Phylogenetic conservation of Trop-2 across species-rodent and primate genomics model anti-Trop-2 therapy for pre-clinical benchmarks.

A phylogenetic conservation analysis of Trop-2 across vertebrate species showed a high degree of sequence conservation, permitting to explore multiple models as pre-clinical benchmarks. Sequence divergence and incomplete conservation of expression patterns were observed in mouse and rat. Primate Trop-2 sequences were found to be 95%-100% identical to the human sequence. Comparative three-dimension primate Trop-2 structures were obtained with AlphaFold and homology modeling. This revealed high structure conservation of Trop-2 (0.66 ProMod3 GMQE, 0.80-0.86 ± 0.05 QMEANDisCo scores), with conservative amino acid changes at variant sites. Primate TACSTD2/TROP2 cDNAs were cloned and transfectants for individual ORF were shown to be efficiently recognized by humanized anti-Trop-2 monoclonal antibodies (Hu2G10, Hu2EF). Immunohistochemistry analysis of Macaca mulatta (rhesus monkey) tissues showed Trop-2 expression patterns that closely followed those in human tissues. This led us to test Trop-2 targeting in vivo in Macaca fascicularis (cynomolgus monkey). Intravenously injected Hu2G10 and Hu2EF were well tolerated from 5 to 10 mg/kg. Neither neurological, respiratory, digestive, urinary symptoms, nor biochemical or hematological toxicities were detected during 28-day observation. Blood serum pharmacokinetic (PK) studies were conducted utilizing anti-idiotypic antibodies in capture-ELISA assays. Hu2G10 (t1/2 = 6.5 days) and Hu2EF (t1/2 = 5.5 days) were stable in plasma, and were detectable in the circulation up to 3 weeks after the infusion. These findings validate primates as reliable models for Hu2G10 and Hu2EF toxicity and PK, and support the use of these antibodies as next-generation anti-Trop-2 immunotherapy tools.

Optical coherence tomography and color fundus photography in the screening of age-related macular degeneration: A comparative, population-based study.

PURPOSE: To analyze the individual value and the contribution of color fundus photography (CFP) and optical coherence tomography (OCT) in the screening of age-related macular degeneration (AMD) of an unselected population. METHODS: CFP and OCT images of 15957 eyes of 8069 subjects older than 55 years, obtained during a population-based screening for AMD using a single diagnostic non-mydriatic imaging device, were analyzed by a blinded examiner. The two techniques were preliminary evaluated considering the dichotomous parameter "gradable/ungradable", then gradable images were classified. CFP were graded according to the standardized classification of AMD lesions. OCT images were also categorized considering the presence of signs of early/intermediate AMD, late AMD, or other retinal diseases. Another blinded operator re-graded 1978 randomly selected images (for both CFP and OCT), to assess test reproducibility. RESULTS: Of the 15957 eyes, 8356 CFP (52.4%) and 15594 (97.7%) OCT scans were gradable. Moreover, most of the eyes with ungradable CFP (7339, 96.6%) were gradable at OCT. AMD signs were revealed in 7.4% of gradable CFP and in 10.4% of gradable OCT images. Moreover, at OCT, AMD signs were found in 1110 (6.9%) eyes whose CFP were ungradable or without AMD (847 and 263 eyes, respectively). The inter-operator agreement was good for the gradable versus ungradable parameter, and optimal for the AMD grading parameter of CFP. The agreement was optimal for all OCT parameters. CONCLUSIONS: OCT provided gradable images in almost all examined eyes, compared to limited CFP efficiency. Moreover, OCT images allowed to detect more AMD eyes compared to gradable photos. OCT imaging appears to significantly improve the power of AMD screening in a general, unselected population, compared to CFP alone.

Cell-block procedure in endoscopic ultrasound-guided-fine-needle-aspiration of gastrointestinal solid neoplastic lesions.

In the present review we have analyzed the clinical applications of endoscopic ultrasound-guided-fine-needle-aspiration (EUS-FNA) and the methodological aspects obtained by cell-block procedure (CBP) in the diagnostic approach to the gastrointestinal neoplastic pathology. CBP showed numerous advantages in comparison to the cytologic routine smears; in particular, better preservation of cell architecture, achievement of routine haematoxylin-eosin staining equivalent to histological slides and possibility to perform immunohistochemistry or molecular analyses represented the most evident reasons to choose this method. Moreover, by this approach, the differential diagnosis of solid gastrointestinal neoplasias may be more easily achieved and the background of contaminant non-neoplastic gastrointestinal avoided. Finally, biological samples collected by EUS-FNA CBP-assisted should be investigated in order to identify and quantify further potential molecular markers.

