IGF-1R pathway activation as putative biomarker for linsitinib therapy to revert tamoxifen resistance in ER-positive breast cancer

Preclinical studies indicate that activated IGF-1R can drive endocrine resistance in ER-positive (ER+) breast cancer, but its clinical relevance is unknown. We studied the effect of IGF-1R signaling on tamoxifen benefit in patients and we searched for approaches to overcome IGF-1R-mediated tamoxifen failure in cell lines. Primary tumor blocks from postmenopausal ER+ breast cancer patients randomized between adjuvant tamoxifen versus nil were recollected. Immunohistochemistry for IGF-1R, p-IGF-1R/InsR, p-ERα(Ser118), p-ERα(Ser167) and PI3K/MAPK pathway proteins was performed. Multivariate Cox models were employed to assess tamoxifen efficacy. The association between p-IGF-1R/InsR and PI3K/MAPK pathway activation in MCF-7 and T47D cells was analyzed with Western blots. Cell proliferation experiments were performed under various growth-stimulating and -inhibiting conditions. Patients with ER+, IGF-1R-positive breast cancer without p-IGF-1R/InsR staining (n = 242) had tamoxifen benefit (HR 0.41, p = 0.0038), while the results for p-IGF-1R/InsR-positive patients (n = 125) were not significant (HR 0.95, p = 0.3). High p-ERα(Ser118) or p-ERα(Ser167) expression was associated with less tamoxifen benefit. In MCF-7 cells, IGF-1R stimulation increased phosphorylation of PI3K/MAPK proteins and ERα(Ser167) regardless of IGF-1R overexpression. This could be abrogated by the dual IGF-1R/InsR inhibitor linsitinib, but not by the IGF-IR-selective antibody 1H7. In MCF-7 and T47D cells, stimulation of the IGF-1R/InsR pathway resulted in cell proliferation regardless of tamoxifen. Abrogation of cell growth was regained by addition of linsitinib. In conclusion, p-IGF-1R/InsR positivity in ER+ breast cancer is associated with reduced benefit from adjuvant tamoxifen in postmenopausal patients. In cell lines, stimulation rather than overexpression of IGF-1R is driving tamoxifen resistance to be abrogated by linsitinib.     

Biperiden and mepazine effectively inhibit MALT1 activity and tumor growth in pancreatic cancer

MALT1 is a key mediator of NF-κB signaling and a main driver of B-cell lymphomas. Remarkably, MALT1 is expressed in the majority of pancreatic ductal adenocarcinomas (PDACs) as well, but absent from normal exocrine pancreatic tissue. Following, MALT1 shows off to be a specific target in cancer cells of PDAC without affecting regular pancreatic cells. Therefore, we studied the impact of pharmacological MALT1 inhibition in pancreatic cancer and showed promising effects on tumor progression. Mepazine (Mep), a phenothiazine derivative, is a known potent MALT1 inhibitor. Newly, we described that biperiden (Bip) is a potent MALT1 inhibitor with even less pharmacological side effects. Thus, Bip is a promising drug leading to reduced proliferation and increased apoptosis in PDAC cells in vitro and in vivo. By compromising MALT1 activity, nuclear translocation of c-Rel is prevented. c-Rel is critical for NF-κB-dependent inhibition of apoptosis. Hence, off-label use of Bip or Mep represents a promising new therapeutic approach to PDAC treatment. Regularly, the Anticholinergicum Bip is used to treat neurological side effects of Phenothiazines, like extrapyramidal symptoms.     

