SignPath Pharma (United States)

Composition of the microbiota in various sections of the gastrointestinal tract of the wild pheasant Phasianus colchicus, collected in the Krasnodar region, was characterized for the first time using high-throughput 16S rRNA gene amplicon sequencing. This species is an important target of sport hunting, an indicator of habitat biocoenosis preservation, and a promising alternative to traditional poultry farming due to the high nutritional value and hypoallergenic properties of its products. The microbiota of wild species exhibited greater diversity and an increased representation of Coriobacteriaceae and Bifidobacteriaceae compared to the microbiota of birds raised under intensive farming conditions. The lower intestine, particularly the cecum, showed a higher presence of these families, whose members are known for their probiotic properties. Assessing the indigenous microbiota of the wild pheasant is of interest for understanding the associations between microbiota and host immunity and as a source of potential probiotic strains for veterinary biopreparations. The highest proportion of potentially probiotic bacteria from the families Coriobacteriaceae, Lactobacteriaceae, and Bifidobacteriaceae was found in the cecal microbiota.

Upon the development of biosimilar drugs, the structural, physical-chemical, and functional similarity to the original drug should be ensured. We have proposed a simple and rapid HPLC-UV method for assessing the content of C-terminal Arg166 in darbepoetin alfa samples, in particular after their isolation from the culture fluid. All the stages of the described method (from the sample extraction to obtaining a chromatographic profile) take about 5 h. The proposed approach will be useful for laboratories developing a technology for producing recombinant darbepoetin alfa, to assess the quality of the product, and for subsequent optimization of the production process.

The 2024 Alzheimer's Association (AA) research diagnostic criteria for Alzheimer's Disease (AD) considers fluid biomarkers, including promising blood-based biomarkers for detecting AD. This study aims to identify dementia subtypes and their cognitive and neuroimaging profiles in older adults with dementia in the Democratic Republic of Congo (DRC) using biomarkers and clinical data. Forty-five individuals with dementia over 65 years old were evaluated using the Community Screening Instrument for Dementia and the informant-based Alzheimer's Questionnaire. Core AD biomarkers (Aβ42/40 and p-tau181) and non-specific neurodegeneration biomarkers (NfL, GFAP) were measured in blood plasma. Neuroimaging structures were assessed using magnetic resonance imaging (MRI). Dementia subtypes were determined based on plasma biomarker pathology and vascular markers. Biomarker cutoff scores were identified to optimize sensitivity and specificity. Individuals were stratified into one of four dementia subtypes - AD only, non-AD vascular, non-AD other, or mixed - based on combinations of abnormalities in these markers. Among the 45 individuals with dementia, mixed dementia had the highest prevalence (42.4%), followed by AD-only (24.4%), non-AD other dementia (22.2%), and non-AD vascular dementia subtypes (11.1%). Both cognitive and neuroimaging profiles aligned poorly with biomarker classifications in the full sample. Cognitive tests varied across dementia subtypes. The cognitive profile of the AD-only and mixed groups suggested relatively low cognitive performance, while the non-AD and other groups had the best scores on average. Consistent with studies in other settings, our preliminary findings suggest that neurodegenerative plasma biomarkers may help to identify dementia subtypes and provide insight into cognitive and neuroimaging profiles among older adults in the DRC.

Abstract Background: HER2 expression level is a key factor in determining the optimal treatment course for breast cancer patients. Roughly 15% of breast cancers are HER2(+), and determination of HER2 status is routinely assessed by immunohistochemistry (IHC). Accurate assessment of the HER2 IHC score (0, 1+, 2+, 3+) by pathologists is therefore critical, especially in light of novel therapeutic approaches demonstrating efficacy in the HER2-low setting (IHC scores 1+, and 2+/FISH-)1,2. To assist pathologists with the consistent provision of reproducible and accurate scores across the entire HER2 scoring range, we developed a machine-learning model (“AIM-HER2”) to generate accurate, slide-level HER2 scores aligned with ASCO-CAP guidelines in clinical breast cancer HER2 IHC specimens. Methods: AIM-HER2 was developed using whole-slide images (WSI; N=4261) from clinical and commercial sources. WSI were split into training (N=2694, 63%) and optimization (N=1567, 37%) sets. An additive multiple instance learning (aMIL) model3 was trained to predict HER2 scores directly from WSI and create interpretable heatmaps that depict HER2 predictions in tissue images. Image artifacts and in situ carcinomas were identified using previously trained artifact and tissue segmentation models and were excluded, leaving only regions of invasive carcinoma to be analyzed. AIM-HER2 performance was assessed on additional slides obtained from five academic or commercial sources (N=804 total, 770 evaluable) on which HER2 IHC was performed. Board-certified pathologists (N=52) with relevant experience provided manual HER2 scores based on ASCO-CAP guidelines. Nested pairwise non-inferiority analysis4 was used to compare model performance to that of pathologists (N=3 pathologists per slide). In the nested pairwise framework, agreement among pathologists was compared to agreement between AIM-HER2 and pathologists via linear kappa, so that summary metrics account for inter-pathologist variability. Results: High concordance was observed between AIM-HER2-predicted and pathologist-labeled slide-level HER2 scores, both overall and for each scoring level. Similar results were observed when assessing AIM-HER2 performance on multiple slide scanners and after IHC with multiple HER2 IHC antibody clones. Results are summarized in Table 1. Conclusions: We developed AIM-HER2, a novel aMIL-based approach for predicting slide-level HER2 IHC scores. AIM-HER2 has similar levels of agreement with pathologists as pathologists have with each other for determining HER2 score. This result is upheld when slides imaged using multiple scanning platforms and stained using multiple HER2 antibody clones. The performance of AIM-HER2 on multiple scanners and after multiple assays supports broad applicability of this algorithm in clinical laboratories, including for the identification of HER2-low cases. Work is ongoing to perform similar analyses in an independent, real-world dataset.

