Early Normal Tissue Effects and Bone Marrow Relative Biological Effectiveness for an Actinium 225-Labeled HER2/neu-Targeting Antibody.

PURPOSE: In this study we determined the dose-independent relative biological effectiveness (RBE2) of bone marrow for an anti-HER2/neu antibody labeled with the alpha-particle emitter actinium 225 (225Ac). Hematologic toxicity is often a consequence of radiopharmaceutical therapy (RPT) administration, and dosimetric guidance to the bone marrow is required to limit toxicity. METHODS AND MATERIALS: Female neu/N transgenic mice (MMTV-neu) were intravenously injected with 0 to 16.65 kBq of the alpha-particle emitter labeled antibody, 225Ac-DOTA-7.16.4, and euthanized at 1 to 9 days after treatment. Complete blood counts were performed. Femurs and tibias were collected, and bone marrow was isolated from 1 femur and tibia and counted for radioactivity. Contralateral intact femurs were fixed, decalcified, and assessed by histology. Marrow cellularity was the biologic endpoint selected for RBE2 determination. For the reference radiation, both femurs of the mice were photon irradiated with 0 to 5 Gy using a small animal radiation research platform. RESULTS: Response as measured by cellularity for the alpha-particle emitter RPT (αRPT) RPT and the external beam radiation therapy were linear and linear quadratic, respectively, as a function of absorbed dose. The resulting dose-independent RBE2 for bone marrow was 6. CONCLUSIONS: As αRPT gains prominence, preclinical studies evaluating RBE in vivo will be important in relating to human experience with beta-particle emitter RPT. Such normal tissue RBE evaluations will help mitigate unexpected toxicity in αRPT.

Bone Marrow Relative Biological Effectiveness for a 212Pb-labeled Anti-HER2/neu Antibody.

PURPOSE: We have determined the in vivo relative biological effectiveness (RBE) of an alpha-particle-emitting radiopharmaceutical therapeutic agent (212Pb-labeled anti-HER2/neu antibody) for the bone marrow, a potentially dose-limiting normal tissue. METHODS AND MATERIALS: The RBE was measured in mice using femur marrow cellularity as the biological endpoint. External beam radiation therapy (EBRT), delivered by a small-animal radiation research platform was used as the reference radiation. Alpha-particle emissions were delivered by 212Bi after the decay of its parent nuclide 212Pb, which was conjugated onto an anti-HER2/neu antibody. The alpha-particle absorbed dose to the marrow after an intravenous administration (tail vein) of 122.1 to 921.3 kBq 212Pb-TCMC-7.16.4 was calculated. The mice were sacrificed at 0 to 7 days after treatment and the radioactivity from the femur bone marrow was measured. Changes in marrow cellularity were assessed by histopathology. RESULTS: The dose response for EBRT and 212Pb-anti-HER2/neu antibody were linear-quadratic and linear, respectively. On transforming the EBRT dose-response relationship into a linear relationship using the equivalent dose in 2-Gy fractions of external beam radiation formalism, we obtained an RBE (denoted RBE2) of 6.4, which is independent of cellularity and absorbed dose. CONCLUSIONS: Because hematologic toxicity is dose limiting in almost all antibody-based RPT, in vivo measurements of RBE are important in helping identify an initial administered activity in phase 1 escalation trials. Applying the RBE2 and assuming typical antibody clearance kinetics (biological half-life of 48 hours), using a modified blood-based dosimetry method, an average administered activity of approximately 185.5 MBq (5.0 mCi) per patient could be administered before hematologic toxicity is anticipated.

Differential gene expression and Ingenuity Pathway Analysis of bronchoalveolar lavage cells from horses with mild/moderate neutrophilic or mastocytic inflammation on BAL cytology.

