Reversal of cancer cachexia and muscle wasting by ActRIIB antagonism leads to prolonged survival.

Muscle wasting and cachexia have long been postulated to be key determinants of cancer-related death, but there has been no direct experimental evidence to substantiate this hypothesis. Here, we show that in several cancer cachexia models, pharmacological blockade of ActRIIB pathway not only prevents further muscle wasting but also completely reverses prior loss of skeletal muscle and cancer-induced cardiac atrophy. This treatment dramatically prolongs survival, even of animals in which tumor growth is not inhibited and fat loss and production of proinflammatory cytokines are not reduced. ActRIIB pathway blockade abolished the activation of the ubiquitin-proteasome system and the induction of atrophy-specific ubiquitin ligases in muscles and also markedly stimulated muscle stem cell growth. These findings establish a crucial link between activation of the ActRIIB pathway and the development of cancer cachexia. Thus ActRIIB antagonism is a promising new approach for treating cancer cachexia, whose inhibition per se prolongs survival.

Integrated COVID-19 Interventions in a Native American Community: Arizona, December 25, 2021-January 31, 2022.

COVID-19 has disproportionately affected Indigenous communities. The Whiteriver Service Unit (WRSU) took an integrated public health-health care system delivery approach in collaboration with the White Mountain Apache Tribe to decrease the case fatality rate (CFR). The WRSU performed daily data analyses identifying risk factors, expeditiously treating and proactively vaccinating people during at-home visits. The WRSU's CFR was 0.3% lower than Arizona's (P = .04). Among communities disproportionally affected, an integrated approach using data to drive real-time decision-making among a culturally competent workforce can contribute to decreased CFR. (Am J Public Health. 2023;113(10):1089-1092. https://doi.org/10.2105/AJPH.2023.307364).

Rational cytokine design for increased lifetime and enhanced potency using pH-activated "histidine switching".

We describe a method for the rational design of more effective therapeutic proteins using amino acid substitutions that reduce receptor binding affinity in intracellular endosomal compartments, thereby leading to increased recycling in the ligand-sorting process and consequently resulting in longer half-life in extracellular medium. We demonstrate this approach for granulocyte colony-stimulating factor by using computationally predicted histidine substitutions that switch protonation states between cell-surface and endosomal pH. Molecular modeling of binding electrostatics indicates two different single-histidine mutants that fulfill our design requirements; experimental assays demonstrate that each mutant indeed exhibits an order-of-magnitude increase in medium half-life along with enhanced potency due to increased endocytic recycling.

Rationale-Based Engineering of a Potent Long-Acting FGF21 Analog for the Treatment of Type 2 Diabetes.

Fibroblast growth factor 21 (FGF21) is a promising drug candidate for the treatment of type 2 diabetes. However, the use of wild type native FGF21 is challenging due to several limitations. Among these are its short half-life, its susceptibility to in vivo proteolytic degradation and its propensity to in vitro aggregation. We here describe a rationale-based protein engineering approach to generate a potent long-acting FGF21 analog with improved resistance to proteolysis and aggregation. A recombinant Fc-FGF21 fusion protein was constructed by fusing the Fc domain of human IgG1 to the N-terminus of human mature FGF21 via a linker peptide. The Fc positioned at the N-terminus was determined to be superior to the C-terminus as the N-terminal Fc fusion retained the ßKlotho binding affinity and the in vitro and in vivo potency similar to native FGF21. Two specific point mutations were introduced into FGF21. The leucine to arginine substitution at position 98 (L98R) suppressed FGF21 aggregation at high concentrations and elevated temperatures. The proline to glycine replacement at position 171 (P171G) eliminated a site-specific proteolytic cleavage of FGF21 identified in mice and cynomolgus monkeys. The derived Fc-FGF21(RG) molecule demonstrated a significantly improved circulating half-life while maintaining the in vitro activity similar to that of wild type protein. The half-life of Fc-FGF21(RG) was 11 h in mice and 30 h in monkeys as compared to 1-2 h for native FGF21 or Fc-FGF21 wild type. A single administration of Fc-FGF21(RG) in diabetic mice resulted in a sustained reduction in blood glucose levels and body weight gains up to 5-7 days, whereas the efficacy of FGF21 or Fc-FGF21 lasted only for 1 day. In summary, we engineered a potent and efficacious long-acting FGF21 analog with a favorable pharmaceutical property for potential clinical development.

Light-induced aggregation of type I soluble tumor necrosis factor receptor.

We evaluated the effect of UV-B light at 302 nm on a model therapeutic protein, 2.6 D type I soluble tumor necrosis factor receptor (sTNF-RI). This protein contains a single Trp at position 97 and seven native disulfide bonds along its interior from the N to the C-terminus. At a protein concentration of 0.1 mg/mL photoirradiation was found to induce the formation of soluble disulfide cross-linked dimers with greater levels of these species formed at pH 8 than at pH 5. Intermolecular disulfide formation was also directly correlated with the photoinduced unfolding of the protein as measured by changes in secondary structure by CD spectroscopy. Trp was implicated as the initiator of the observed photoreactions by the detection of the Trp oxidation products and the absence of dimer formation when Trp97 was replaced with Gln. Reactive oxygen species or triplet state species of Trp were not involved in the reaction suggesting that disulfides were cleaved through one-electron reduction by either hydrated or peptide bound electrons produced by the photoirradiated Trp resulting in thiyl radical formation with disruption of the protein structure and intermolecular cross-linking. Photodegradation was not prevented by deoxygenation, methionine or sucrose commonly used for formulation of biopharmaceuticals. To our knowledge this is the first report directly documenting disulfide mediated aggregation through thiyl radical formation initiated by photoirradiation of Trp.

Regulatory effects of novel neurotrophin-1/b cell-stimulating factor-3 (cardiotrophin-like cytokine) on B cell function.

We describe regulatory effects that a novel neurotrophin-1/B cell-stimulating factor-3 (NNT-1/BSF-3; also reported as cardiotrophin-like cytokine) has on B cell function. NNT-1/BSF-3 stimulates B cell proliferation and Ig production in vitro. NNT-1/BSF-3-transgenic mice, engineered to express NNT-1/BSF-3 in the liver under control of the apolipoprotein E promoter, show B cell hyperplasia with particular expansion of the mature follicular B cell subset in the spleen and the prominent presence of plasma cells. NNT-1/BSF-3-transgenic mice show high serum levels of IgM, IgE, IgG2b, IgG3, anti-dsDNA Abs, and serum amyloid A. NNT-1/BSF-3-transgenic mice also show non-amyloid mesangial deposits that contain IgM, IgG, and C3 and are characterized by a distinctive ultrastructure similar to that of immunotactoid glomerulopathy. NNT-1/BSF-3-transgenic mice produce high amounts of Ag-specific IgM, IgA, and IgE and low amounts of IgG2a and IgG3. Normal mice treated with NNT-1/BSF-3 also produce high amounts of Ag-specific IgE. NNT-1/BSF-3 regulates immunity by stimulating B cell function and Ab production, with preference for Th2 over Th1 Ig types.