Guidalberto Fabris

Epidermal Growth Factor Receptor (EGFR) Status in Primary Colorectal Tumors Does Not Correlate With EGFR Expression in Related Metastatic Sites: Implications for Treatment With EGFR-Targeted Monoclonal Antibodies

PURPOSE We hypothesized that the detection of epidermal growth factor receptor (EGFR) expression performed in primary tumors for treatment with EGFR-targeted monoclonal antibodies could not always correlate with EGFR status in metastatic sites, thus making cancer cells in these sites resistant to therapy. The aim of our study was to correlate EGFR expression on primary tumors and related metastases in order to find out whether assessing EGFR status on primary cancer is to be considered an effective tool for planning treatment with EGFR-targeted antibodies. PATIENTS AND METHODS We retrospectively evaluated EGFR immunohistochemistry from primary tumors and related metastatic sites in 99 colorectal cancer patients. The site of primary tumor was colon in 77 patients (78%) and rectum in 22 patients (22%). Metastatic sites analyzed were liver in 84 patients (81%), lung in 13 patients (13%), bone in one patient (1%), and brain in five patients (5%). EGFR status was defined as positive if the percentage of malignant cells stained was > or = 1%. RESULTS EGFR status was positive in 53 primary tumors (53%). In 19 primary tumors expressing EGFR (36%), the corresponding metastatic site was found negative, whereas it was found positive in seven metastases (15%) from EGFR-negative primary cancers. The difference between these two groups of patients (ie, EGFR-positive to EGFR-negative v EGFR-negative to EGFR-positive) was statistically significant (P = .036). CONCLUSION Our results suggest that the detection of the EGFR in primary colorectal cancer could be inadequate for planning therapy with EGFR-targeted monoclonal antibodies in a considerable proportion of both EGFR-positive and -negative primary tumors (36% and 15%, respectively).

HER‐2 status discrepancy between primary breast cancer and metastatic sites. Impact on target therapy

In this prospective study, we determined HER‐2 status in primary breast invasive carcinomas and in the paired lymph node metastases (synchronous and metachronous), local recurrence and metachronous distant metastases, to verify the percentage of discordant cases. HercepTest™ and Fluorescence in situ hybridization (FISH) were used to determine HER‐2 status on 119 cases of primary infiltrating breast carcinoma and paired metastases (45 cases with synchronous lymph node metastases, 9 cases with metachronous lymph node metastases, 30 cases with local recurrence, and 35 cases with metachronous distant metastases). A therapeutically significant HER‐2 status discordance was demonstrated between primary carcinoma and synchronous lymph node metastases (6.7%), local recurrence (13.3%) and metachronous distant metastases (28.6%). In the first comparison, there was a normal HER‐2 status in primary tumours and HER‐2 amplification in paired metastases, in the second the opposite phenomenon was present, and both types of discordance were evident in the third comparison. Considering the cases of local recurrences and metachronous distant metastases all together, 14 out of 65 cases (21.5%) showed a therapeutically significant discordance of HER‐2 status between the primary tumour and the paired metachronous recurrence or metastasis (p < 0.001), the 15.4% of cases showing normal HER‐2 status in the primary tumour and HER‐2 amplification in the neoplastic relapse. For the treatment of metastatic patients, the evaluation of HER‐2 status should be performed in neoplastic tissue from metastatic site, whenever possible. This procedure could be also suggested in the patients that are metastatic at the time of diagnosis.

Long-term experience with low-dose interferon-α and PUVA in the management of early mycosis fungoides

Abstract:  Objectives: Combined high‐dose Interferon‐α and psoralen plus ultraviolet A irradiation (PUVA) have been reported to be effective in the treatment of early mycosis fungoides (MF); however, our study is the first controlled prospective study in the literature exploring the activity and tolerability of the combination with low dosages and evaluating further clinical outcome of early‐MF patients. Methods: We carried out a multicentric prospective Phase II clinical study on 89 patients with early‐stage IA to IIA MF treated for 14 months with low‐dose IFN‐α2b (6–18 MU/wk) and PUVA. Treatment success was analysed in terms of freedom from treatment failure. Results and conclusions: Complete remission (CR) was achieved in 84% and an overall response rate in 98% of cases: six‐month CR was associated with a non‐confluent skin infiltrate at histology (P = 0.044) and 14‐month CR with high epidermal CD1a+ dendritic‐cell density (P = 0.030). The combination protocol was successfully tolerated and the most common reason of ‘failure’ was related to relapse and not to toxicity. Sustained remissions were achieved in 20% of patients. High CD8+ lymphoid T‐cell density was associated with a lower relapse rate (P = 0.002). We think that our combination therapy can be considered an alternative approach compared with other modalities. Good immunological host surveillance in the skin lesions seems to be an optimal basis for the therapeutic success.

