Optimization of patient selection for EGFR-TKIs in advanced non-small cell lung cancer by combined analysis of KRAS, PIK3CA, MET, and non-sensitizing EGFR mutations.

BACKGROUND: We present a comprehensive analysis of KRAS, PIK3CA, MET, and non-sensitizing EGFR mutations in advanced non-small cell lung cancer (NSCLC) patients treated with tyrosine kinase inhibitors (TKIs), with the aim of clarifying the relative contribution of these molecular alterations to resistance. PATIENTS AND METHODS: One hundred and sixty-six patients with advanced NSCLC treated with EGFR-TKIs with available archival tissue specimens were included. EGFR (exons 18-21), KRAS (exons 2, 3), PIK3CA (exons 9, 20), and MET (exons 14, 15) mutations were analyzed using PCR-based sequencing. Among all the mutations evaluated, only KRAS, PIK3CA, MET, and non-sensitizing EGFR mutations, defined as "TKI non-sensitizing mutations" were used for response, time to progression (TTP), and overall survival (OS) analysis. RESULTS: TKI non-sensitizing mutations were associated with disease progression (p = 0.001), shorter TTP (p < 0.0001), and worse OS (p = 0.03). Cox's multivariate analysis including histology and performance status showed that TKI non-sensitizing mutations were independent factors for shorter TTP (p < 0.0001) and worse OS (p = 0.01). CONCLUSIONS: When KRAS, PIK3CA, MET, and non-sensitizing EGFR mutations are concomitant, up to 96.0% of NSCLC patients unlikely to respond to TKIs can be identified, and they represented independent negative prognostic factors. Comprehensive molecular dissection of EGFR signaling pathways should be considered to select advanced NSCLC patients for TKIs therapies.

A useful ultrasound score to select thyroid nodules requiring fine needle aspiration in an iodine-deficient area.

BACKGROUND: The high prevalence of thyroid nodules in iodine-deficient areas is a practical problem because of the large number of patients requiring fine needle aspiration (FNA) to detect malignant nodules. AIM: To obtain an ultrasound (US) score for predicting malignant nodules and reduce the number of unnecessary and expensive FNA. SUBJECT AND METHOD: All nodules observed from September 2001 to March 2006 were evaluated by US: echostructure, echogenicity, halo, microcalcifications and ratio between antero-posterior and transversal diameters (AP/TR). Two thousand six hundred and forty-two consecutive patients underwent US-guided FNA for a total of 3645 nodules. RESULTS: Logistic regression analysis showed a potent predictive role for solitary nodules and absence/ incomplete halo (p=0.000). A significant predictive role for microcalcifications and AP/TR ratio was also observed. A 10-point score was constructed using the standardized regression coefficient. Nodules with US score or=5.5 had a frequency of malignancy of 0.4, 1.1 and 5.6% (p<0.001), respectively. Nodules with >or=5.5 US score were characterized by a 66% sensitivity and a 76% specificity compared to the diagnostic values of single parameters which were either sensitive or specific. CONCLUSIONS: According to our data, we suggest FNA for nodules reaching a >5.4 US score, whereas a clinical judgement should be used for the intermediate category nodules. When the score is lower than 2.5 we do not recommend FNA. The practical use of this US score can help reduce unnecessary and expensive FNA in iodine-deficient areas.

Fine needle aspiration cytology of thyroid nodules: conventional vs thin layer technique.

