Mathematical Modeling Predicts Enhanced Growth of X-Ray Irradiated Pigmented Fungi

Ionizing radiation is known for its cytotoxic and mutagenic properties. However, recent evidence suggests that chronic sub-lethal irradiation stimulates the growth of melanin-pigmented (melanized) fungi, supporting the hypothesis that interactions between melanin and ionizing photons generate energy useful for fungal growth, and/or regulate growth-promoting genes. There are no quantitative models of how fungal proliferation is affected by ionizing photon energy, dose rate, and presence versus absence of melanin on the same genetic background. Here we present such a model, which we test using experimental data on melanin-modulated radiation-induced proliferation enhancement in the fungus Cryptococcus neoformans, exposed to two different peak energies (150 and 320 kVp) over a wide range of X-ray dose rates. Our analysis demonstrates that radiation-induced proliferation enhancement in C. neoformans behaves as a binary “on/off” phenomenon, which is triggered by dose rates 5000 mGy/h. Proliferation enhancement of irradiated cells compared with unirradiated controls occurs at both X-ray peak energies, but its magnitude is modulated by X-ray peak energy and cell melanization. At dose rates <5000 mGy/h, both melanized and non-melanized cells exposed to 150 kVp X-rays, and non-melanized cells exposed to 320 kVp X-rays, all exhibit the same proliferation enhancement: on average, chronic irradiation stimulates each founder cell to produce 100 (95% CI: 83, 116) extra descendants over 48 hours. Interactions between melanin and 320 kVp X-rays result in a significant (2-tailed p-value = 4.8×10−5) additional increase in the number of radiation-induced descendants per founder cell: by 55 (95% CI: 29, 81). These results show that both melanin-dependent and melanin-independent mechanisms are involved in radiation-induced fungal growth enhancement, and implicate direct and/or indirect interactions of melanin with high energy ionizing photons as an important pro-proliferative factor

Treating Cancer as an Infectious Disease—Viral Antigens as Novel Targets for Treatment and Potential Prevention of Tumors of Viral Etiology

Nearly 20% of human cancers worldwide have an infectious etiology with the most prominent examples being hepatitis B and C virus-associated hepatocellular carcinoma and human papilloma virus-associated cervical cancer. There is an urgent need to find new approaches to treatment and prevention of virus-associated cancers.Viral antigens have not been previously considered as targets for treatment or prevention of virus-associated cancers. We hypothesized that it was possible to treat experimental HPV16-associated cervical cancer (CC) and Hepatitis B-associated hepatocellular carcinoma (HCC) by targeting viral antigens expressed on cancer cells with radiolabeled antibodies to viral antigens. Treatment of experimental CC and HCC tumors with (188)Re-labeled mAbs to E6 and HBx viral proteins, respectively, resulted in significant and dose-dependent retardation of tumor growth in comparison with untreated mice or mice treated with unlabeled antibodies.This strategy is fundamentally different from the prior uses of radioimmunotherapy in oncology, which targeted tumor-associated human antigens and promises increased specificity and minimal toxicity of treatment. It also raises an exciting possibility to prevent virus-associated cancers in chronically infected patients by eliminating cells infected with oncogenic viruses before they transform into cancer

Ionizing Radiation Changes the Electronic Properties of Melanin and Enhances the Growth of Melanized Fungi

BACKGROUND: Melanin pigments are ubiquitous in nature. Melanized microorganisms are often the dominating species in certain extreme environments, such as soils contaminated with radionuclides, suggesting that the presence of melanin is beneficial in their life cycle. We hypothesized that ionizing radiation could change the electronic properties of melanin and might enhance the growth of melanized microorganisms. METHODOLOGY/PRINCIPAL FINDINGS: Ionizing irradiation changed the electron spin resonance (ESR) signal of melanin, consistent with changes in electronic structure. Irradiated melanin manifested a 4-fold increase in its capacity to reduce NADH relative to non-irradiated melanin. HPLC analysis of melanin from fungi grown on different substrates revealed chemical complexity, dependence of melanin composition on the growth substrate and possible influence of melanin composition on its interaction with ionizing radiation. XTT/MTT assays showed increased metabolic activity of melanized C. neoformans cells relative to non-melanized cells, and exposure to ionizing radiation enhanced the electron-transfer properties of melanin in melanized cells. Melanized Wangiella dermatitidis and Cryptococcus neoformans cells exposed to ionizing radiation approximately 500 times higher than background grew significantly faster as indicated by higher CFUs, more dry weight biomass and 3-fold greater incorporation of (14)C-acetate than non-irradiated melanized cells or irradiated albino mutants. In addition, radiation enhanced the growth of melanized Cladosporium sphaerospermum cells under limited nutrients conditions. CONCLUSIONS/SIGNIFICANCE: Exposure of melanin to ionizing radiation, and possibly other forms of electromagnetic radiation, changes its electronic properties. Melanized fungal cells manifested increased growth relative to non-melanized cells after exposure to ionizing radiation, raising intriguing questions about a potential role for melanin in energy capture and utilization

