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Experts point out that “A quarter of all overhead power lines outages are caused by birds, and 86% of grid companies name birds as main problem”. Unfortunately, this is the key factor in the problem of mass death of birds: power engineers are forced to protect lines from birds, while the issue of protecting birds from lines has lower priority. In any case, it all comes down to one question: what should we do?

Introduction: We have developed a delivery approach that uses two pHLIP peptides that collaborate in the targeted intracellular delivery of a single payload, dimeric STINGa (dMSA).Methods: dMSA was conjugated with two pHLIP peptides via S-S cleavable self-immolating linkers to form 2pHLIP-dMSA.Results: Biophysical studies were carried out to confirm pH-triggered interactions of the 2pHLIP-dMSA with membrane lipid bilayers. The kinetics of linker self-immolation and dMSA release, the pharmacokinetics, the binding to plasma proteins, the stability of the agent in plasma, the targeting and resulting cytokine activation in tumors, and the biodistribution of the construct was investigated. This is the first study demonstrating that combining the energy of the membrane-associated folding of two pHLIPs can be utilized to enhance the targeted intracellular delivery of large therapeutic cargo payloads.Discussion: Linking two pHLIPs to the cargo extends blood half-life, and targeted delivery of dimeric STINGa induces tumor eradication and the development of robust anti-cancer immunity.

The family of pH (Low) Insertion Peptides (pHLIP) comprises a tumor-agnostic technology that uses the low pH (or high acidity) at the surfaces of cells within the tumor microenvironment (TME) as a targeted biomarker. pHLIPs can be used for extracellular and intracellular delivery of a variety of imaging and therapeutic payloads. Unlike therapeutic delivery targeted to specific receptors on the surfaces of particular cells, pHLIP targets cancer, stromal and some immune cells all at once. Since the TME exhibits complex cellular crosstalk interactions, simultaneous targeting and delivery to different cell types leads to a significant synergistic effect for many agents. pHLIPs can also be positioned on the surfaces of various nanoparticles (NPs) for the targeted intracellular delivery of encapsulated payloads. The pHLIP technology is currently advancing in pre-clinical and clinical applications for tumor imaging and treatment.

Dysregulated extracellular pH, the universal feature of tumor, works as an evolutional force to drive dissemination of tumor cells. It is well-established that tumor acidity is associated with tumor growth and metastasis. However, the pH of pre-metastatic niche remains unclear. We hypothesized that primary tumor cells remotely prime acidity in secondary organ to achieve metastatic colonization. Herein, we demonstrated that the pH responsive probe pH Low Insertion Peptide (pHLIP) was notably accumulated in pre-metastatic lungs of 4T1.2 breast tumor-bearing mice. The pHLIP-targeted lungs showed high amounts of lactate and overexpressed glycolysis-related proteins. Pharmacological inhibition of glycolysis suppressed the lung acidification induced by 4T1.2 cancer cell culture supernatant and delayed subsequent metastatic burden of disseminated tumor cells. In the acidic lungs, pHLIP was primarily localized in alveolar type 2 cells which strongly expressed glycolysis-related proteins. 4T1.2-derived extracellular vesicles expressed some of the glycolysis-related proteins, and their administration increased pHLIP accumulation and glycolytic enhancement in lungs. pHLIP-conjugated dexamethasone effectively attenuated lung metastatic burden by disrupting pro-inflammatory response in the acidic lungs. From these results, targeting the metastasis-supporting microenvironment by pHLIP technology creates possibility to identify pre-metastatic organ and prevent metastatic recurrence.

