=0.0077). In univariate analysis, the expression of p53 as well as high proliferation and lymph node involvement after preoperative chemotherapy were predictors of a worse disease-free survival. Patients with p53 positivity also had a worse overall survival. In multivariate analysis, both p53 expression and nodal status after preoperative chemotherapy were significantly associated with disease-free and overall survival: the hazard ratios for relapse and death in patients with p53-expressing versus non-p53-expressing tumors were 2.29 (p=0.015) and 7.74 (p=0.002), respectively. The hazard ratios for relapse and death in node-positive versus node-negative patients were 3.63 (p=0.003) and 3.64 (p=0.041), respectively. Conclusions In this series of patients, p53 expression was significantly associated with markers of aggressive tumor biology, and with a higher likelihood of attaining pCR. p53 expression was a negative prognostic parameter for disease-free and overall survival in univariate and multivariate analysis.