Pre-Clinical Evaluation of a 213Bi-Labeled 2556 Antibody to HIV-1 gp41 Glycoprotein in HIV-1 Mouse Models as a Reagent for HIV Eradication

Any strategy for curing HIV infection must include a method to eliminate viral-infected cells. Based on our earlier proof-of-principle results targeting HIV-1 infected cells with radiolabeled antibody (mAb) to gp41 viral antigen, we embarked on identifying a suitable candidate mAb for preclinical development.Among the several human mAbs to gp41 tested, mAb 2556 was found to have high affinity, reactivity with multimeric forms of gp41 present on both the surface of virus particles and cells expressing HIV-1 Env, and recognition of a highly conserved epitope of gp41 shared by all HIV-1 subtypes. Also, mAb 2556 was the best in competition with HIV-1+ serum antibodies, which is an extremely important consideration for efficacy in the treatment of HIV patients. When radiolabeled with alpha-emitting radionuclide 213-Bismuth ((213)Bi) - (213)Bi-2556 efficiently and specifically killed ACH-2 human lymphocytes chronically infected with HIV-1, and HIV-1 infected human peripheral blood mononuclear cells (hPBMCs). The number of binding sites for (213)Bi-2556 on the surface of the infected cells was >10(6). The in vivo experiments were performed in two HIV-1 mouse models--splenic and intraperitoneal. In both models, the decrease in HIV-1 infected hPBMCs from the spleens and peritoneum, respectively, was dose-dependent with the most pronounced killing of hPBMCs observed in the 100 µCi (213)Bi-2556 group (P = 0.01). Measurement of the blood platelet counts and gross pathology of the treated mice demonstrated the lack of toxicity for (213)Bi-2556.We describe the preclinical development of a novel radiolabeled mAb reagent that could potentially be part of an HIV eradication strategy that is ready for translation into the clinic as the next step in its development. As viral antigens are very different from "self" human antigens - this approach promises high selectivity, increased efficacy and low toxicity, especially in comparison to immunotoxins

The second International Symposium on Fungal Stress: ISFUS

The topic of ‘fungal stress’ is central to many important disciplines, including medical mycology, chronobiology, plant and insect pathology, industrial microbiology, material sciences, and astrobiology. The International Symposium on Fungal Stress (ISFUS) brought together researchers, who study fungal stress in a variety of fields. The second ISFUS was held in May 8-11 2017 in Goiania, Goiás, Brazil and hosted by the Instituto de Patologia Tropical e Saúde Pública at the Universidade Federal de Goiás. It was supported by grants from CAPES and FAPEG. Twenty-seven speakers from 15 countries presented their research related to fungal stress biology. The Symposium was divided into seven topics: 1. Fungal biology in extreme environments; 2. Stress mechanisms and responses in fungi: molecular biology, biochemistry, biophysics, and cellular biology; 3. Fungal photobiology in the context of stress; 4. Role of stress in fungal pathogenesis; 5. Fungal stress and bioremediation; 6. Fungal stress in agriculture and forestry; and 7. Fungal stress in industrial applications. This article provides an overview of the science presented and discussed at ISFUS-2017.Sao Paulo Research Foundation (FAPESP) 2010/06374-1, 2013/50518-6, 2014/01229-4Brazilian National Council for Scientific and Technological Development (CNPq) PQ2 302312/2011-0, PQ1D 308436/2014-8Coordenação de Aperfeiçoãmento de Pessoal de Nível Superior (CAPES) PAEP 88881.123209/2016-01Fundação de Amparo à Pesquisa do Estado de Goiás Brazil 20171026700011

Protection of Melanized Cryptococcus neoformans from Lethal Dose Gamma Irradiation Involves Changes in Melanin\u27s Chemical Structure and Paramagnetism

Certain fungi thrive in highly radioactive environments including the defunct Chernobyl nuclear reactor. Cryptococcus neoformans (C. neoformans), which uses L-3,4-dihydroxyphenylalanine (L-DOPA) to produce melanin, was used here to investigate how gamma radiation under aqueous aerobic conditions affects the properties of melanin, with the aim of gaining insight into its radioprotective role. Exposure of melanized fungal cell in aqueous suspensions to doses of γ-radiation capable of killing 50 to 80% of the cells did not lead to a detectable loss of melanin integrity according to EPR spectra of melanin radicals. Moreover, upon UV-visible (Xe-lamp) illumination of melanized cells, the increase in radical population was unchanged after γ-irradiation. Gamma-irradiation of frozen cell suspensions and storage of samples for several days at 77 K however, produced melanin modification noted by a reduced radical population and reduced photoresponse. More direct evidence for structural modification of melanin came from the detection of soluble products with absorbance maxima near 260 nm in supernatants collected after γ-irradiation of cells and cell-free melanin. These products, which include thiobarbituric acid (TBA)-reactive aldehydes, were also generated by Fenton reagent treatment of cells and cell-free melanin. In an assay of melanin integrity based on the metal (Bi+3) binding capacity of cells, no detectable loss in binding was detected after γ-irradiation. Our results show that melanin in C. neoformans cells is susceptible to some damage by hydroxyl radical formed in lethal radioactive aqueous environments and serves a protective role in melanized fungi that involves sacrificial breakdown