OBJECTIVE: Liquid-based cytology using the thin layer technique has recently been introduced in thyroid fine needle aspiration cytology together with or in substitution of direct smears, but its usefulness is still controversial and relatively few studies have been published in this field. The aim of the present study was to compare the results obtained from conventional smears with those from thin layer smears. DESIGN: In 3875 thyroid nodules, a double cytologic sampling was taken in randomized order, to prepare conventional or thin layer smears. MAIN OUTCOME: The diagnoses agreed in 2934 (75.7%) cases and disagreed in 941 (24.3%). The analysis of discordant data showed there were fewer non-diagnostic cases in the thin layer smears (377 vs 541, p<0.001) whereas in conventional smears there were more cases positive for carcinoma (27 vs 4, p<0.001). The cytohistologic correlation was available for 194 cases and showed that conventional smears had a greater capacity for revealing carcinomas (44 vs 31). Finally, diagnoses based on conventional smears were more sensitive than thin layer smears (93.6% vs 65.9%) whereas specificity was constant. CONCLUSIONS: From our experience, the conventional smear offers a greater possibility of diagnosis when suspecting malignancy or diagnosing malignancy cases, whereas thin layer smears significantly reduce the number of non-diagnostic cases. For this reason, we suggest combining the two techniques in routine cytologic diagnosis.

Intraoperative autologous blood recovery in prostate cancer surgery: in vivo validation using a tumour marker.

BACKGROUND AND OBJECTIVE: Intraoperative autologous blood recovery during radical retro pubic prostatectomy has the potential of contamination with tumour cells. Its safety is proved by similar survival rates between allogeneic and autologous transfusion to oncology patients without standardization. Silencing of the gene encoding pi class of gluthatione-S transferase is a specific and sensitive molecular marker for prostate cancer, because it is present in more than 90% of prostate tumours. Using such tumour marker, we aimed to demonstrate that viable tumour cells could be eliminated using leucodepletion filters followed by irradiation. MATERIALS AND METHODS: Fifty patients with pi class of gluthatione-S transferase promoter hypermethylation in their primary prostate tumours were included in the analysis. Peripheral blood samples were collected during anaesthetic induction and recovered blood was collected throughout the surgery and then submitted to washing, leucoreduction and irradiation. Samples were analysed stepwise for the presence of promoter hypermethylation using real-time methylation-specific polymerase chain reaction. RESULTS: Positive hypermethylation was found in recovered blood (two samples), recovered and washed blood (three samples), and recovered washed and filtered blood (two samples). After filtration and irradiation of the recovered blood, this marker could not be detected in any of the cases analysed, suggesting the absence of viable tumour cells. CONCLUSION: Even though the risk of disseminating tumour cells in prostate cancer surgery by intraoperative autologous blood recovery is not yet fully established, no tumour-specific gene amplification was found after the association of blood filtration and irradiation, suggesting a significant reduction of such risk.

Determination of the mechanism for resonant scattering in LaMnO3.

The resonant multiple Bragg x-ray diffraction is used to study the forbidden (104) reflection in LaMnO3. Using the interference between the three-beam scattering and resonant scattering we can determine the phase of the resonant scattering. This phase is shown to be consistent with a model in which the resonant scattering is caused by the influence of the Mn-O bond length distortion rather than directly by the orbital ordering on the Mn 4p band structure.

[Cutaneous angiomyolipoma: a case report and literature review]. / Angiomiolipoma cutaneo: descrizione di un caso e revisione della letteratura.

Angiomyolipoma is a lesion usually observed in the kidney of patients with tuberous sclerosis. Extrarenal sites are very unusual with sporadic cases in internal organ, soft tissue and skin (fifteen cases have been described in this site). Herein we describe an adding case located on the ear in 58-year-old man reviewing the pertinent literature. The main diagnostic differential criteria are also discussed.

Matrigel invasion by the prostate cancer cell lines, PC3 and DU145, and cathepsin L+B activity.

