A phase I dose-escalation study of enzalutamide in combination with the AKT inhibitor AZD5363 (capivasertib) in patients with metastatic castration-resistant prostate cancer.

BACKGROUND: Activation of the PI3K/AKT/mTOR pathway through loss of phosphatase and tensin homolog (PTEN) occurs in approximately 50% of patients with metastatic castration-resistant prostate cancer (mCRPC). Recent evidence suggests that combined inhibition of the androgen receptor (AR) and AKT may be beneficial in mCRPC with PTEN loss. PATIENTS AND METHODS: mCRPC patients who previously failed abiraterone and/or enzalutamide, received escalating doses of AZD5363 (capivasertib) starting at 320 mg twice daily (b.i.d.) given 4 days on and 3 days off, in combination with enzalutamide 160 mg daily. The co-primary endpoints were safety/tolerability and determining the maximum tolerated dose and recommended phase II dose; pharmacokinetics, antitumour activity, and exploratory biomarker analysis were also evaluated. RESULTS: Sixteen patients were enrolled, 15 received study treatment and 13 were assessable for dose-limiting toxicities (DLTs). Patients were treated at 320, 400, and 480 mg b.i.d. dose levels of capivasertib. The recommended phase II dose identified for capivasertib was 400 mg b.i.d. with 1/6 patients experiencing a DLT (maculopapular rash) at this level. The most common grade ≥3 adverse events were hyperglycemia (26.7%) and rash (20%). Concomitant administration of enzalutamide significantly decreased plasma exposure of capivasertib, though this did not appear to impact pharmacodynamics. Three patients met the criteria for response (defined as prostate-specific antigen decline ≥50%, circulating tumour cell conversion, and/or radiological response). Responses were seen in patients with PTEN loss or activating mutations in AKT, low or absent AR-V7 expression, as well as those with an increase in phosphorylated extracellular signal-regulated kinase (pERK) in post-exposure samples. CONCLUSIONS: The combination of capivasertib and enzalutamide is tolerable and has antitumour activity, with all responding patients harbouring aberrations in the PI3K/AKT/mTOR pathway. CLINICAL TRIAL NUMBER: NCT02525068.

Application of Liquid Biopsies in Cancer Targeted Therapy.

As a growing body of evidence demonstrates intertumoral and intratumoral heterogeneity and clonal evolution, both during carcinogenesis and also throughout treatment resulting in acquired drug resistance, the utility of blood-based assays or "liquid biopsies" is becoming increasingly recognized in clinical practice and trial design. "Liquid biopsies" provide a less invasive approach to the current gold standard of interrogating tumors by tissue biopsies, which are frequently unfeasible, associated with morbidity, and cannot be performed as often.

Analysis of DNA methylation in single circulating tumor cells.

Direct analysis of circulating tumor cells (CTCs) can inform on molecular mechanisms underlying systemic spread. Here we investigated promoter methylation of three genes regulating epithelial-to-mesenchymal transition (EMT), a key mechanism enabling epithelial tumor cells to disseminate and metastasize. For this, we developed a single-cell protocol based on agarose-embedded bisulfite treatment, which allows investigating DNA methylation of multiple loci via a multiplex PCR (multiplexed-scAEBS). We established our assay for the simultaneous analysis of three EMT-associated genes miR-200c/141, miR-200b/a/429 and CDH1 in single cells. The assay was validated in solitary cells of GM14667, MDA-MB-231 and MCF-7 cell lines, achieving a DNA amplification efficiency of 70% with methylation patterns identical to the respective bulk DNA. Then we applied multiplexed-scAEBS to 159 single CTCs from 11 patients with metastatic breast and six with metastatic castration-resistant prostate cancer, isolated via CellSearch (EpCAMpos/CKpos/CD45neg/DAPIpos) and subsequent FACS sorting. In contrast to CD45pos white blood cells isolated and processed by the identical approach, we observed in the isolated CTCs methylation patterns resembling more those of epithelial-like cells. Methylation at the promoter of microRNA-200 family was significantly higher in prostate CTCs. Data from our single-cell analysis revealed an epigenetic heterogeneity among CTCs and indicates tumor-specific active epigenetic regulation of EMT-associated genes during blood-borne dissemination.