Physico-Chemical Evaluation of Rationally Designed Melanins as Novel Nature-Inspired Radioprotectors

Melanin, a high-molecular weight pigment that is ubiquitous in nature, protects melanized microorganisms against high doses of ionizing radiation. However, the physics of melanin interaction with ionizing radiation is unknown.We rationally designed melanins from either 5-S-cysteinyl-DOPA, L-cysteine/L-DOPA, or L-DOPA with diverse structures as shown by elemental analysis and HPLC. Sulfur-containing melanins had higher predicted attenuation coefficients than non-sulfur-containing melanins. All synthetic melanins displayed strong electron paramagnetic resonance (2.14.10(18), 7.09.10(18), and 9.05.10(17) spins/g, respectively), with sulfur-containing melanins demonstrating more complex spectra and higher numbers of stable free radicals. There was no change in the quality or quantity of the stable free radicals after high-dose (30,000 cGy), high-energy ((137)Cs, 661.6 keV) irradiation, indicating a high degree of radical stability as well as a robust resistance to the ionizing effects of gamma irradiation. The rationally designed melanins protected mammalian cells against ionizing radiation of different energies.We propose that due to melanin's numerous aromatic oligomers containing multiple pi-electron system, a generated Compton recoil electron gradually loses energy while passing through the pigment, until its energy is sufficiently low that it can be trapped by stable free radicals present in the pigment. Controlled dissipation of high-energy recoil electrons by melanin prevents secondary ionizations and the generation of damaging free radical species

Protection of Melanized Cryptococcus neoformans from Lethal Dose Gamma Irradiation Involves Changes in Melanin's Chemical Structure and Paramagnetism

Certain fungi thrive in highly radioactive environments including the defunct Chernobyl nuclear reactor. Cryptococcus neoformans (C. neoformans), which uses L-3,4-dihydroxyphenylalanine (L-DOPA) to produce melanin, was used here to investigate how gamma radiation under aqueous aerobic conditions affects the properties of melanin, with the aim of gaining insight into its radioprotective role. Exposure of melanized fungal cell in aqueous suspensions to doses of γ-radiation capable of killing 50 to 80% of the cells did not lead to a detectable loss of melanin integrity according to EPR spectra of melanin radicals. Moreover, upon UV-visible (Xe-lamp) illumination of melanized cells, the increase in radical population was unchanged after γ-irradiation. Gamma-irradiation of frozen cell suspensions and storage of samples for several days at 77 K however, produced melanin modification noted by a reduced radical population and reduced photoresponse. More direct evidence for structural modification of melanin came from the detection of soluble products with absorbance maxima near 260 nm in supernatants collected after γ-irradiation of cells and cell-free melanin. These products, which include thiobarbituric acid (TBA)-reactive aldehydes, were also generated by Fenton reagent treatment of cells and cell-free melanin. In an assay of melanin integrity based on the metal (Bi+3) binding capacity of cells, no detectable loss in binding was detected after γ-irradiation. Our results show that melanin in C. neoformans cells is susceptible to some damage by hydroxyl radical formed in lethal radioactive aqueous environments and serves a protective role in melanized fungi that involves sacrificial breakdown

Human monoclonal antibodies against Staphylococcus aureus surface antigens recognize in vitro and in vivo biofilm

Implant-associated Staphylococcus aureus infections are difficult to treat because of biofilm formation. Bacteria in a biofilm are often insensitive to antibiotics and host immunity. Monoclonal antibodies (mAbs) could provide an alternative approach to improve the diagnosis and potential treatment of biofilm-related infections. Here, we show that mAbs targeting common surface components of S. aureus can recognize clinically relevant biofilm types. The mAbs were also shown to bind a collection of clinical isolates derived from different biofilm-associated infections (endocarditis, prosthetic joint, catheter). We identify two groups of antibodies: one group that uniquely binds S. aureus in biofilm state and one that recognizes S. aureus in both biofilm and planktonic state. Furthermore, we show that a mAb recognizing wall teichoic acid (clone 4497) specifically localizes to a subcutaneously implanted pre-colonized catheter in mice. In conclusion, we demonstrate the capacity of several human mAbs to detect S. aureus biofilms in vitro and in vivo