Cathepsins L and B are lysosomal cysteine proteinases whose activities and cellular location are altered in many types of cancers and cancer cell lines. Cathepsins L and B play an unspecified role in cancer invasion and metastasis. The purpose of our study was to determine whether cathepsins L and B are important for the ability of two prostate cancer cell lines, PC3 and DU 145, to invade the basement membrane-like preparation, Matrigel. Exposure of PC3 and DU145 to the irreversible cysteine proteinase inhibitor, E64, decreases the invasive ability of DU145, but not PC3. PC3 and DU145 were treated with the phorbol ester analogue, phorbol 12-myristate 13-acetate (PMA), a known tumor promoter that activates protein kinase C and contributes to the metastatic phenotype. PMA increased secreted cathepsin L+B activity and the invasive ability of PC3 and DU145; co-exposure to E64 and PMA decreased both cathepsin L+B activity and invasion. We conclude that DU145 requires cathepsin L+B activity more than PC3 for the invasion of the Matrigel. When the amount of secreted cathepsin L+B activity is increased by PMA treatment, however, PC3 becomes dependent on cathepsin L+B for invasion. Our study demonstrates that modulation of the amount of secreted cathepsin L+B activity influences the invasive phenotype of PC3 and DU145.

Molecular evidence of tumour cell removal from salvaged blood after irradiation and leucocyte depletion.

Intra-operative autologous blood recovery offers many advantages. However, blood salvage during cancer surgery is of limited use due to the potential presence of circulating tumour cells. It was the aim of this study to show that intra-operative salvage blood can be freed of cells and cellular DNA after leucoreduction by filtration and irradiation of washed blood. Known amounts of tissue culture derived from carcinoma, melanoma and osteosarcoma were added to whole blood bags. This mixture was then submitted to washing, leucoreduction and irradiation. Samples were studied stepwise in relation to the integrity and size of DNA by the polymerase chain reaction (PCR). After filtration and irradiation, PCR targeting the beta-globin gene (268 bp amplicon) was negative. Our results were corroborated by studying plasma samples added with tumoural cells. Using PCR methodology, we showed the absence of DNA from cells in experimentally contaminated blood and plasma bags after filtration and irradiation. This experimental study is an effort to ensure the safety of intra-operative autologous transfusion.

Decreased activity of cathepsins L + B and decreased invasive ability of PC3 prostate cancer cells.

Cancer metastasis involves multiple factors, one of which is the production and secretion of matrix degrading proteases by the cancer cells. Many metastasizing cancer cells secrete the lysosomal proteases, cathepsins L and B, which implicates them in the metastatic process. Cathepsins L and B are regulated by endogenous cysteine proteinase inhibitors (CPI) known as cystatins. An imbalance between cathepsin L and/or B and cystatin expression/activity may be a characteristic of the metastatic phenotype. To determine whether cystatins can attenuate the invasive ability of PC3 prostate cancer cells, cells were transfected with a cDNA coding for chicken cystatin. Expression of chicken cystatin mRNA was determined by PCR analysis. Total cysteine proteinase inhibitory activity, cathepsins L + B activity, and invasion through a Matrigel matrix were assessed. Stably transfected cells expressed the chicken cystatin mRNA and exhibited a significant decrease in secreted cathepsin L + B activity and a small increase in secreted cysteine proteinase inhibitor activity. The ability of cystatin transfected cells to invade the reconstituted basement membrane, Matrigel, was attenuated compared to nontransfected cells or cells transfected with vector alone. We have demonstrated that the cysteine proteinases cathepsins L and B participate in the invasive ability of the PC3 prostate cancer cell line, and we discuss here the potential of using cysteine proteinase inhibitors such as the cystatins as anti-metastatic agents.

Absolute structure-factor measurements of an Al-Pd-Mn quasicrystal.

A number of X-ray reflections from an icosahedral quasicrystal Al-Pd-Mn have been measured with great accuracy on an absolute basis by making use of Bragg-case diffraction. Since the specimen had high crystal quality, the dynamical theory was used for analyzing the results and to extract structure factors from measured integrated intensities. Good agreement was found between theory and experiment for strong reflections. Anomalous transmission was found to be strong in the 'good' regions of the quasicrystalline specimen and it was measured on an absolute basis, but the small residual strains present in the specimen prevented an accurate comparison between theory and experiment. A detailed discussion is presented on the parameters that mostly affect anomalous transmission in relationship to the adopted structural model.