Preclinical medical student observations associated with later professionalism concerns.

PURPOSE: Professionalism is a core physician competency and identifying students at risk for poor professional development early in their careers may allow for mentoring. This study identified indicators in the preclinical years associated with later professionalism concerns. METHODS: A retrospective analysis of observable indicators in the preclinical and clinical years was conducted using two classes of students (n = 226). Relationships between five potential indicators of poor professionalism in the preclinical years and observations related to professional concerns in the clinical years were analyzed. RESULTS: Fifty-three medical students were identified with at least one preclinical indicator and one professionalism concern during the clinical years. Two observable preclinical indicators were significantly correlated with unprofessional conduct during the clinical years: Three or more absences from attendance-required sessions (odds ratio 4.47; p=.006) and negative peer assessment (odds ratio 3.35; p=.049). CONCLUSIONS: We identified two significant observable preclinical indicators associated with later professionalism concerns: excessive absences and negative peer assessments. Early recognition of students at risk for future professionalism struggles would provide an opportunity for proactive professional development prior to the clinical years, when students' permanent records may be affected. Peer assessment, coupled with attention to frequent absences, may be a method to provide early recognition.

Identification of novel determinants of resistance to lapatinib in ERBB2-amplified cancers.

The gene encoding the receptor tyrosine kinase ERBB2, also known as HER2, is amplified and/or overexpressed in up to 15% of breast cancers. These tumours are characterised by an aggressive phenotype and poor clinical outcome. Although therapies targeted at ERBB2 have proven effective, many patients fail to respond to treatment or become resistant and the reasons for this are still largely unknown. Using a high-throughput functional screen we assessed whether genes found to be recurrently amplified and overexpressed in ERBB2+ve breast cancers mediate resistance to the ERBB2-targeted agent lapatinib. Lapatinib-resistant ERBB2-amplified breast cancer cell lines were screened, in the presence or absence of lapatinib, with an RNA interference library targeting 369 genes recurrently amplified and overexpressed in both ERBB2-amplified breast cancer tumours and cell lines. Small interfering RNAs targeting a number of genes caused sensitivity to lapatinib in this context. The mechanisms of resistance conferred by the identified genes were further investigated and in the case of NIBP (TRAPPC9), lapatinib resistance was found to be mediated through NF-κB signalling. Our results indicate that specific amplified and/ or overexpressed genes found in ERBB2-amplified breast cancer may mediate response to ERBB2-targeting agents.

Integrative molecular and functional profiling of ERBB2-amplified breast cancers identifies new genetic dependencies.

Overexpression of the receptor tyrosine kinase ERBB2 (also known as HER2) occurs in around 15% of breast cancers and is driven by amplification of the ERBB2 gene. ERBB2 amplification is a marker of poor prognosis, and although anti-ERBB2-targeted therapies have shown significant clinical benefit, de novo and acquired resistance remains an important problem. Genomic profiling has demonstrated that ERBB2+ve breast cancers are distinguished from ER+ve and 'triple-negative' breast cancers by harbouring not only the ERBB2 amplification on 17q12, but also a number of co-amplified genes on 17q12 and amplification events on other chromosomes. Some of these genes may have important roles in influencing clinical outcome, and could represent genetic dependencies in ERBB2+ve cancers and therefore potential therapeutic targets. Here, we describe an integrated genomic, gene expression and functional analysis to determine whether the genes present within amplicons are critical for the survival of ERBB2+ve breast tumour cells. We show that only a fraction of the ERBB2-amplified breast tumour lines are truly addicted to the ERBB2 oncogene at the mRNA level and display a heterogeneous set of additional genetic dependencies. These include an addiction to the transcription factor gene TFAP2C when it is amplified and overexpressed, suggesting that TFAP2C represents a genetic dependency in some ERBB2+ve breast cancer cells.