Mild to moderate equine asthma syndrome (mEAS) affects horses of all ages and breeds. To date, the etiology and pathophysiology of mEAS are active areas of research, and it remains incompletely understood whether mEAS horses with different immune cell 'signatures' on BAL cytology represent different phenotypes, distinct pathobiological mechanisms (endotypes), varied environmental conditions, disease severity, genetic predispositions, or all of the above. In this descriptive study, we compared gene expression data from BAL cells isolated from horses with normal BALF cytology (n = 5), to those isolated from horses with mild/moderate neutrophilic inflammation (n = 5), or mild/moderate mastocytic inflammation (n = 5). BAL cell protein lysates were analyzed for cytokine/chemokine levels using Multiplex Bead Immunoassay, and for select proteins using immunoblot. The transcriptome, determined by RNA-seq and analyzed with DEseq2, contained 20, 63, and 102 significantly differentially expressed genes in horses with normal vs. neutrophilic, normal vs. mastocytic, and neutrophilic vs. mastocytic BALF cytology, respectively. Pathway analyses revealed that BAL-isolated cells from horses with neutrophilic vs. normal cytology showed enrichment in inflammation pathways, and horses with mastocytic vs. normal cytology showed enrichment in pathways involved in fibrosis and allergic reaction. BAL cells from horses with mastocytic mEAS, compared to neutrophilic mEAS, showed enrichment in pathways involved in alteration of tissue structures. Cytokine analysis determined that IL-1ß was significantly different in the lysates from horses with neutrophilic inflammation compared to those with normal or mastocytic BAL cytology. Immunoblot revealed significant difference in the relative level of MMP2 in horses with neutrophilic vs. mastocytic mEAS. Upregulation of mRNA transcripts involved in the IL-1 family cytokine signaling axis (IL1a, IL1b, and IL1R2) in neutrophilic mEAS, as well as KIT mRNA in mastocytic mEAS, are novel, potentially clinically relevant, findings of this study. These findings further inform our understanding of inflammatory cell subtypes in mEAS.

The Role of Neutrophils in the Pathophysiology of Asthma in Humans and Horses.

Asthma is a common and debilitating chronic airway disease that affects people and horses of all ages worldwide. While asthma in humans most commonly involves an excessive type 2 immune response and eosinophilic inflammation, neutrophils have also been recognized as key players in the pathophysiology of asthma, including in the severe asthma phenotype where neutrophilic inflammation predominates. Severe equine asthma syndrome (sEAS) features prominent neutrophilic inflammation and has been increasingly used as a naturally occurring animal model for the study of human neutrophilic asthma. This comparative review examines the recent literature in order to explore the role of neutrophil inflammatory functions in the pathophysiology and immunology of asthma in humans and horses.

Comparative Review of Asthma in Farmers and Horses.

PURPOSE OF REVIEW: Farmers are routinely exposed to organic dusts and aeroallergens that can have adverse respiratory health effects including asthma. Horses are farm-reared large animals with similar exposures and can develop equine asthma syndrome (EAS). This review aims to compare the etiology, pathophysiology, and immunology of asthma in horses compared to farmers and highlights the horse as a potential translational animal model for organic dust-induced asthma in humans. RECENT FINDINGS: Severe EAS shares many clinical and pathological features with various phenotypes of human asthma including allergic, non-allergic, late onset, and severe asthma. EAS disease features include variable airflow obstruction, cough, airway hyperresponsiveness, airway inflammation/remodeling, neutrophilic infiltrates, excess mucus production, and chronic innate immune activation. Severe EAS is a naturally occurring and biologically relevant, translational animal disease model that could contribute to a more thorough understanding of the environmental and immunologic factors contributing to organic dust-induced asthma in humans.

Bronchoalveolar Lavage Cytology Characteristics and Seasonal Changes in a Herd of Pastured Teaching Horses.

Equine asthma syndrome (EAS) is a common problem that affects horses of any age. Severe EAS is reported to affect 10-20% of adult horses in the northern hemisphere, while mild/moderate EAS is reported to affect 60-100% of adult horses, depending on the population and geographic region. For both severe and mild/moderate EAS, the presence of lower airway inflammation is attributed to airborne "triggers" such as dust, mold, and bacterial components that horses encounter in hay and stable-environments; and treatment recommendations for horses with EAS often include full-time pasture turnout. The caveat to this recommendation is horses with summer-pasture associated EAS (SP-EAS), who experience allergic lower airway inflammation when exposed to summer pasture. The prevalence of EAS in horses on pasture that do not have SP-EAS has not been reported. The purpose of this study was to use bronchoalveolar lavage (BAL) cytology to determine the prevalence of EAS in a herd of pastured, adult research horses with no history of respiratory disease. The horses were members of a teaching animal herd housed on pasture in the southeastern United States and fed round-bale Bermuda-grass hay. BAL fluid (BALF) cytology was analyzed in both summer (May-August 2017) and winter (November 2017-February 2018). Similar to previous reports, the prevalence of severe EAS in our study population was 10% in summer and 4.3% in winter. The prevalence of mild/moderate EAS was 60% in summer and 87% in winter. The high prevalence of mild/moderate EAS in this population was unexpected, given the 24-h, year-round pasture environment and the lack of history of respiratory disease. Additionally, 61.1% of horses with both summer and winter data had a different BALF cytology profile between the two seasons. To the authors' knowledge, this is the first study to use BAL cytology to diagnose, and monitor changes in, EAS phenotype in pastured adult horses. These results help to inform discussions regarding prevalence of EAS in pastured, adult horses in the southeastern region of North America.