Leveraging PET to image folate receptor α therapy of an antibody-drug conjugate

Background: The folate receptor α (FRα)-targeting antibody-drug conjugate (ADC), IMGN853, shows great antitumor activity against FRα-expressing tumors in vivo, but patient selection and consequently therapy outcome are based on immunohistochemistry. The aim of this study is to develop an antibody-derived immuno-PET imaging agent strategy for targeting FRα in ovarian cancer as a predictor of treatment success. Methods: We developed [89Zr]Zr-DFO-M9346A, a humanized antibody-based radiotracer targeting tumorassociated FRα in the preclinical setting. [89Zr]Zr-DFO-M9346A’s binding ability was tested in an in vitro uptake assay using cell lines with varying FRα expression levels. The diagnostic potential of [89Zr]Zr-M9346A was evaluated in KB and OV90 subcutaneous xenografts. Following intravenous injection of [89Zr]Zr-DFO-M9346A (~90 μCi, 50 μg), PET imaging and biodistribution studies were performed. We determined the blood half-life of [89Zr]Zr-DFO-M9346A and compared it to the therapeutic, radioiodinated ADC [131I]-IMGN853. Finally, in vivo studies using IMG853 as a therapeutic, paired with [89Zr]Zr-DFO-M9346A as a companion diagnostic were performed using OV90 xenografts. Results: DFO-M9346A was labeled with Zr-89 at 37 °C within 60 min and isolated in labeling yields of 85.7 ± 5.7%, radiochemical purities of 98.0 ± 0.7%, and specific activities of 3.08 ± 0.43 mCi/mg. We observed high specificity for binding FRα positive cells in vitro. For PET and biodistribution studies, [89Zr]Zr-M9346A displayed remarkable in vivo performance in terms of excellent tumor uptake for KB and OV xenografts (45.8 ± 29.0 %IA/g and 26.1 ± 7.2 %IA/g), with low non-target tissue uptake in other organs such as kidneys (4.5 ± 1.2 %IA/g and 4.3 ± 0.7 %IA/g). A direct comparison of the blood half life of [89Zr]Zr-M9346A and [131I]-IMGN853 corroborated the equivalency of the radiopharmaceutical and the ADC, paving the way for a companion PET imaging study. Conclusions: We developed a new folate receptor-targeted 89Zr-labeled PET imaging agent with excellent pharmacokinetics in vivo. Good tumor uptake in subcutaneous KB and OV90 xenografts were obtained, and ADC therapy studies were performed with the precision predictor

Effect of mixing and feed batch sequencing on the prevalence and distribution of African swine fever virus in swine feed

It is critical to have methods that can detect and mitigate the risk of African swine fever virus (ASFV) in potentially contaminated feed or ingredients bound for the United States. The purpose of this work was to evaluate feed batch sequencing as a mitigation technique for ASFV contamination in a feed mill, and to determine if a feed sampling method could identify ASFV following experimental inoculation. Batches of feed were manufactured in a BSL-3Ag room at Kansas State University's Biosafety Research Institute in Manhattan, Kansas. First, the pilot feed manufacturing system mixed, conveyed, and discharged an ASFV-free diet. Next, a diet was manufactured using the same equipment, but contained feed inoculated with ASFV for final concentration of 5.6 × 104 TCID50/g. Then, four subsequent ASFV-free batches of feed were manufactured. After discharging each batch into a collection container, 10 samples were collected in a double ‘X’ pattern. Samples were analysed using a qPCR assay for ASFV p72 gene then the cycle threshold (Ct) and Log10 genomic copy number (CN)/g of feed were determined. The qPCR Ct values (p < .0001) and the Log10 genomic CN/g (p < .0001) content of feed samples were impacted based on the batch of feed. Feed samples obtained after manufacturing the ASFV-contaminated diet contained the greatest amounts of ASFV p72 DNA across all criteria (p < .05). Quantity of ASFV p72 DNA decreased sequentially as additional batches of feed were manufactured, but was still detectable after batch sequence 4. This subsampling method was able to identify ASFV genetic material in feed samples using p72 qPCR. In summary, sequencing batches of feed decreases concentration of ASFV contamination in feed, but does not eliminate it. Bulk ingredients can be accurately evaluated for ASFV contamination by collecting 10 subsamples using the sampling method described herein. Future research is needed to evaluate if different mitigation techniques can reduce ASFV feed contamination