Assessing the hemodynamic influence between multiple lesions in a realistic right coronary artery segment: A computational study.

Coronary artery disease is the primary cause of morbidity and mortality worldwide. Therefore, detailed assessment of lesions in the coronary vasculature is critical in current clinical practice. Fractional flow reserve (FFR) has been proven as an efficient method for assessing the hemodynamic severity of a coronary stenosis. However, functional assessment of a coronary segment with multiple stenoses (≥ 2) remains complex for guiding the strategy of percutaneous coronary intervention due to the hemodynamic interplay between adjacent stenoses. In this work, we created four 3-dimensional (3D) arterial models that derive from a healthy patient-specific right coronary artery segment. The initial healthy model was reconstructed using fusion of intravascular ultrasound (IVUS) and biplane angiographic patient data. The healthy 3D model presented a measured FFR value of 0.96 (pressure-wire) and a simulated FFR value of 0.98. We then created diseased models with two artificial sequential stenoses of 90% lumen area reduction or with the proximal and distal stenosis separately. We calculated the FFR value for each case: 0.65 for the case with the two stenoses, 0.73 for the case with the distal stenosis and 0.90 for the case with the proximal stenosis. This leads to the conclusion that although both stenoses had the same degree of lumen area stenosis, there was a large difference in hemodynamic severity, thereby indicating that angiographic lumen assessment by itself is often not adequate for accurate assessment of coronary lesions.

Computational assessment of the fractional flow reserve from intravascular ultrasound and coronary angiography data: a pilot study.

Cardiovascular disease is one of the primary causes of morbidity and mortality around the globe. Thus, the diagnosis of critical lesions in coronary arteries is of utmost importance in clinical practice. One useful and efficient method to assess the functional severity of one or multiple lesions in a coronary artery is the calculation of the fractional flow reserve (FFR). In the current work, we present a method which allows the calculation of the FFR value computationally, without the use of a pressure wire and the induction of hyperemia, using intravascular ultrasound (IVUS) and biplane angiography images for three-dimensional (3D) coronary artery reconstruction and measurements of the volumetric flow rate derived from angiographic sequences. The simulated FFR values were compared to the invasively measured FFR values in 7 cases, presenting high correlation (r=0.85) and good agreement (mean difference=0.002). FFR assessment without employing a pressure wire and the induction of hyperemia is feasible using 3D reconstructed coronary artery models from angiographic and IVUS data coupled with computational fluid dynamics.

Integrative molecular profiling of triple negative breast cancers identifies amplicon drivers and potential therapeutic targets.

Triple negative breast cancers (TNBCs) have a relatively poor prognosis and cannot be effectively treated with current targeted therapies. We searched for genes that have the potential to be therapeutic targets by identifying genes consistently overexpressed when amplified. Fifty-six TNBCs were subjected to high-resolution microarray-based comparative genomic hybridization (aCGH), of which 24 were subjected to genome-wide gene expression analysis. TNBCs were genetically heterogeneous; no individual focal amplification was present at high frequency, although 78.6% of TNBCs harboured at least one focal amplification. Integration of aCGH and expression data revealed 40 genes significantly overexpressed when amplified, including the known oncogenes and potential therapeutic targets, FGFR2 (10q26.3), BUB3 (10q26.3), RAB20 (13q34), PKN1 (19p13.12) and NOTCH3 (19p13.12). We identified two TNBC cell lines with FGFR2 amplification, which both had constitutive activation of FGFR2. Amplified cell lines were highly sensitive to FGFR inhibitor PD173074, and to RNAi silencing of FGFR2. Treatment with PD173074 induced apoptosis resulting partly from inhibition of PI3K-AKT signalling. Independent validation using publicly available aCGH data sets revealed FGFR2 gene was amplified in 4% (6/165) of TNBC, but not in other subtypes (0/214, P=0.0065). Our analysis demonstrates that TNBCs are heterogeneous tumours with amplifications of FGFR2 in a subgroup of tumours.