We detail a cross-sectional, multi-plasma biomarker characterization of the AIBL cohort samples using plasma Aβ42/40 ratio, pTau217, pTau181, GFAP, and Nf-L assays across the Alzheimer's disease (AD) continuum, and determine discriminatory performance with CNS amyloid PET imaging status.

Abstract Introduction/Objective Cancer Cytogenetics (CG) and Fluorescence in-situ Hybridization (FISH) testing are essential parts of molecular genetics (MG) in clinical diagnosis for patients suffering from hematopoietic disorders. Even though MG assays are clinically offered, and technical expertise is available, the need for expertise in CG and FISH cannot be understated. Due to budget constraints, deviated focus on innovation by academic programs and lack of awareness in the undergraduate programs, only a handful of centers offer formal training programs. Therefore, the need for qualified CG and FISH technologists to support the industry pipeline is continuously needed. Methods/Case Report With the increased workload and the need to combat workforce shortages, we continue to support academic centers as an on-site training facility. In addition, we have initiated academic collaborations with accredited training programs, both national and international, by training of our own technicians with a pilot program. These individuals hold bachelor’s degrees in biological sciences and are qualified to perform high-complexity testing under Clinical Laboratory Improvement Amendments (CLIA) and College of American Pathologists (CAP) regulations. Results (if a Case Study enter NA) Since 2009, we have had 32 qualified CG/FISH technologists with a CG (ASCP) board certification, most of whom graduated through academic programs. We took the initiative to train four staff members who acquired job experience while working in our CLIA/CAP accredited lab. After the ASCP year requirement was fulfilled, they became eligible to take the ASCP registry exam in CG. All four have passed and are certified cytogenetic technologists. Conclusion Challenged by a reduction in training programs and increasing demand in the industry, we have been successful in training and certifying our technologists with academic and private collaborations. More collaborative efforts are needed to fulfill such an important need in the field of CG.

e14525 Background: The efficacy of tumor-infiltrating lymphocyte (TIL) therapy is limited by the rarity of anti-tumor TCRs and exhausted status of the product. Its improvement should focus on a high frequency of diverse anti-tumor TCRs with stem-like phenotype, which can be achieved synthetically by identifying multiple anti-tumor TCRs from the patient's intra-tumor TCR repertoire and subsequently engineering them into the patient’s peripheral T cells. Such a ‘synthetic TIL’ is cost-prohibitive using existing technologies. To enable a synthetic TIL product, we developed ultra-high throughput TCR gene synthesis and screening methods to synthesize and screen > 50,000 TCRs from scRNA-Seq data from > 30 patients and identified > 100 tumor/NeoAg-reactive TCRs. The gene expression patterns of the T cells carrying such experimentally validated tumor/NeoAg-reactive TCRs gave rise to an algorithm called Interpreter to predict potentially anti-tumor TCRs from scRNA-Seq data. Methods: To translate synthetic TIL to the clinic, we developed a method to generate up to 100 sequence-verified TCR-encoding plasmids (selected by Interpreter) from fresh tumor biopsy in just 3 weeks. Our GMP-grade production uses a collective knock-in approach, where a mixture of up to 100 plasmids (each encoding a distinct TCR) at equal proportion are packaged into AAV which serve as template to precisely insert the TCR genes into a primarily monoallelic locus and simultaneously inhibit the endogenous TCR. This gene editing is characterized by a modified scRNA-Seq method called EntrySeq. In first-in-human, N-of-1 trials, 109 to 6x109 synthetic TIL cells were administered to solid tumor patients with or without lymphodepletion and IL2. An investigator-initiated Phase 1 trial was then opened to evaluate safety, preliminary efficacy, and cellular activity of synthetic TIL and to determine RP2D and MTD. Subjects with relapsed, unresectable locally advanced solid tumors who received at least 1 standard systemic therapy are allowed to enroll in this study. Those patients with no standard treatment regimens available, or those who are unable to obtain effective treatment due to economic conditions are also permitted. Results: EntrySeq confirmed that > 80% of the T cells express only 1 TCR, avoiding mispairing of different TCRs. The frequencies of different TCRs were largely uniform and were stable through plasmid, AAV and T-cell mixtures. Up to 1.5x1010 synthetic TIL cells were produced from a patient’s leukopak. 40 to 60% of the T cells in the product for infusion express an exogenous TCR. In first-in-human trials, the administered T cells showed favorable persistence and safety profile. The Phase 1 trial is ongoing. Conclusions: We established a fast, affordable, and GMP-compliant process to manufacture synthetic TIL products, enabling a Phase 1 trial.