Proteomic alterations associated with residual disease in neoadjuvant chemotherapy treated ovarian cancer tissues.

BACKGROUND: Optimal cytoreduction to no residual disease (R0) correlates with improved disease outcome for high-grade serous ovarian cancer (HGSOC) patients. Treatment of HGSOC patients with neoadjuvant chemotherapy, however, may select for tumor cells harboring alterations in hallmark cancer pathways including metastatic potential. This study assessed this hypothesis by performing proteomic analysis of matched, chemotherapy naïve and neoadjuvant chemotherapy (NACT)-treated HGSOC tumors obtained from patients who had suboptimal (R1, n = 6) versus optimal (R0, n = 14) debulking at interval debulking surgery (IDS). METHODS: Tumor epithelium was harvested by laser microdissection from formalin-fixed, paraffin-embedded tissues from matched, pre- and post-NACT treated tumors for twenty HGSOC patients and analyzed by quantitative mass spectrometry-based proteomics. RESULTS: Differential analysis of patient matched pre- and post-NACT treated tumors revealed proteins associated with cell survival and metabolic signaling to be significantly altered in post-NACT treated tumor cells. Comparison of pre-NACT treated tumors from suboptimal (R1) versus optimally (R0) debulked patients identified proteins associated with tumor cell viability and invasion signaling enriched in R1 patients. We identified five proteins altered between R1 and R0 patients in pre- NACT treated tumors that significantly correlated with PFS in an independent cohort of HGSOC patients, including Fermitin family homolog 2 (FERMT2), a protein elevated in R1 that correlated with disease progression in HGSOC patients (multivariate Cox HR = 1.65, Wald p = 0.022) and increased metastatic potential in solid-tumor malignancies. CONCLUSIONS: This study identified distinct proteome profiles in patient matched pre- and post-NACT HGSOC tumors that correlate with NACT resistance and that may predict residual disease status at IDS that collectively warrant further pre-clinical investigation.

Inclusion of lipopeptides into the DMPC lipid bilayers prevents Aß peptide insertion.

Using an all-atom explicit water model and replica exchange with solute tempering molecular dynamics we have studied the binding of Aß10-40 peptides to the mixed cationic bilayers composed of DMPC lipids and C14-KAAK lipopeptides (LPs). Using as a control our previous replica exchange simulations probing binding of the same peptide to the zwitterionic DMPC bilayer we assessed the impact of lipopeptides on the Aß binding mechanism. We found that binding to the mixed DMPC + LP bilayers does not enhance the Aß helix propensity as much as binding to the pure DMPC bilayers. Tertiary interactions also differ in the peptide bound to the DMPC + LP bilayers due to a reduced helix content, salt bridge disruption, and the formation of new long-range hydrophobic interactions. More importantly, we showed that mixing lipopeptides into the DMPC bilayers prevents Aß10-40 insertion forcing the peptide to reside on the bilayer surface and considerably destabilizes Aß-bilayer interactions leading to the formation of a shallow water layer between the peptide and the bilayer. Furthermore, we demonstrated that Aß10-40 peptides cause minor DMPC + LP bilayer thinning and perturbation beneath their binding footprint. These observations stand in sharp contrast to Aß10-40 binding to the pure DMPC bilayers, which results in deep peptide insertion and significant disruption of the bilayer structure. We argued that lipopeptides expel Aß10-40 peptides from the bilayer due to strong, mostly electrostatic, interfacial interactions introduced by the lipopeptides into the bilayers. We therefore propose that C14-KAAK lipopeptides can be used to engineer lipid bilayers, which withstand binding of Alzheimer's Aß peptides.

Standardization and harmonization of distributed multi-center proteotype analysis supporting precision medicine studies.

Cancer has no borders: Generation and analysis of molecular data across multiple centers worldwide is necessary to gain statistically significant clinical insights for the benefit of patients. Here we conceived and standardized a proteotype data generation and analysis workflow enabling distributed data generation and evaluated the quantitative data generated across laboratories of the international Cancer Moonshot consortium. Using harmonized mass spectrometry (MS) instrument platforms and standardized data acquisition procedures, we demonstrate robust, sensitive, and reproducible data generation across eleven international sites on seven consecutive days in a 24/7 operation mode. The data presented from the high-resolution MS1-based quantitative data-independent acquisition (HRMS1-DIA) workflow shows that coordinated proteotype data acquisition is feasible from clinical specimens using such standardized strategies. This work paves the way for the distributed multi-omic digitization of large clinical specimen cohorts across multiple sites as a prerequisite for turning molecular precision medicine into reality.

Signal integration by the two-component signal transduction response regulator CpxR.

The CpxAR two-component signal transduction system in Escherichia coli and other pathogens senses diverse envelope stresses and promotes the transcription of a variety of genes that remedy these stresses. An important member of the CpxAR regulon is cpxP. The CpxA-dependent transcription of cpxP has been linked to stresses such as misfolded proteins and alkaline pH. It also has been proposed that acetyl phosphate, the intermediate of the phosphotransacetylase (Pta)-acetate kinase (AckA) pathway, can activate the transcription of cpxP in a CpxA-independent manner by donating its phosphoryl group to CpxR. We tested this hypothesis by measuring the transcription of cpxP using mutants with mutations in the CpxAR pathway, mutants with mutations in the Pta-AckA pathway, and mutants with a combination of both types of mutations. From this epistasis analysis, we learned that CpxR integrates diverse stimuli. The stimuli that originate in the envelope depend on CpxA, while those associated with growth and central metabolism depend on the Pta-AckA pathway. While CpxR could receive a phosphoryl group from acetyl phosphate, this global signal was not the primary trigger for CpxR activation associated with the Pta-AckA pathway. On the strength of these results, we contend that the interactions between central metabolism and signal transduction can be quite complex and that successful investigations of such interactions must include a complete epistatic analysis.

Molecular Mechanisms of Alzheimer's Biomarker FDDNP Binding to Aß Amyloid Fibril.

Using isobaric-isothermal replica exchange molecular dynamics and the all-atom explicit water model, we examined the binding of FDDNP biomarkers to the Aß amyloid fibril fragment. Our results can be summarized as follows. First, FDDNP ligands bind with high affinity to the Aß fibril, and the hydrophobic effect together with π-stacking interactions are the dominant factors governing FDDNP binding. In comparison, electrostatic interactions and hydrogen bonding play a minor role. Second, our simulations reveal a strong tendency of bound FDDNP molecules for self-aggregation. Accordingly, about two-thirds of all bound ligands form aggregated clusters of various sizes, and ligand-ligand interactions make considerable contribution to FDDNP binding. Third, FDDNP ligands bind to two distinct sites on the Aß fibril. Primary binding sites (NT) are located at the N-terminals of Aß10-40 peptides, whereas secondary ones (CE) occur on the concave fibril edge near fibril channels. The NT sites are characterized by strong hydrophobic and π-stacking interactions, favorable binding entropy resulting from multiple FDDNP binding orientations and propensity for self-aggregation but relatively weak van der Waals interactions. In contrast, the CE sites offer stronger van der Waals binding interactions but weaker hydrophobic and aromatic interactions and less favorable binding entropy. By comparing our data with previous studies, we suggest that the primary binding locations identified by us are likely to occur in other Aß fibril polymorphic structures. We also show that FDDNP binds via distinct mechanisms to Aß fibrils and monomers. We argue that FDDNP binds with stronger affinity to benign Aß monomers than to the fibrils, raising questions about the ability of FDDNP to selectively label amyloid deposits.