Endoscopic ultrasound-guided fine-needle aspiration cytology in pancreaticobiliary carcinomas: diagnostic efficacy of cell-block immunocytochemistry.

BACKGROUND: Endoscopic ultrasound-guided fine-needle aspiration cytology was demonstrated to be a useful tool for the diagnosis and staging of pancreaticobiliary neoplastic lesions. Nonetheless, the diagnostic value of this procedure may be limited by low cellularity of the specimen, contamination of intestinal cells and unfeasibility of ancillary immunocytochemical procedures. The present study was to evaluate its usefulness in the diagnosis of neoplastic lesions. METHODS: A series of 46 pancreaticobiliary carcinomas with available cell block preparations was submitted to immunocytochemistry against cytokeratins, carcinoembryonic antigen, E-cadherin, CD10 and p53. The sensitivity, specificity, positive and negative predictive values of the cytological smear in the discrimination of malignant lesions were calculated and compared with those of cell block preparation with the immunocytochemical stains against p53 and CD10. RESULTS: According to our findings, the use of cell block preparations together with immunostains against p53 and CD10 allowed to discriminate malignant versus benign specimens with higher sensitivity than the only cytological examination. In detail, CD10 immunostaining was of significant help for the discrimination between cytological contaminants, such as benign gastrointestinal cells, and the neoplastic elements of pancreaticobiliary well differentiated adenocarcinomas. Also, intense nuclear immunoreactivity for p53 was encountered in about 2/3 of the cases and identified pancreatic malignancy with high sensitivity. CONCLUSIONS: We suggest that immunocytochemistry against both CD10 and p53 could be applied case by case, mainly to differentiate gastrointestinal and pancreatic benign cellular contaminants showing hyperplasia or reactive changes from differentiated pancreaticobiliary adenocarcinomas.

Lymph node fine needle cytology in the diagnosis of infectious diseases and instrumental guides: ultrasound and computed tomography.

Imaging techniques, such as ultrasound imaging (US), computed tomography (CT), positron emission tomography-CT (PET-CT) or PET-magnetic resonance imaging (MRI), are highly accurate procedure in the lymph node enlargement detection, but none of them has the same sensitivity in the biological definition and in the cause of enlargement identification. Therefore, a direct evaluation of corresponding lymph nodes is necessary in much of the cases and Fine Needle Cytology (FNC) is one of the most frequently used technique for this purpose. The same imaging procedures are often used to perform targeted biopsies including FNC. This study discusses procedures, indications, advantages and limitations of imaging techniques as a support to FNC.

Fine needle cytology, infectious diseases and non-Hodgkin lymphoma.

The association between some infections and non-Hodgkin lymphomas (NHL) is well known. Human T-cell leukemia/lymphoma virus type 1 (HTLV-1) was the first oncogenic human retrovirus to be discovered and has been found to be associated with adult T-cell leukemia/lymphoma (ATLL). Epstein-Barr virus (EBV) has consistently been linked to both endemic and sporadic Burkitt lymphoma (BL), as well as to Hodgkin lymphoma (HL), post-transplantation proliferative disorders, extra-nodal NK-T-cell lymphoma (nasal type) and B-cell NHL arising in HIV patients; HCV infection is also associated to low-grade lymphoproliferative disorders that can progress to NHL. Bacterial infections have also been associated to NHL; chronic gastritis caused by Helicobacter pylori is responsible for mucosa-associated lymphoid tissue (MALT) NHL and high prevalence of Chlamydia psittaci infections has been reported in ocular adnexal lymphomas. In both these conditions, infection may contribute to the development of lymphomas, as proven by the clinical responses eradicating antibiotic therapies. Histological diagnosis coupled with immunohistochemical and molecular procedures are needed for a definitive diagnosis, but Fine Needle Cytology (FNC) combined with ancillary techniques can also produce correct diagnoses in most cases. In patients suffering from NHL, FNC also plays an important role in differential diagnosis between relapse of primary disease and reactive lymph nodes enlargement. This review explores the role of FNC in the diagnosis and classification of NHL trying to highlight possibilities and the limitations of the technique.