Expression, mutation and copy number analysis of platelet-derived growth factor receptor A (PDGFRA) and its ligand PDGFA in gliomas.

BACKGROUND: Malignant gliomas are the most prevalent type of primary brain tumours but the therapeutic armamentarium for these tumours is limited. Platelet-derived growth factor (PDGF) signalling has been shown to be a key regulator of glioma development. Clinical trials evaluating the efficacy of anti-PDGFRA therapies on gliomas are ongoing. In this study, we intended to analyse the expression of PDGFA and its receptor PDGFRA, as well as the underlying genetic (mutations and amplification) mechanisms driving their expression in a large series of human gliomas. METHODS: PDGFA and PDGFRA expression was evaluated by immunohistochemistry in a series of 160 gliomas of distinct World Health Organization (WHO) malignancy grade. PDGFRA-activating gene mutations (exons 12, 18 and 23) were assessed in a subset of 86 cases by PCR-single-strand conformational polymorphism (PCR-SSCP), followed by direct sequencing. PDGFRA gene amplification analysis was performed in 57 cases by quantitative real-time PCR (QPCR) and further validated in a subset of cases by chromogenic in situ hybridisation (CISH) and microarray-based comparative genomic hybridisation (aCGH). RESULTS: PDGFA and PDGFRA expression was found in 81.2% (130 out of 160) and 29.6% (48 out of 160) of gliomas, respectively. Its expression was significantly correlated with histological type of the tumours; however, no significant association between the expression of the ligand and its receptor was observed. The absence of PDGFA expression was significantly associated with the age of patients and with poor prognosis. Although PDGFRA gene-activating mutations were not found, PDGFRA gene amplification was observed in 21.1% (12 out of 57) of gliomas. No association was found between the presence of PDGFRA gene amplification and expression, excepting for grade II diffuse astrocytomas. CONCLUSION: The concurrent expression of PDGFA and PDGFRA in different subtypes of gliomas, reinforce the recognised significance of this signalling pathway in gliomas. PDGFRA gene amplification rather than gene mutation may be the underlying genetic mechanism driving PDGFRA overexpression in a portion of gliomas. Taken together, our results could provide in the future a molecular basis for PDGFRA-targeted therapies in gliomas.

Genomic profile of a secretory breast cancer with an ETV6-NTRK3 duplication.

BACKGROUND: Secretory breast cancer (SBC) is a rare entity characterised by indolent clinical behaviour, distinctive histological features and the presence of a recurrent chromosomal translocation t(12;15)(p13;q25), leading to the formation of the ETV6-NTRK3 fusion gene. AIM: To describe the molecular genetic features of a case of SBC which harbours a duplication of the t(12;15) translocation. METHODS: Tiling path array comparative genomic hybridisation (aCGH) analysis and fluorescence in situ hybridisation (FISH) using in-house-generated probes for ETV6, NTRK3 and the fusion genes, centromeric probes for chromosomes 12 and 15, and a commercially available split-apart ETV6/NTRK3 probe. RESULTS: FISH revealed the presence of a duplication of the translocation t(12;15), which resulted from the gain of one copy of the derivative chromosome der(15)t(12;15), retention of one normal copy of both ETV6 and NTRK3 genes and deletion of the derivative chromosome der(12)t(12;15). Consistent with FISH findings, aCGH revealed copy number gains of ETV6 and NTRK3 and deletions encompassing the regions centromeric to ETV6 and telomeric to NTRK3. Additional regions of copy number changes included gains of 10q21, 10q26.3, 12p13.3-p13.31 15q11-q25.3 and 16pq and losses of 6q24.1-q27, 12p13.2-q12 and 15q25.3-q26.3. CONCLUSIONS: To the best of our knowledge, this is the first time a carcinoma has been shown to harbour a duplication of the ETV6-NTRK3 translocation. The presence of an additional copy of the derivative chromosome der(15)t(12;15) coupled with deletion of the other derivative der(12)t(12;15) in the modal population of cancer cells suggests that this was either an early phenomenon or conferred additional growth advantage on neoplastic cells.

