RAD51 foci as a functional biomarker of homologous recombination repair and PARP inhibitor resistance in germline BRCA-mutated breast cancer.

Background: BRCA1 and BRCA2 (BRCA1/2)-deficient tumors display impaired homologous recombination repair (HRR) and enhanced sensitivity to DNA damaging agents or to poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi). Their efficacy in germline BRCA1/2 (gBRCA1/2)-mutated metastatic breast cancers has been recently confirmed in clinical trials. Numerous mechanisms of PARPi resistance have been described, whose clinical relevance in gBRCA-mutated breast cancer is unknown. This highlights the need to identify functional biomarkers to better predict PARPi sensitivity. Patients and methods: We investigated the in vivo mechanisms of PARPi resistance in gBRCA1 patient-derived tumor xenografts (PDXs) exhibiting differential response to PARPi. Analysis included exome sequencing and immunostaining of DNA damage response proteins to functionally evaluate HRR. Findings were validated in a retrospective sample set from gBRCA1/2-cancer patients treated with PARPi. Results: RAD51 nuclear foci, a surrogate marker of HRR functionality, were the only common feature in PDX and patient samples with primary or acquired PARPi resistance. Consistently, low RAD51 was associated with objective response to PARPi. Evaluation of the RAD51 biomarker in untreated tumors was feasible due to endogenous DNA damage. In PARPi-resistant gBRCA1 PDXs, genetic analysis found no in-frame secondary mutations, but BRCA1 hypomorphic proteins in 60% of the models, TP53BP1-loss in 20% and RAD51-amplification in one sample, none mutually exclusive. Conversely, one of three PARPi-resistant gBRCA2 tumors displayed BRCA2 restoration by exome sequencing. In PDXs, PARPi resistance could be reverted upon combination of a PARPi with an ataxia-telangiectasia mutated (ATM) inhibitor. Conclusion: Detection of RAD51 foci in gBRCA tumors correlates with PARPi resistance regardless of the underlying mechanism restoring HRR function. This is a promising biomarker to be used in the clinic to better select patients for PARPi therapy. Our study also supports the clinical development of PARPi combinations such as those with ATM inhibitors.

A dominant-negative N-terminal fragment of HER2 frequently expressed in breast cancers.

The transmembrane tyrosine kinase HER2 (ErbB2, neu) is a prototypical biomarker for breast cancers and a therapeutic target. Although anti-HER2 therapies are remarkably effective, HER2-positive tumors are heterogeneous and some subtypes do not respond or develop resistance to these therapies. Here we show that H2NTF, a novel N-terminal fragment of HER2, is expressed at variable levels in 60% of the breast cancer samples analyzed. Characterization of H2NTF shows that it is devoid of the tyrosine kinase domain but it readily interacts with full-length HER2 and other HER receptors. As a consequence, H2NTF acts as a dominant-negative, attenuating the signaling triggered by full-length HER receptors. Expression of H2NTF results in resistance to the treatment with low concentrations of trastuzumab in vitro. However, cells expressing H2NTF and non-expressing cells have similar sensitivity to trastuzumab in vivo, indicating that H2NTF/trastuzumab complexes trigger antibody-dependent cell-mediated cytotoxicity.

Study on some factors affecting survivability of airborne fungi.

The aim of the present study was to investigate the effect of some air pollutants and meteorological parameters on the survivability of airborne fungi. Fungi were collected by using a slit impactor sampler calibrated to draw 20 L/min, for 3 min. Nitrogen dioxide (NO(2)), sulfur dioxide (SO(2)), particulate matter (PM), relative humidity (RH %), temperature (T °C) and wind speed (WS) were also measured. Air samples were taken during the period from March 2006 to February 2007. Fungal concentrations ranged between 45 and 451 CFU/m(3) with an annual mean concentration of 216 CFU/m(3). The lowest fungal concentration was found in the summer, however the highest one was found in the autumn. NO(2,) SO(2) and PM averaged 83.66 µg/m(3), 67.01 µg/m(3), and 237.69 µg/m(3), respectively. T °C was positively and negatively correlated with Aspergillus (P = 0.000) and Penicillium (P = 0.007), respectively. RH% was positively correlated with total fungi (P = 0.001), Aspergillus (P = 0.002) and Cladosporium (P = 0.047). Multiple regression analysis showed that T °C and RH% were the most predicted variants. Non-significant correlations were found between fungal concentrations and air pollutants. Meteorological parameters were the critical factors affecting fungal survivability.

PI3K inhibition results in enhanced HER signaling and acquired ERK dependency in HER2-overexpressing breast cancer.

There is a strong rationale to therapeutically target the phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mTOR) pathway in breast cancer since it is highly deregulated in this disease and it also mediates resistance to anti-HER2 therapies. However, initial studies with rapalogs, allosteric inhibitors of mTORC1, have resulted in limited clinical efficacy probably due to the release of a negative regulatory feedback loop that triggers AKT and ERK signaling. Since activation of AKT occurs via PI3K, we decided to explore whether PI3K inhibitors prevent the activation of these compensatory pathways. Using HER2-overexpressing breast cancer cells as a model, we observed that PI3K inhibitors abolished AKT activation. However, PI3K inhibition resulted in a compensatory activation of the ERK signaling pathway. This enhanced ERK signaling occurred as a result of activation of HER family receptors as evidenced by induction of HER receptors dimerization and phosphorylation, increased expression of HER3 and binding of adaptor molecules to HER2 and HER3. The activation of ERK was prevented with either MEK inhibitors or anti-HER2 monoclonal antibodies and tyrosine kinase inhibitors. Combined administration of PI3K inhibitors with either HER2 or MEK inhibitors resulted in decreased proliferation, enhanced cell death and superior anti-tumor activity compared with single agent PI3K inhibitors. Our findings indicate that PI3K inhibition in HER2-overexpressing breast cancer activates a new compensatory pathway that results in ERK dependency. Combined anti-MEK or anti-HER2 therapy with PI3K inhibitors may be required in order to achieve optimal efficacy in HER2-overexpressing breast cancer. This approach warrants clinical evaluation.

Comparison of once-daily versus twice-daily administration of cefdinir against typical bacterial respiratory tract pathogens.

In an in vitro pharmacodynamic model, a twice-daily cefdinir dosing regimen was more effective than a once-daily regimen against common bacterial respiratory pathogens in producing 3-log(10) killing and preventing the occurrence of regrowth at 24 h. Twice-daily administration is likely the more appropriate cefdinir dosing strategy for the treatment of community-acquired pneumonia.

In vitro pharmacodynamic analysis of single daily dosing versus conventional dosing of gentamicin administered with penicillin against Enterococcus faecalis.

STUDY OBJECTIVE: To compare the effectiveness of single daily dosing (SDD) versus conventional dosing of gentamicin when administered with penicillin to treat enterococcal infections. DESIGN: In vitro pharmacodynamic model. SETTING: Hospital laboratory. MEASUREMENTS AND MAIN RESULTS: A 24-hour in vitro pharmacodynamic model was employed to simulate SDD and 3 times/day dosing of gentamicin, in conjunction with continuously infused penicillin, against Enterococcus faecalis. Duplicate 24-hour kill curves were generated with varying concentrations of penicillin and gentamicin alone and in combination. No difference in the rate of kill was seen between any combination of penicillin and gentamicin. Regrowth occurred only with drug combinations in which penicillin was administered continuously at the minimum inhibitory concentration. Variations in the gentamicin dosing regimen did not affect regrowth. CONCLUSION: In the treatment of enterococcal infections, an SDD regimen for gentamicin shows no efficacy benefit compared with conventional dosing.