Targeted antigen delivery allows activation of the immune system to kill cancer cells. Here we report the targeted delivery of various epitopes, including a peptide, a small molecule, and a sugar, to tumors by pH Low Insertion Peptides (pHLIPs), which respond to surface acidity and insert to span the membranes of metabolically activated cancer and immune cells within tumors. Epitopes linked to the extracellular ends of pH Low Insertion Peptide peptides were positioned at the surfaces of tumor cells and were recognized by corresponding anti-epitope antibodies. Special attention was devoted to the targeted delivery of the nine residue HA peptide epitope from the Flu virus hemagglutinin. The HA sequence is not present in the human genome, and immunity is readily developed during viral infection or immunization with KLH-HA supplemented with adjuvants. We tested and refined a series of double-headed HA-pHLIP agents, where two HA epitopes were linked to a single pH Low Insertion Peptide peptide via two Peg12 or Peg24 polymers, which enable HA epitopes to engage both antibody binding sites. HA-epitopes positioned at the surfaces of tumor cells remain exposed to the extracellular space for 24–48 h and are then internalized. Different vaccination schemes and various adjuvants, including analogs of FDA approved adjuvants, were tested in mice and resulted in a high titer of anti-HA antibodies. Anti-HA antibody binds HA-pHLIP in blood and travels as a complex leading to significant tumor targeting with no accumulation in organs and to hepatic clearance. HA-pHLIP agents induced regression of 4T1 triple negative breast tumor and B16F10 MHC-I negative melanoma tumors in immunized mice. The therapeutic efficacy potentially is limited by the drop of the level of anti-HA antibodies in the blood to background level after three injections of HA-pHLIP. We hypothesize that additional boosts would be required to keep a high titer of anti-HA antibodies to enhance efficacy. pH Low Insertion Peptide-targeted antigen therapy may provide an opportunity to treat tumors unresponsive to T cell based therapies, having a small number of neo-antigens, or deficient in MHC-I presentation at the surfaces of cancer cells either alone or in combination with other approaches.

Despite significant progress in the development of novel STING agonists (STINGa), applications appear to be challenged by the low efficiency and poor selectivity of these agents. A pH Low Insertion Peptide (pHLIP) extends the lifetime of a STINGa in the blood and targets it to acidic cancer-associated fibroblasts (CAFs), tumor-associated macrophages (TAMs), myeloid derived suppressor cells (mMDSCs) and dendritic cells (DCs). CAFs constitute 25% of all live cells within CT26 tumors, and M2-type TAMs and mMDSCs are the most abundant among the immune cells. The resulting activation of cytokines within the tumor microenvironment (TME) triggers the eradication of small (100 mm3) and large (400-700 mm3) CT26 tumors in mice after a single dose of pHLIP-STINGa. The tumor stroma was destroyed (the number of CAFs was reduced by 98%), intratumoral hemorrhage developed, and the level of acidity within the TME was reduced. Further, no tumors developed in 20 out of 25 tumor-free mice re-challenged by an additional injection of cancer cells. The therapeutic effect on CT26 tumors was insignificant in nude mice, lacking T-cells. Thus, targeted delivery of STINGa to tumor stroma and TAMs induces activation of signaling, potentially resulting in the recruitment and infiltration of T-cells, which gain access to the tumor core. The cytotoxic activity of T-cells is not impaired by an acidic environment and immune memory is developed.

SrLnCuSe3 (Ln = Dy, Ho, Er, Tm) compounds crystallize in the Pnma and Cmcm orthorhombic space group and belong to the Eu2CuS3 and KCuZrS3 structural type, respectively. According to a single-crystal XRD study, the SrTmCuSe3 unit cell parameters are a = 4.0631 (4), b = 13.4544 (14), c = 10.4430 (10) Å, and V = 570.88 (10) Å3. All the studied SrLnCuSe3 samples in the temperature range of 1.77–300 K demonstrate paramagnetic behavior without any features pointing to magnetic ordering. The measured Curie constants coincide with the values expected for Ln3+ ions with good accuracy, which confirms the stoichiometric composition of the samples and the non-magnetic state of the copper ions, Cu1+ (S = 0). The conducted optical absorption study showed that the polycrystalline SrLnCuSe3 (Ln = Dy, Ho, Er, Tm) samples are semiconductors with a direct bandgap ranging from 2.14 eV to 2.31 eV. Two indirect bandgaps were revealed and explained by the presence of optical transitions to highly dispersive subbands in the conduction band. The compounds demonstrate two reversible phase transitions α⇆β, β⇆γ and an incongruent melting at 1606 K (Dy), 1584 K (Ho), 1634 K (Er), and 1620 K (Tm) associated with the formation of solid solutions of SrSe, Cu2-XSe, and Ln2Se3 binary compounds.