ESR1 amplification in endometrial carcinomas: hope or hyperbole?

The ESR1 gene maps 6q25 and encodes for oestrogen receptor alpha, which has been shown to play a pivotal role in the development of breast and endometrial cancer. It has recently been reported that oestrogen receptor alpha expression may be driven in some cases by ESR1 gene amplification and that this phenomenon may be an early event in breast and endometrial carcinogenesis. Although copy number gains of 6q have been reported by several groups, their prevalence, association with oestrogen receptor alpha expression, and clinical implications have been a matter of controversy. Here we discuss the key issues regarding the methods employed in the identification of ESR1 amplification, and briefly review the current literature and recent controversies on the subject of ESR1 amplification in endometrial and breast cancers.

The genomic profile of HER2-amplified breast cancers: the influence of ER status.

Expression profiling studies have suggested that HER2-amplified breast cancers constitute a heterogeneous group that may be subdivided according to their ER status: HER2-amplified ER-positive breast carcinomas that fall into the luminal B cluster; and HER2-amplified ER-negative cancers which form a distinct molecular subgroup, known as the erbB2 or HER2 subgroup. ER-negative breast cancer differs significantly from ER-positive disease in the pattern, type, and complexity of genetic aberrations. Here we have compared the genomic profiles of ER-positive and ER-negative HER2-amplified cancers using tiling path microarray-based comparative genomic hybridization (aCGH). Validation of the differentially amplified regions was performed in an independent series of 70 HER2-amplified breast cancers. Although HER2-amplified cancers had remarkably complex patterns of molecular genetic aberrations, ER-positive and ER-negative HER2-amplified breast carcinomas shared most molecular genetic features as defined by aCGH. Genome-wide Fisher's exact test analysis revealed that less than 1.5% of the genome was significantly differentially gained or lost in ER-positive versus ER-negative HER2-amplified cancers. However, two regions of amplification were significantly associated with ER-positive carcinomas, one of which mapped to 17q21.2 and encompassed GJC1, IGFBP4, TNS4, and TOP2A. Chromogenic in situ hybridization analysis of an independent validation series confirmed the association between ER status and TOP2A amplification. In conclusion, although hormone receptor status does not determine the overall genetic profile of HER2-amplified breast cancers, specific genetic aberrations may be characteristic of subgroups of HER2 breast cancers.

Molecular profiling pleomorphic lobular carcinomas of the breast: evidence for a common molecular genetic pathway with classic lobular carcinomas.

Pleomorphic lobular carcinomas (PLC) of the breast display histological features associated with classic invasive lobular carcinoma (ILC), yet they also exhibit more conspicuous nuclear atypia and pleomorphism, and an aggressive clinical behaviour. From a breast cancer progression perspective, it is unclear whether PLC is a variant of ILC or is a high-grade invasive ductal carcinoma (IDC) that has lost E-cadherin. The molecular features of 26 PLC were studied using immunohistochemistry [oestrogen receptor (ER), progesterone receptor (PR), HER2, p53 and E-cadherin], 0.9 Mb resolution, microarray-based comparative genomic hybridization (aCGH), fluorescent (FISH) and chromogenic (CISH) in situ hybridization and loss of heterozygosity. Comparative analysis was performed with aCGH data from PLC with classic ILC (16 cases) and high grade IDC (35 cases). PLCs were frequently ER- and PR-positive, E-cadherin-negative and occasionally HER2- and p53-positive. Recurrent copy number changes identified by aCGH included gains on 1q, 8q, 11q, 12q, 16p and 17q and losses on 8p, 11q, 13q, 16q and Xq. Highly recurrent 1q+ (100% of cases), 16p+ (93%), 11q- (53%) and 16q- (93%) and evidence of the der(1;16)/der(16)t(1;16) rearrangement, as detected by FISH, suggested that PLC had a 'lobular genotype'. Focal amplifications were evident at 8p12-p11, 8q24, 11q13.1-q14.1, 12q14, 17q12 and 20q13. Loss of BRCA2 was detected in 40% of PLC by LOH. Comparative analysis of aCGH data suggested the molecular features of PLC (ER/PR-positive, E-cadherin-negative, 1q+, 11q(-), 16p+ and 16q(-)) were more closely related to those of ILC than IDC, implicating an overlapping developmental pathway for these lobular tumour types. Molecular alterations found in PLC that are more typical of high-grade IDC than ILC (p53 and HER2 positivity, 8q+, 17q24-q25+, 13q(-) and amplification of 8q24, 12q14, 17q12 and 20q13) are likely to drive the high-grade and more aggressive biology of PLC.