Is Secretory Activation Delayed in Women with Type Two Diabetes? A Pilot Study

(1) Background: Breastfeeding duration may be reduced in women with type 2 diabetes. Delayed secretory activation (SA) is associated with poorer breastfeeding outcomes; however, no prior studies have examined SA in women with type 2 diabetes. This pilot study aimed to assess SA in women with type 2 diabetes by assessing breastmilk constituents. Secondary aims were to assess breastfeeding rates postpartum, and contributory factors. (2) Methods: A prospective cohort of pregnant women with type 2 diabetes (n = 18) and two control groups with age- and parity-matched nondiabetic pregnant women (body mass index (BMI)) matched (n = 18) or normal-range BMI (n = 18)) were recruited. Breastmilk constituents (citrate, lactose, protein, and fat) were measured twice daily for 5 days postpartum and compared between groups. Associations between peripartum variables, breastmilk constituents, and breastfeeding at 4 months postpartum were explored. (3) Results: Women with type 2 diabetes had a slower increase in breastmilk citrate concentration postpartum, indicative of delayed SA, compared to both control groups. Higher predelivery insulin doses in women with type 2 diabetes were associated with increasing time to SA. Both women with type 2 diabetes and BMI-matched controls were less likely to fully breastfeed at 4 months, compared with normal-BMI controls. (4) Conclusion: SA is delayed in women with type 2 diabetes when compared to BMI-matched and normal-BMI women. Women with type 2 diabetes are less likely to fully breastfeed, at hospital discharge and by 4 months postpartum, compared to women with normal-BMI.     

Reduced Abundance of Nitrate-Reducing Bacteria in the Oral Microbiota of Women with Future Preeclampsia

The oral microbiota can contribute to the regulation of blood pressure by increasing the availability of nitric oxide through the reduction of nitrate to nitrite, which can be converted into nitric oxide in the stomach and then enter the circulation. It is unclear if the composition of the oral microbiota is different between women who do and do not develop preeclampsia. This study aimed to compare the composition of the buccal microbiota just prior to the development of symptoms at 36 weeks gestation in 12 women who developed late-onset preeclampsia and 24 matched women who remained normotensive throughout pregnancy by 16S rRNA gene amplicon sequencing. The abundance of the nitrate-reducing Veillonella spp V. parvula and V. dispar and a subunit of nitrate reductase narH was compared using real-time PCR. The abundance of bacteria was correlated with maternal blood pressure and dietary intake of nitrate-containing vegetables. The results showed that the abundance of nitrate-reducing bacteria including Veillonella, specifically V. parvula, and Prevotella was reduced in women who developed preeclampsia. Veillonella but not Prevotella abundance was negatively correlated with maternal blood pressure. The dietary intake of nitrate-containing vegetables did not differ between the groups and was not correlated with the abundance of Veillonella. There was no difference in the abundance of the nitrate reductase subunit narH between the groups. These results suggest that the abundance of nitrate-reducing bacteria is reduced in the oral microbiota of women who later develop preeclampsia, indicating a potential pathway for prevention.     

The effect of nightshift on emergency registrars' clinical skills

Objective: The effect of nightshift on ED staff performance is of clinical and risk‐management significance. Previous studies have demonstrated deterioration in psychomotor skills but the present study specifically assessed the impact of nightshift on clinical performance. Methods: The ED registrars in a tertiary hospital were enrolled in a prospective observational study and served as their own controls. During nightshift, subjects were presented simulated scenarios and tested with eight clinical questions developed to Fellowship examination standard. Matched scenarios and questions for the same subjects during dayshift served as controls. Two investigators, blinded to subject identity and the setting in which questions were attempted, independently collated answers. Results: Of 22 eligible subjects, all were recruited; four were excluded owing to incomplete data. A correlation of 0.99 was observed between the independent scoring investigators. Of a possible score of 17, the median result for nightshift was 9.5 (interquartile range: 8–11); corresponding value for dayshift was 12 (interquartile range: 10–13); P= 0.047. Conclusion: Nightshift effect on clinical performance is anecdotally well known. The present study quantifies such effects, specifically for the ED setting, and paves the way for focused research. The implications for clinical governance strategies are significant, as the fraternity embraces the mandate to maintain quality emergency care 24 h per day.     