The oxidation reactions of BaLaCuS3 in the artificial air atmosphere were studied at different heating rates in the temperature range of 50–1200 °C. The oxidation stages were determined by DSC-TG, XRD and IR–vis methods. The kinetic characteristics of the proceeding reactions were obtained with the use of the Kissinger model in a linearized form. Compound BaLaCuS3 was stable in the air up to 280 °C. Upon further heating up to 1200 °C, this complex sulfide underwent three main oxidation stages. The first stage is the formation of BaSO4 and CuLaS2. The second stage is the oxidation of CuLaS2 to La2O2SO4 and copper oxides. The third stage is the destruction of La2O2SO4. The final result of the high-temperature treatment in the artificial air atmosphere was a mixture of barium sulfate, copper (II) oxide and La2CuO4. The mechanism and stages of BaLaCuS3 oxidation and further interactions of the components were discussed.

T-cell activation primarily occurs in the lymph nodes, highly organized and specialized secondary lymphoid organs. Here the authors show that the acidic extracellular pH in lymph node paracortical zones limits cytokine production by effector T-cells, but does not alter their activation by antigen-presenting cells.

Acidosis of the tumor microenvironment is a hallmark of tumor progression and has emerged as an essential biomarker for cancer diagnosis, prognosis, and evaluation of treatment response. A tool for quantitatively visualizing the acidic tumor environment could significantly advance our understanding of the behavior of aggressive tumors, improving patient management and outcomes. 89Zr-labeled pH-low insertion peptides (pHLIP) are a class of radiopharmaceutical imaging probes for the in vivo analysis of acidic tumor microenvironments via positron emission tomography (PET). Their unique structure allows them to sense and target acidic cancer cells. In contrast to traditional molecular imaging agents, pHLIP’s mechanism of action is pH-dependent and does not rely on the presence of tumor-specific molecular markers. In this study, one promising acidity-imaging PET probe ([89Zr]Zr-DFO-Cys-Var3) was identified as a candidate for clinical translation.

Praseodymium sulfate was obtained by the precipitation method and the crystal structure was determined by Rietveld analysis. Pr2(SO4)3 is crystallized in the monoclinic structure, space group C2/c, with cell parameters a = 21.6052 (4), b = 6.7237 (1) and c = 6.9777 (1) Å, β = 107.9148 (7)°, Z = 4, V = 964.48 (3) Å3 (T = 150 °C). The thermal expansion of Pr2(SO4)3 is strongly anisotropic. As was obtained by XRD measurements, all cell parameters are increased on heating. However, due to a strong increase of the monoclinic angle β, there is a direction of negative thermal expansion. In the argon atmosphere, Pr2(SO4)3 is stable in the temperature range of T = 30–870 °C. The kinetics of the thermal decomposition process of praseodymium sulfate octahydrate Pr2(SO4)3·8H2O was studied as well. The vibrational properties of Pr2(SO4)3 were examined by Raman and Fourier-transform infrared absorption spectroscopy methods. The band gap structure of Pr2(SO4)3 was evaluated by ab initio calculations, and it was found that the valence band top is dominated by the p electrons of oxygen ions, while the conduction band bottom is formed by the d electrons of Pr3+ ions. The exact position of ZPL is determined via PL and PLE spectra at 77 K to be at 481 nm, and that enabled a correct assignment of luminescent bands. The maximum luminescent band in Pr2(SO4)3 belongs to the 3P0 → 3F2 transition at 640 nm.

The advantages of targeted therapy have motivated many efforts to find distinguishing features between the molecular cell surface landscapes of diseased and normal cells. Typically, the features have been proteins, lipids or carbohydrates, but other approaches are emerging. In this discussion, we examine the use of cell surface acidity as a feature that can be exploited by using pH-sensitive peptide folding to target agents to diseased cell surfaces or cytoplasms.