DNA Methylation Profiles of Ovarian Clear Cell Carcinoma

BACKGROUND: Ovarian clear cell carcinoma (OCCC) is a rare ovarian cancer histotype that tends to be resistant to standard platinum-based chemotherapeutics. We sought to better understand the role of DNA methylation in clinical and biological subclassification of OCCC. METHODS: We interrogated genome-wide methylation using DNA from fresh frozen tumors from 271 cases, applied non-smooth non-negative matrix factorization (nsNMF) clustering, and evaluated clinical associations and biological pathways. RESULTS: Two approximately equally sized clusters that associated with several clinical features were identified. Compared to Cluster 2 (N=137), Cluster 1 cases (N=134) presented at a more advanced stage, were less likely to be of Asian ancestry, and tended to have poorer outcomes including macroscopic residual disease following primary debulking surgery (p-values <0.10). Subset analyses of targeted tumor sequencing and immunohistochemical data revealed that Cluster 1 tumors showed TP53 mutation and abnormal p53 expression, and Cluster 2 tumors showed aneuploidy and ARID1A/PIK3CA mutation (p-values <0.05). Cluster-defining CpGs included 1,388 CpGs residing within 200 bp of the transcription start sites of 977 genes; 38% of these genes (N=369 genes) were differentially expressed across cluster in transcriptomic subset analysis (p-values <10(−4)). Differentially expressed genes were enriched for six immune-related pathways, including interferon alpha and gamma responses (p-values < 10(−6)). CONCLUSIONS: DNA methylation clusters in OCCC correlate with disease features and gene expression patterns among immune pathways. IMPACT: This work serves as a foundation for integrative analyses that better understand the complex biology of OCCC in an effort to improve potential for development of targeted therapeutics

Prevalence and Distribution of African Swine Fever Virus in Swine Feed After Mixing and Feed Batch Sequencing

As the United States maintains trade with countries where African swine fever virus (ASFV) is endemic, it is critical to have methods that can detect and mitigate the risk of ASFV in potentially contaminated feed or ingredients. Therefore, the objectives of this study were to 1) evaluate feed batch sequencing as a mitigation technique for ASFV contamination in a feed mill, and 2) determine if a feed sampling method could identify ASFV following experimental inoculation. Batches of feed were manufactured in a BSL-3Ag room at Kansas State University’s Biosafety Research Institute in Manhattan, KS. First, the pilot feed manufacturing system mixed, conveyed, and discharged an ASFV-free diet. Next, a diet was manufactured using the same equipment, but contained feed inoculated with ASFV for a final concentration of 5.6 × 104 TCID50/g. Then, four subsequent ASFV-free batches of feed were manufactured. After discharging each batch into a biohazard tote, 10 samples were collected in a double ‘X’ pattern. Samples were analyzed using a qPCR assay specific for the ASFV p72 gene to determine the cycle threshold (Ct) and log10 genomic copy number (CN)/g of feed. Batch of feed affected the qPCR Ct values (P \u3c 0.0001) and the log10 genomic CN/g (P \u3c 0.0001) content of feed. Feed samples obtained after manufacturing the ASFV-contaminated diet contained the greatest (P \u3c 0.05) amounts of ASFV p72 DNA across all criteria. Quantity of ASFV p72 DNA decreased sequentially as additional batches of initially ASFV-free feed were manufactured, but it was still detectable after batch sequence 4, suggesting cross contamination between batches. This subsampling method was able to identify ASFV genetic material in feed samples using the PCR assay specific for the ASFV p72 gene. In summary, sequencing batches of feed decreases concentration of ASFV contamination in feed, but does not eliminate it. Bulk ingredients or feed can be accurately evaluated for ASFV contamination by collecting 10 evenly distributed subsamples, representing 0.05% of the volume of the container, using the sampling method described herein

Evaluating the distribution of African swine fever virus within a feed mill environment following manufacture of inoculated feed

11 Pág. Centro de Investigación en Sanidad Animal (CISA)It is critical to understand the role feed manufacturing may have regarding potential African swine fever virus (ASFV) transmission, especially given the evidence that feed and/or ingredients may be potential vectors. The objective of the study was to evaluate the distribution of ASFV in a feed mill following manufacture of contaminated feed. To accomplish this, a pilot-scale feed mill consisting of a mixer, bucket elevator, and spouting was constructed in a BSL-3Ag facility. First, a batch of ASFV-free feed was manufactured, followed by a batch of feed that had an ASFV-contaminated ingredient added to feed, which was then mixed and discharged from the equipment. Subsequently, four additional ASFV-free batches of feed were manufactured using the same equipment. Environmental swabs from 18 locations within the BSL-3Ag room were collected after each batch of feed was discharged. The locations of the swabs were categorized into four zones: 1) feed contact surface, 2) non-feed contact surface 1 meter from feed, and 4) transient surfaces. Environmental swabs were analyzed using a qPCR specific for the ASFV p72 gene and reported as genomic copy number (CN)/mL of environmental swab processing buffer. Genomic copies were transformed with a log10 function for statistical analysis. There was no evidence of a zone × batch interaction for log10 genomic CN/mL (P = 0.625) or cycle threshold (Ct) value (P = 0.608). Sampling zone impacted the log10 p72 genomic CN/mL (P < 0.0001) and Ct values (P < 0.0001), with a greater amount of viral genome detected on transient surfaces compared to other surfaces (P < 0.05). This study illustrates that once ASFV enters the feed mill environment it becomes widespread and movement of people can significantly contribute to the spread of ASFV in a feed mill environment.Funding for this work was obtained from the NBAF Transition Funds from the state of Kansas (JAR), the National Pork Board under award number 20-018 (CKJ), the Department of Homeland Security Center of Excellence for Emerging and Zoonotic Animal Diseases under grant number HSHQDC 16-A-B0006 (JAR), and the AMP Core of the NIGMS COBRE Center on Emerging and Zoonotic Infectious Diseases (CEZID) under award number P20GM13044 (JAR)Peer reviewe

Heterogeneous Tumor-Immune Microenvironments among Differentially Growing Metastases in an Ovarian Cancer Patient.