Immunophenotypic features of MELF pattern invasion in endometrial adenocarcinoma: evidence for epithelial–mesenchymal transition

Aims:  Endometrial endometrioid adenocarcinomas (EEC) may show a distinctive morphological alteration characterized by the presence of microcystic, elongated and fragmented ('MELF') glands. These changes share features of epithelial–mesenchymal transition (EMT) in carcinomas arising at other sites. The aim was to compare the immunophenotypic profile of MELF-type epithelium with conventional glandular areas of EEC.
Methods and results:  Twenty-one EEC were stained immunohistochemically for cytokeratin (CK) AE1/AE3, CK7, vimentin, oestrogen receptor, progesterone receptor and E-cadherin. Conventional tumour glands usually showed preserved membranous E-cadherin immunoreactivity with peripheral accentuation of vimentin and hormone receptor expression. MELF-type invasion was characterized by strong CK7 expression, sometimes in contrast to adjacent unstained tumour glands. MELF areas were usually negative for hormone receptors and showed reduced E-cadherin expression.
Conclusions:  The expression of hormone receptors and intermediate filaments shows specific distribution patterns within EEC. MELF pattern invasion shows an altered immunophenotype compared with conventional glandular tumour areas. These findings suggest that MELF-type invasion represents a specific tumour alteration, and the reduction in hormone receptor and E-cadherin expression would be consistent with EMT. Immunohistochemical studies of EEC should consider micro anatomical variations in immunoreactivity, since these may be relevant to tumour invasion and progression.     

Characterization of the anti-HIV effects of native lactoferrin and other milk proteins and protein-derived peptides

In a search for natural proteins with anti-HIV activity, we screened a large set of purified proteins from bovine milk and peptide fragments thereof. Because several charged proteins and peptides are known to inhibit the process of virus entry, we selected proteins with an unusual charge composition or hydrophobicity profile. In contrast with some chemically modified (strongly negative) milk proteins, unmodified alpha(s2)-, beta- and kappa-casein, as well as several negatively and positively charged fragments thereof, did not show significant inhibition of virus replication. In fact, HIV-1 replication was elevated in the presence of beta-casein or amphiphilic fragments thereof. Bovine lactoferrin (bLF), a milk protein of 80 kDa, showed considerable inhibitory activity against HIV-1 with an IC50 of 0.4 microM. Modest inhibition was obtained with lactoferricin, a highly positively charged loop domain of bLF, indicating that other domains within the native bLF protein may also be required for inhibition. bLF blocked HIV-1 variants that use either the CXCR4 or the CCR5 coreceptor. In order to obtain further insight into the mechanism of action of this antiviral protein, we selected a bLF-resistant HIV-1 variant. The bLF-resistance phenotype is mediated by the viral envelope protein, which contains two interesting mutations that have previously been associated with an altered virus-host interaction and a modified receptor-coreceptor interaction. These results demonstrate that bLF targets the HIV-1 entry process.     

The Antiviral Protein Human Lactoferrin Is Distributed in the Body to Cytomegalovirus (CMV) Infection-Prone Cells and Tissues

Purpose. Lactoferrin has anti-Cytomegalovirus (CMV) and -HIV properties in vitro. However, the pharmacokinetic behavior of the 80-kD protein has not been well defined. We, therefore, assessed the plasma decay and body distribution of lactoferrin after intravenous administration to freely moving rats. Furthermore, the systemic availability of lactoferrin after intraperitoneal dosing was determined. Methods and Results. After intravenous injection, human lactoferrin (hLF) was rapidly cleared from the plasma, but higher doses resulted in prolonged plasma levels. Immunohistochemical analysis revealed a pronounced distribution of hLF to endothelial cells in the liver whereas diffuse staining in hepatocytes indicated the presence of considerable amounts in this large cell population. This endothelial association, which also was found in other organ/tissues, including blood vessels, was confirmed by in vitro cell-binding studies. In addition, leukocytes in plasma that were infiltrated in various organs showed binding of hLF. A small fraction of hLF was transported into the lymphatic system. Western blot analysis revealed that hLF, present in the various organs, mainly consisted of an 80-kD protein. After intraperitoneal administration, small amounts of 80-kD hLF distributed to the general circulation. The bioavailability was 0.6% but increased to 3.6% after multiple administrations. Conclusions. The affinity of hLF for endothelial cells and leukocytes, and its penetration into the lymphatic system, indicates that this protein reaches target cells and body compartments that are crucial for CMV and HIV replication. The ability to reach the blood compartment after intraperitoneal dosing offers opportunities for parenteral administration of the protein in future studies on its antiviral effects in vivo.