Abstract Acidity at surface of cancer cells is a hallmark of tumor microenvironments, which does not depend on tumor perfusion, thus it may serve as a general biomarker for targeting tumor cells. We used the pH (low) insertion peptide (pHLIP) for decoration of liposomes and niosomes. pHLIP senses pH at the surface of cancer cells and inserts into the membrane of targeted cells, and brings nanomaterial to close proximity of cellular membrane. DMPC liposomes and Tween 20 or Span 20 niosomes with and without pHLIP in their coating were fully characterized in order to obtain fundamental understanding on nanocarrier features and facilitate the rational design of acidity sensitive nanovectors. The samples stability over time and in presence of serum was demonstrated. The size, ζ-potential, and morphology of nanovectors, as well as their ability to entrap a hydrophilic probe and modulate its release were investigated. pHLIP decorated vesicles could be useful to obtain a prolonged (modified) release of biological active substances for targeting tumors and other acidic diseased tissues.

EuLnCuSe3 (Ln = Nd, Sm, Gd, Er), due to their complex composition, should be considered new materials with the ability to purposefully change the properties. Samples of the EuLnCuSe3 were prepared using Cu, rare earth metal, Se (99.99%) by the ampoule method. The samples were obtained by the crystallization from a melt and annealed at temperatures 1073 and 1273 K. The EuErCuSe3 crystal structure was established using the single-crystal particle. EuErCuSe3 crystallizes in the orthorhombic system, space group Cmcm, KCuZrS3 structure type, with cell parameters a = 4.0555 (3), b = 13.3570 (9), and c = 10.4602 (7) Å, V = 566.62 (6) Å3. In structure EuErCuSe3, erbium ions are coordinated by selenium ions in the octahedral polyhedron, copper ions are in the tetrahedral coordination, europium ions are between copper and erbium polyhedra layers and are coordinated by selenium ions as two-cap trigonal prisms. The optical band gap is 1.79 eV. At 4.7 K, a transition from the ferrimagnetic state to the paramagnetic state was detected in EuErCuSe3. At 85 and 293 K, the compound is in a paramagnetic state. According to XRPD data, EuLnCuSe3 (Ln = Nd, Sm, Gd) compounds have a Pnma orthorhombic space group of the Eu2CuS3 structure type. For EuSmCuSe3, a = 10.75704 (15) Å, b = 4.11120 (5) Å, c = 13.37778 (22) Å. In the series of EuLnCuSe3 compounds, the optical band gap increases 1.58 eV (Nd), 1.58 eV (Sm), 1.72 eV (Gd), 1.79 eV (Er), the microhardness of the 205 (Nd), 210 (Sm), 225 (Gd) 235 ± 4 HV (Er) phases increases, and the thermal stability of the phases increases significantly. According to the measurement data of differential scanning calorimetry, the EuNdCuSe3 decomposes, according to the solid-phase reaction T = 1296 K, ΔH = 8.2 ± 0.8 kJ/mol. EuSmCuSe3 melts incongruently T = 1449 K, ΔH = 18.8 ± 1.9 kJ/mol. For the EuGdCuSe3, two (Tα↔β = 1494 K, ΔHα↔β = 14.8 kJ/mol, Tβ↔γ = 1530 K, ΔHβ↔γ = 4.8 kJ/mol) and for EuErCuSe3 three polymorphic transitions (Tα↔β = 1561 K, ΔHα↔β = 30.3 kJ/mol, Tβ↔γ = 1579 K, ΔHβ↔γ = 4.4 kJ/mol, and Tγ↔δ = 1600 K, ΔHγ↔δ = 10.1 kJ/mol). The compounds melt incongruently at the temperature of 1588 K, ΔHmelt = 17.9 ± 1.8 kJ/mol and 1664 K, ΔHmelt = 25.6 ± 2.5 kJ/mol, respectively. Incongruent melting of the phases proceeds with the formation of a solid solution of EuSe and a liquid phase.