Is acinic cell carcinoma a variant of secretory carcinoma? A FISH study using ETV6'split apart' probes.

AIMS: Acinic cell carcinomas (ACCs) and secretory carcinomas (SCs) of the breast are rare, low-grade malignancies that preferentially affect young female patients. Owing to the morphological and immunohistochemical similarities between these lesions, they have been proposed to be two morphological variants of the same entity. It has been demonstrated that SCs of the breast consistently harbour the t(12;15)ETV6-NTRK3 translocation. The aim was to determine whether ACCs also harbour ETV6 gene rearrangements and are thus variants of SCs. METHODS AND RESULTS: Using the ETV6 fluorescence in situ hybridization DNA Probe Split Signal (Dako), the presence of ETV6 rearrangements in three SCs and six ACCs was investigated. Cases were considered as harbouring an ETV6 gene rearrangement if >10% of nuclei displayed 'split apart signals' (i.e. red and green signals were separated by a distance greater than the size of two hybridization signals). Whereas the three SCs displayed ETV6 split apart signals in >10% of the neoplastic cells, no ACC showed any definite evidence of ETV6 gene rearrangement. CONCLUSIONS: Based on the lack of ETV6 rearrangements in ACCs, our results strongly support the concept that SCs and ACCs are distinct entities and should be recorded separately in breast cancer taxonomy schemes.

Comparative analysis of six different antibodies against Her2 including the novel rabbit monoclonal antibody (SP3) and chromogenic in situ hybridisation in breast carcinomas.

AIMS: To compare the sensitivity and specificity of new rabbit monoclonal antibody SP3 with those of mouse monoclonal and rabbit polyclonal antibodies using HER2 amplification defined by chromogenic in situ hybridisation (CISH) as the gold standard. METHODS: Serial sections from tissue microarrays (TMAs) containing 84 breast carcinomas were submitted to CISH (Zymed HER2 Spot-Light kit) and immunohistochemistry, using NeoMarkers SP3 (rabbit monoclonal), DAKO A0485 and DAKO HercepTest (polyclonal), Novocastra NCL-CB11, Cell Marque CM-CB11, and Genentech 4D5 (mouse monoclonal). RESULTS: The best antibody concordance was between SP3 and HercepTest (kappa = 0.74). SP3, A0485 and HercepTest detected all HER2 amplified tumours, but were less specific than mouse monoclonal antibodies. 3/38 (7.9%) and 8/38 (21.0%) non-amplified tumours were scored as 3+ using SP3 and A0485, respectively. 3/46 (6.5%) amplified tumours were negative for NCL-CB11. SP3, HercepTest and A0485 showed no gene amplification on 55%, 62.5% and 92.3% of the 2+ scored tumours, but most of the 2+ scored tumours using monoclonal antibodies were amplified by CISH (80-92.3%). CONCLUSIONS: SP3 is more sensitive than mouse monoclonal antibodies for Her2 assessment. However, HercepTest, CB11 and 4D5 show higher specificity than SP3 for the identification of HER2 gene amplification. Mouse monoclonal antibodies show less Her2 2+ tumours; most are amplified by CISH.

Genomic and immunophenotypical characterization of pure micropapillary carcinomas of the breast.