Detection and genotyping of human papillomavirus in gynaecologic outpatients of Messina, eastern Sicily, Italy.

In order to determine the prevalence of human papillomavirus (HPV) infection in sexually active female population in Messina, we tested cervical scrapes of women referred to university clinics for routine gynaecologic care. Between March and December 2008, a total of 680 cervical samples of 598 patients (573 Italian from province of Messina and 25 resident aliens) were examined consecutively from laboratory of molecular biology at the Department of Human Pathology. For each sample, cervical cells were collected by centrifugation and DNA was extracted (QIAamp DNA mini kit, Qiagen), followed by a PCR-based HPV DNA assay and reverse dot blot genotyping (HPV-HS Bio plus HPV-strip, AB Analytica or HPV-type, AB Analytica). The overall rate of HPV DNA detection in Italian patients (mean age 34 years; range 15-69) was 70.5% (404/573), with 163 cases of multiple infections (40.3%). In 335 patients (82.9%) a high-risk HPV infection was detected. In this group the coexistence of a low-risk HPV infection was documented in 97 cases while 65 patients exhibited only a low-risk HPV infection. HPV-16 was the most prevalent (33.4%), followed by HPV-6 (28.0%), HPV-31 (24.3%), HPV-58 (11.4%), HPV-66 (11.1%), HPV-53 (6.4%), HPV-18 (6.2%), HPV-56 (5.4%), HPV-33 (5.2%) while the other genotypes identified (HPV-11, -40, -42, -43, -44, -54, -61, -70, -81, -26, -35, -39, -45, -51, -52, -59, -68, -73, -82) were below 5%. HPV prevalence (any type) was 78.7% at age < or =24 years, 73.4% at 25-34 years and 67.1% at 35-44 years and 58.1% at age > or =45 years. A significant association (chi2=12.718; P=0.006) between HPV DNA detection and the younger age was encountered. Since available data on the prevalence and distribution of HPV infection in Italy are somewhat discordant, this study represents a helpful contribution to the knowledge on the circulation of precise genotypes in east Sicily in order to improve new HPV vaccines.

An unusual presentation of secondary involvement of B-cell chronic lymphocytic leukemia. A case report.

Extramammary tumors rarely metastasize to the breast. The commonest tumors to metastasize in breast tissue are lymphoproliferative diseases, melanoma, lung cancer and gynecological malignancies. Primary breast lymphoma has been reported in the literature with a maximum percentage of about 0.5% of all breast malignancies, while secondary localizations of lymphomas in the breast are less well studied in the literature than primary ones. The authors report a rare case of a secondary localization of B-cell chronic lymphocytic leukemia to the breast in which the diagnosis was obtained by histopathology and immunohistochemistry and further confirmed by molecular data. This occurrence must be considered in the differential diagnosis of a breast lump so that the primary hematological disease can be adequately treated and the correct type of breast surgery performed.

Lactoferrin immunoexpression in endometrial carcinomas: relationships with sex steroid hormone receptors (ER and PR), proliferation indices (Ki-67 and AgNOR) and survival.

We have investigated Lf immunoexpression as well as its biological meaning in 71 formalin-fixed, paraffin-embedded surgical samples of endometrial carcinomas (EC); 64 EC were endometrioid type, whereas 7 were non-endometrioid carcinomas. Immunohistochemistry was performed by primary antibodies against Lactoferrin (Lf), estrogen receptor (ER), progesterone receptor (PR) and Ki-67 antigen. Quantification of Lf immunoreactivity was performed using an intensity-distribution (ID) score. Moreover, the AgNOR technique according to guidelines of the Committee on AgNOR Quantification was used to assess the proliferation rate (NORA). A variable expression of Lf was revealed in 43 cases (61%) of EC. Endometrioid type carcinoma showed a significant higher Lf ID-score than non-endometrioid type; in contrast, no relationships were demonstrated between Lf immunoexpression and histologic grade, stage, clinical course as well as proliferative activity of EC. Moreover, a significantly higher Lf ID-score was encountered in ER-positive carcinomas. Survival analysis in EC indicated the architectural, nuclear and combined histologic grades as well as the stage, PR, Ki-67 and NORA as significant parameters. The utilization of Lf as a prognostic marker, able to identify patients at different risk of death, or alternatively, its clinical application as therapeutic agent, must be considered with great caution.