Novel approaches in synthesis of spherical and multispiked gold nanoparticles coated with polyethylene glycol (PEG) and pH Low Insertion Peptide (pHLIP®) were introduced. The presence of a tumor-targeting pHLIP® peptide in the nanoparticle coating enhances the stability of particles in solution and promotes a pH-dependent cellular uptake. The spherical particles were prepared with sodium citrate as a gold reducing agent to form particles of 7.0±2.5 nm in mean metallic core diameter and ∼43 nm in mean hydrodynamic diameter. The particles that were injected into tumors in mice (21 µg of gold) were homogeneously distributed within a tumor mass with no staining of the muscle tissue adjacent to the tumor. Up to 30% of the injected gold dose remained within the tumor one hour post-injection. The multispiked gold nanoparticles with a mean metallic core diameter of 146.0±50.4 nm and a mean hydrodynamic size of ~161 nm were prepared using ascorbic acid as a reducing agent and disk-like bicelles as a template. Only the presence of a soft template, like bicelles, ensured the appearance of spiked nanoparticles with resonance in the near infrared region. The irradiation of spiked gold nanoparticles by an 805 nm laser led to the time- and concentration-dependent increase of temperature. Both pHLIP® and PEG coated gold spherical and multispiked nanoparticles might find application in radiation and thermal therapies of tumors.

Crystalline hydrate of double cesium europium sulfate [CsEu(H2O)3(SO4)2]·H2O was synthesized by the crystallization from an aqueous solution containing equimolar amounts of 1Cs+:1Eu3+:2SO42− ions. Anhydrous salt CsEu(SO4)2 was formed as a result of the thermal dehydration of the crystallohydrate. The unusual effects observed during the thermal dehydration were attributed to the specific coordination of water molecules in the [CsEu(H2O)3(SO4)2]·H2O structure. The crystal structure of [CsEu(H2O)3(SO4)2]·H2O was determined by a single crystal X-ray diffraction analysis, and the crystal structure of CsEu(SO4)2 was obtained by the Rietveld method. [CsEu(H2O)3(SO4)2]·H2O crystallizes in the monoclinic system, space group P21/c (a = 6.5574(1) Å, b = 19.0733(3) Å, c = 8.8364(2) Å, β = 93.931(1)°, V = 1102.58(3) Å3). The anhydrous sulfate CsEu(SO4)2 formed as a result of the thermal destruction crystallizes in the monoclinic system, space group C2/c (a = 14.327(1) Å, b = 5.3838(4) Å, c = 9.5104(6) Å, β = 101.979(3) °, V = 717.58(9) Å3). The vibration properties of the compounds are fully consistent with the structural models and are mainly determined by the deformation of non-rigid structural elements, such as H2O and SO42−. As shown by the diffused reflection spectra measurements and DFT calculations, the structural transformation from [CsEu(H2O)3(SO4)2]·H2O to CsEu(SO4)2 induced a significant band gap reduction. A noticeable difference of the luminescence spectra between cesium europium sulfate and cesium europium sulfate hydrate is detected and explained by the variation of the extent of local symmetry violation at the crystallographic sites occupied by Eu3+ ions, namely, by the increase in inversion asymmetry in [CsEu(H2O)3(SO4)2]·H2O and the increase in mirror asymmetry in CsEu(SO4)2. The chemical shift of the 5D0 energy level in cesium europium sulfate hydrate, with respect to cesium europium sulfate, is associated with the presence of H2O molecules in the vicinity of Eu3+ ion.

The discovery of the pH Low Insertion Peptides (pHLIPs) provides an opportunity to develop imaging and drug delivery agents targeting extracellular acidity. Extracellular acidity is associated with many pathological states, such as those in cancer, ischemic stroke, neurotrauma, infection, lacerations and others. The metabolism of cells in injured or diseased tissues often results in the acidification of the extracellular environment, so acidosis might be useful as a general marker for the imaging and treatment of diseased states if an effective targeting method can be developed. The molecular mechanism of a pHLIP peptide is based on pH-dependent membrane-associated folding. pHLIPs, being moderately hydrophobic peptides, have high affinities for cellular membranes at normal pH, but fold and insert across membranes at low pH, allowing them to sense pH at the surfaces of cells in diseased tissues, where it is the lowest. Here we discuss the main principles of pHLIP interactions with membrane lipid bilayers at neutral and low pHs, the possibility of tuning the folding and insertion pH by peptide sequence variation, and potential applications of pHLIPs for imaging, therapy and image-guided interventions.