Benign, borderline, and well-differentiated malignant intestinal mucinous tumors of the ovary: a clinicopathologic, histochemical, immunohistochemical, and nuclear quantitative study of 57 cases

Mucinous ovarian tumors are still a subject of controversy because they can show either intestinal or endocervical differentiation. Morphologic distinction between borderline and malignant tumors is sometimes difficult, and their clinical behavior has not been definitively ascertained. We selected 10 mucinous cystadenomas (MCAs), 32 intestinal mucinous borderline tumors (IMBTs), and 15 well-differentiated mucinous carcinomas (MCCs), all with goblet cells, at least focally. In all cases, we studied the clinicopathologic features, mucin content, intermediate filament expression, and some nuclear quantitative features, namely, the volume-corrected mitotic index (M/Vi), percentage of nucleolated nuclei, mean number of nucleoli per nucleus, percentage of nucleoli touching the nuclear membrane, and mean nuclear area. The quantitative nuclear study included cytometric DNA analysis and the results were expressed as relative mean ploidy value (RMPV) and as diploid-tetraploid or aneuploid histograms. The results of the quantitative study were evaluated statistically. All patients had stage IA tumors, had received surgical therapy only, and were alive and well after a follow-up of more than 5 years. Light microscopic examination revealed that destructive stromal invasion was not present in any MCCs and that IMBTs and MCCs were easily recognizable using the Hart and Norris criteria, later expanded by Hart. Mucin histochemistry and intermediate filament immunohistochemistry failed to detect substantial differences between the diagnostic categories. DNA analysis demonstrated an increase in aneuploid tumors going from IMBTs to MCCs, but these differences were not statistically significant. On the other hand, nuclear quantitative morphology showed significant differences among the three groups of tumors for all features considered. Forward stepwise discriminant analysis highlighted that MCAs, IMBTs, and MCCs were contiguous but different categories. These data support the separation of IMBTs and MCCs into morphologically different categories as underlined by the results of quantitative nuclear morphologic analysis. The favorable outcome of all patients confirms the excellent prognosis of stage I IMBTs and suggests that well-differentiated MCCs without destructive stromal invasion at stage IA could be assimilated, in terms of prognosis and therapy, into stage I IMBTs.

Expression of Vascular Endothelial Growth Factor (VEGF), Hypoxia Inducible Factor-1α (HIF-1α), and Microvessel Density in Endometrial Tissue in Women With Adenomyosis

Adenomyosis is a disease with a mysterious pathogenesis, defined by an abnormal displacement of the eutopic endometrium deeply and haphazardly inside the myometrium. Angiogenesis has been indicated to play an important role and our aim was to investigate whether vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) expression and microvessel density (MVD) were different in women with and without adenomyosis. Immunohistochemistry was performed in endometrial tissues in 23 patients who underwent radical hysterectomy for adenomyosis (14) and for ovarian cysts and fibroids (9) at an Academic Hospital. Compared to women without the disease, VEGF expression was increased in endometrium with a normal location in patients with adenomyosis, although not associated to a significant increase of HIF-1α and MVD. Moreover, the endometrium with an abnormal location in patients with adenomyosis showed an increased VEGF and HIF-1α expression, particularly in the epithelial cells, associated to an increase of MVD, compared with the endometrium in a normal location in the same group of patients. Our present findings suggest that VEGF-mediated angiogenesis might be associated with the development of adenomyosis. In the ectopic foci the abnormal location might contribute to increased HIF-1a expression, stimulation of VEGF production, and increased vessel formation. In endometrium with a normal location, instead, where VEGF increased expression seems not to be correlated with HIF-1α increased expression nor with an increased MVD, other mechanisms might be reasonably postulated. Additional studies are required to explore new targeted and more effective treatment modalities.

Bone marrow histopathological and molecular changes of small B-cell lymphomas after rituximab therapy: comparison with clinical response and patients outcome.

This study correlates bone marrow changes after Rituximab (RTX) treatment with the clinical characteristics and outcome of 26 patients with small B-cell lymphomas. The percentage, phenotypic profile and clonality pattern of bone marrow lymphoid infiltrate were analysed before and after RTX treatment. Clinical, histological and molecular responses to RTX were correlated to the clinical outcome of the patients. Sixteen out of twenty-six patients obtained a complete clinical remission (CR). A favourable histology - follicular lymphoma (FL), hairy cell leukaemia (HCL) and marginal zone lymphoma (MZL) - was associated with a higher frequency of clinical CR and histological remission (HR), in comparison with mantle cell lymphoma (MCL), chronic lymphocytic leukaemia (CLL) and lymphoplasmacytic lymphoma (LPL). Two patterns of bone marrow HR were observed: 1) complete lymphoid cell disappearance (9 patients); or 2) nodular/interstitial T-cell infiltration (10 patients). Three histological persistence (HP) patterns were observed: 1) persistence of CD20+ small lymphoid cells in 1 patient with MCL; 2) loss of CD20 antigen expression in 4 patients with CLL; or 3) persistence only of clusters of monotypic plasma cells in 2 patients with LPL. CR and HR were strongly correlated. The percentage of lymphomatous infiltrate after RTX was higher in patients who subsequently died of the disease. Molecular response showed no correlations with the further clinical course in 12 patients achieving a complete clinical remission. In conclusion, bone marrow morphological and immunohistochemical analysis with a restricted panel of antibodies is useful to avoid 42% false positive and 85% false negative interpretations. Persistence of monoclonality after RTX might have a role in evaluating the molecular pattern of CD20-negative clones that can emerge after RTX as a tumoral escape to therapy.