We present an exceptional case of a patient with high-grade serous ovarian cancer, treated with multiple chemotherapy regimens, who exhibited regression of some metastatic lesions with concomitant progression of other lesions during a treatment-free period. Using immunogenomic approaches, we found that progressing metastases were characterized by immune cell exclusion, whereas regressing and stable metastases were infiltrated by CD8+ and CD4+ T cells and exhibited oligoclonal expansion of specific T cell subsets. We also detected CD8+ T cell reactivity against predicted neoepitopes after isolation of cells from a blood sample taken almost 3 years after the tumors were resected. These findings suggest that multiple distinct tumor immune microenvironments co-exist within a single individual and may explain in part the heterogeneous fates of metastatic lesions often observed in the clinic post-therapy. VIDEO ABSTRACT.Cancer Research UK core grant (C14303/A17197), Memorial Sloan Kettering Cancer Cencter core grant (P30 CA008748

Association of cetuximab with adverse pulmonary events in cancer patients: a comprehensive review

Compounds derived from biologic sources, or biologicals, are increasingly utilized as therapeutic agents in malignancy. Development of anti-cancer targeted therapies from biologics is increasingly being utilized. Cetuximab, a chimeric monoclonal antibody, is one such anti-cancer targeted therapeutic that has shown efficacy in quelling the rate of patient decline in colorectal, head/neck, and non-small cell lung cancer. However, due to the relatively recent addition of biologic compounds to the therapeutic arsenal, information related to adverse reactions is less well known than those seen in traditional chemotherapeutics. Dermatologic reactions have been demonstrated as the most frequent side effect cited during cetuximab therapy for malignancy; however, other effects may lead to greater morbidity. In general, pulmonary complications of therapeutics can lead to significant morbidity and mortality. The purpose of this review is to compile the various pulmonary side effects seen in patients treated with cetuximab for various malignancies, and to compare the incidence of these adverse reactions to standard therapies

Diverse BRCA1 and BRCA2 Reversion Mutations in Circulating Cell-Free DNA of Therapy-Resistant Breast or Ovarian Cancer

Purpose:; Resistance to platinum-based chemotherapy or PARP inhibition in germline; BRCA1; or; BRCA2; mutation carriers may occur through somatic reversion mutations or intragenic deletions that restore BRCA1 or BRCA2 function. We assessed whether; BRCA1/2; reversion mutations could be identified in circulating cell-free DNA (cfDNA) of patients with ovarian or breast cancer previously treated with platinum and/or PARP inhibitors.; Experimental Design:; cfDNA from 24 prospectively accrued patients with germline; BRCA1; or; BRCA2; mutations, including 19 patients with platinum-resistant/refractory ovarian cancer and five patients with platinum and/or PARP inhibitor pretreated metastatic breast cancer, was subjected to massively parallel sequencing targeting all exons of 141 genes and all exons and introns of; BRCA1; and; BRCA2; Functional studies were performed to assess the impact of the putative; BRCA1/2; reversion mutations on BRCA1/2 function.; Results:; Diverse and often polyclonal putative; BRCA1; or; BRCA2; reversion mutations were identified in cfDNA from four patients with ovarian cancer (21%) and from two patients with breast cancer (40%).; BRCA2; reversion mutations were detected in cfDNA prior to PARP inhibitor treatment in a patient with breast cancer who did not respond to treatment and were enriched in plasma samples after PARP inhibitor therapy. Foci formation and immunoprecipitation assays suggest that a subset of the putative reversion mutations restored BRCA1/2 function.; Conclusions:; Putative; BRCA1/2; reversion mutations can be detected by cfDNA sequencing analysis in patients with ovarian and breast cancer. Our findings warrant further investigation of cfDNA sequencing to identify putative; BRCA1/2; reversion mutations and to aid the selection of patients for PARP inhibition therapy.; Clin Cancer Res; 23(21); 6708-20. ©2017 AACR;

A united statement of the global chiropractic research community against the pseudoscientific claim that chiropractic care boosts immunity.

BACKGROUND: In the midst of the coronavirus pandemic, the International Chiropractors Association (ICA) posted reports claiming that chiropractic care can impact the immune system. These claims clash with recommendations from the World Health Organization and World Federation of Chiropractic. We discuss the scientific validity of the claims made in these ICA reports. MAIN BODY: We reviewed the two reports posted by the ICA on their website on March 20 and March 28, 2020. We explored the method used to develop the claim that chiropractic adjustments impact the immune system and discuss the scientific merit of that claim. We provide a response to the ICA reports and explain why this claim lacks scientific credibility and is dangerous to the public. More than 150 researchers from 11 countries reviewed and endorsed our response. CONCLUSION: In their reports, the ICA provided no valid clinical scientific evidence that chiropractic care can impact the immune system. We call on regulatory authorities and professional leaders to take robust political and regulatory action against those claiming that chiropractic adjustments have a clinical impact on the immune system