Pure invasive micropapillary carcinoma (MPC) is a special histological type that accounts for 0.7-3% of all breast cancers. MPC has a distinctive growth pattern and a more aggressive clinical behaviour than invasive ductal carcinomas of no special type (IDC-NSTs). To define the molecular characteristics of MPCs, we profiled a series of 12 MPCs and 24 grade and oestrogen receptor (ER)-matched IDC-NSTs using high-resolution microarray comparative genomic hybridization (aCGH). In addition, we generated a tissue microarray containing a series of 24 MPCs and performed immunohistochemical analysis with ER, PR, Ki-67, HER2, CK5/6, CK14, CK17, EGFR, topoisomerase-IIalpha, cyclin D1, caveolin-1, E-cadherin, and beta-catenin antibodies. In situ hybridization probes were employed to evaluate the prevalence of amplification of HER2, TOP2A, EGFR, CCND1, MYC, ESR1, and FGFR1 genes. aCGH analysis demonstrated that MPCs significantly differed from IDC-NSTs at the genomic level. Gains of 1q, 2q, 4p, 6p, 6q23.2-q27, 7p, 7q, 8p, 8q, 9p, 10p, 11q, 12p, 12q, 16p, 17p, 17q, 19p, 20p, 20q, and 21q, and losses of 1p, 2p, 6q11.1-q16.3, 6q21-q22.1, 9p, 11p, 15q, and 19q were more prevalent in MPCs. High-level gains/amplifications of 8p12-p11, 8q12, 8q13, 8q21, 8q23, 8q24, 17q21, 17q23, and 20q13 were significantly associated with MPCs. A comparison between 24 MPCs and a series of 48 grade and ER-matched IDC-NSTs revealed that high cyclin D1 expression, high proliferation rates, and MYC (8q24) amplification were significantly associated with MPCs. Our results demonstrate that MPCs have distinct histological features and molecular genetic profiles supporting the contention that they constitute a distinct pathological entity.

EGFR amplification and lack of activating mutations in metaplastic breast carcinomas.

Metaplastic breast carcinomas are reported to harbour epidermal growth factor receptor (EGFR) overexpression in up to 80% of the cases, but EGFR gene amplification is the underlying genetic mechanism in around one-third of these. In this study, EGFR gene amplification as defined by chromogenic in situ hybridization and protein overexpression was examined in a cohort of 47 metaplastic breast carcinomas. Furthermore, the presence of activating EGFR mutations in exons 18, 19, 20, and 21 was investigated. Thirty-two cases showed EGFR overexpression and of these, 11 (34%) harboured EGFR gene amplification. In addition, EGFR amplification showed a statistically significant association with EGFR overexpression (p < 0.0094) and was restricted to carcinomas with homologous metaplasia. Ten cases, five with and five without EGFR amplification, were subjected to microarray-based CGH, which demonstrated that EGFR copy number gain may occur by amplification of a discrete genomic region or by gains of the short arm of chromosome 7 with a breakpoint near the EGFR gene locus, the minimal region of amplification mapping to EGFR, LANCL2, and SEC61G. No activating EGFR mutations were identified, suggesting that this is unlikely to be a common alternative underlying genetic mechanism for EGFR expression in metaplastic breast carcinomas. Given that metaplastic breast carcinomas are resistant to conventional chemotherapy or hormone therapy regimens and that tumours with EGFR amplification are reported to be sensitive to EGFR tyrosine kinase inhibitors, these findings indicate that further studies are warranted to explore EGFR tyrosine kinase inhibitors as potential therapeutic agents for metaplastic breast carcinomas harbouring amplification of 7p11.2.

An unexpected cause of an acute hypersensitivity reaction during recovery from anaesthesia.

Acute hypersensitivity reactions to chlorhexidine in the operating room are probably more likely to occur during the early phases of anaesthesia because chlorhexidine is often used for cleaning the surgical field or during placement of indwelling catheters. We report a case of an acute hypersensitivity reaction that occurred in the post anaesthetic care unit. Subsequent skin testing suggested sensitivity to chlorhexidine, which had been applied over the vaginal mucosa at the end of surgery. Relevant issues in the investigation of acute hypersensitivity reactions in the post anaesthetic period are discussed.