HER-2/neu antigen loss in metastatic breast tumors—A pathological point of view

Dear Sir, Our study, such as other previous ones, demonstrated that in about 21% of the cases there is a discordance in the HER-2/neu status between primary breast cancer and paired metastases or local recurrences. This discordance may be explained by some biological phenomena, such as primary tumor heterogeneity and neoplastic genetic instability, more frequent as a consequence of adjuvant endocrine therapy and/or chemotherapy. In their very interesting Letter to the Editor, Manjili and coworkers, reporting their preclinical results, suggested that HER-2/neu antigen loss may be due to the immune pressure in vivo and in vitro. They pointed out the attention on the role of HER-2/neu specific IFN-g-secreting T cells in the induction of HER-2/neu antigen loss or down-regulation of HER-2/neu expression. They also demonstrated, with in vitro studies, that IL-6 inflammatory cytokine would upregulate the expression of IFN-g Ra in SKBR3 human breast tumor line. On the basis of these results, Manjili and coworkers suggested that IFN-g Ra positive tumor clones had lost HER-2/neu expression in the presence of HER-2/neu-specific immune responses, IFN-g-producing T cells in particular, and became metastatic, while IFN-g Ra negative tumor clones continued to express this antigen in primary tumor lesions. This hypothesis is very exciting also from a pathological point of view. The relation between immune system and breast tumor is evident in many cases as chronic inflammatory infiltrate around and between the neoplastic cells. A classical example is represented by medullary carcinoma in which an heavy lymphoplasmacytic infiltrate, with a predominance of T lymphocytes, is present around and between solid sheets of very atypical syncytial neoplastic cells (grade 3 according to Elston-Ellis). This type of tumor has a biological behavior less aggressive than a grade 3 infiltrating ductal carcinoma not otherwise specified (IDC). Another pathological example, that can support the results of Manjili and coworker, is represented by the evidence that many high grade in situ ductal carcinomas (DCIS) have a positive HER-2/neu status, whereas in the majority of the cases the associated IDCs have a negative one. If we consider that in many cases early stromal infiltration from a high grade DCIS is morphologically associated to an heavy lymphocytic infiltrate (mainly T lymphocytes) around the ducts and the microinvasive foci, may be quite easy to hypothesize a strong correlation between T cell products, inflammatory cytokines and HER-2/neu status change. In our study we demonstrated that in the 11.4% of cases, HER-2/neu positive primary tumors had HER-2/neu negative paired distant metastases; otherwise, in 17.2% of cases the opposite phenomenon was described. Similarly, HER-2/neu negative primary tumors had HER-2/neu positive local recurrences in 13.3% of the cases. Considering our results, the hypothesis of Manjili and coworkers may explain the 11.4% of HER-2/neu positive primary tumors (4 cases) in which the paired metachronous distant metastases (3 in the pleura and 1 in the retroperitoneum) were HER-2/neu negative. We revised microscopically these 4 primary tumors and the paired metastases and we found an evident inflammatory infiltrate, mainly composed by T lymphocytes, in 2 primary tumors and in all the 4 metastatic sites. In conclusion, the discordances between primary tumors and paired distant metastases or local recurrences regarding the HER-2/neu status may have some different possible biological explanations, one of the most interesting and stimulating being surely represented by the hypothesis advanced by Manjili and coworkers. Nevertheless, to confirm the results obtained by Manjili and coworkers on in vitro studies, it is necessary to study the expression of IFN-g Ra on tissue sections in an adequate number of cases that, for statistical reasons, must be necessarily greater than the four cases described in our article.

Gangliosides, sialoglycoproteins and glucocerebroside in the spleen and bone marrow of patients with β-thalassemia major

SummaryBone marrow and spleen samples were studied biochemically to elucidate the nature of the material stored in the large histiocytes present in β-thalassemic patients. Only the protein-bound N-acetylneuraminic acid (NANA) content of the spleen of homozygous β-thalassemic patients was significantly higher than that of controls. In the bone marrow, which contained a large number of thalassemic storage cells, the lipid-bound NANA and the glucocerebroside showed a moderate increase, while the amount of protein-bound NANA was much greater than that in controls. These results, in agreement with ultrastructural and histochemical observations, differentiate thalasseminc storage cells from other storage cells found in diseases characterized by an increased destruction of blood cells and suggest that there may also be an impairment of the sialoglycoprotein metabolism of the red blood cells in homozygous β-thalassemia.