Maxygen (United States)

To evaluate the safety of Aliya pulsed electric field (PEF) ablation in lung lesions at the time of biopsy before first-line standard of care (SOC) therapy in patients with metastatic cancer. Eligible patients requiring diagnostic biopsy for suspected metastatic lung lesions were enrolled. Treatment-naive lesions ≤5cm were PEF ablated bronchoscopically or percutaneously. Procedure and/or device-related adverse events (AE) or serious adverse events (SAEs) and their effects on subsequent therapy were assessed from PEF ablation through 30 days. Forty-four lesions from 30 enrolled patients were ablated using PEF with 3-dimensional image guidance immediately following diagnostic biopsy under general anesthesia. Ninety percent (27/30) were ablated bronchoscopically with robotic navigation. One to 3 lesions (range: 0.4 to 2.4cm longest diameter) were ablated per patient, including 17 within 1cm of the pleura or a fissure. One procedure-related SAE (pneumothorax) occurred at the time of percutaneous biopsy and PEF ablation and resolved without sequelae. No AE impacted planned therapy initiation. Limited changes to the lung parenchyma in and around the PEF ablation zone were observed on CT scans 30 days post-PEF ablation. This study demonstrates PEF ablation of lung lesions at the time of diagnostic biopsy is safe. One PEF-related SAE occurred within 30 days of PEF and resolved without sequelae. Ablation adjacent to sensitive structures was feasible, safe, and without impact on subsequent therapy.

Abstract Rust fungi comprise thousands of species many of which cause disease on important crop plants. The flax rust fungus Melampsora lini has been a model species for the genetic dissection of plant immunity since the 1940s, however the highly fragmented and incomplete reference genome has so far hindered progress in effector gene discovery. Here, we generate a fully-phased, chromosome-scale assembly of the two nuclear genomes of M. lini strain CH5, resolving an additional 320 Mbp of sequence compared to the previous short-read assembly. The 482 Mbp dikaryotic genome is at least 79% repetitive with a large proportion (∼40%) of the genome comprised of young, highly similar transposable elements. The assembly resolves the known effector gene loci some of which carry complex duplications that were collapsed in the previous assembly. Using a genetic map followed by manual correction of gene models, we identify the AvrM3 and AvrN genes which encode unusually large fungal effector proteins and trigger defense responses when co-expressed with the corresponding resistance genes in Nicotiana tabacum . We locate the genes linked to the tetrapolar mating system on chromosomes 4 and 9, but in contrast to the cereal rusts which have one pheromone receptor gene per haplotype, in flax rust three pheromone receptor genes are found with two of them closely linked on one haplotype. Taken together, we show that a high-quality assembly is crucial for resolving complex gene loci and given the increasing number of fungal effectors of large size, the commonly applied criterion for effector candidates as being small proteins as needs to be reconsidered.

BackgroundICB have shown remarkable clinical efficacy with durable responses in many cancers. However, many patients are resistant, or develop resistance to ICB. For these patients’ treatment options are limited. Thus, treatments to overcome ICB resistance are necessary.WU-NK-101 (WUNK) is a PBMC-derived, cytokine-reprogrammed, expanded, and cryopreserved off-the-shelf memory-like (ML) NK cell product. WUNK exhibits enhanced cytotoxicity, metabolic fitness/flexibility, and resistance to tumor microenvironment (TME) immunosuppression compared to conventional NK cells.1 2 WUNK cells are active against multiple solid tumor cell lines in vitro and demonstrate additive cytotoxicity in combination with ICB. Furthermore, ML-NK cell treatment in vivo promotes T cell migration/activation in the TME, IFN-γ and chemokine release, and MHC-I and PD-L1 upregulation on residual tumor cells.3MethodsHerein, we investigate the utility of WUNK in reversing ICB, namely pembrolizumab, resistance. We report that WUNK therapy rescues pembrolizumab failure and show that combination of WUNK and ICB synergize leading to increased cytotoxicity in a novel xenograft mouse model of immune escape.ResultsMDA-MB-231 TNBC cells and human PBMC’s were co-engrafted in NSG MHC-I/II dKO mice. At baseline 88.7±02% of tumor cells expressed MHC-I molecules and 7.6±0.4% were positive for PD-L1. Tumor engraftment was noted on day 2, while the presence of hPBMC in the circulation was confirmed by D7 with continued expansion (1000-fold) through day 40. Initial tumor control, ~40-fold decrease in tumor burden, was noted between D7–16. However, after two weeks, tumor growth resumed and correlated with a 68% decrease in MHC-1 and ~52% increase of PD-L1 positivity in tumor cells indicating escape from immune surveillance. Administration of pembrolizumab (5mg/kg dose) at the time of immune evasion (D18) resulted in tumor growth inhibition (TGI=13.8%; figure 1). Remarkably, co-administration of WUNK with pembrolizumab resulted in a synergistic activity, significantly reducing tumor growth (TGI=94.4%; figure 1) and improving overall survival. Further histological analysis of tumor tissue confirmed enhanced infiltration of T-cells, and interaction between ML-NK cells within the TME.ConclusionsIn summary, using a novel model of ICB resistance, our study demonstrates that treatment with WUNK led to increased infiltration and activation of T-cells into the TME. Furthermore, WUNK synergized with ICB to enhance anti-tumor activity and rescue pembrolizumab resistance leading to a durable adaptive response. Overall, these findings provide a strong rationale for exploring the combination of WUNK with ICBs clinically to overcome immune evasion and improve patient outcomes.ReferencesMuth JJ, Mathyer ME, Cruz B, DuFour C, Leedom T, Hamil AS, Pouyanfard S, Berrien-Elliot M, Luukkonen L, Tran M, Tumala B, Pomper N, Arthur L, Foster M, Gorrochategui J, Pennacchi PC, Primo D, Ballesteros J, Tolstikov V, Kiebish MA, Shah P, Vadakekolathu J, Rutella S, Fehniger TA, Sullivan R, Kabakibi A, Cooper ML, Davidson-Moncada J; WU-NK-101, an allogeneic memory NK cell, for the treatment of relapse or refractory (R/R) acute myeloid leukemia (AML). HemaSphere 2022;6(S3):2497.Rutella S, Muth J, Vadakekulathu J, Mathyer ME, Tumala BR, Foster M, Baughman J, Pennacchi PC, Tolstikov V, Carter A, Primo D, Gorrochategui J, Magee K, Shah P, Kiebish M, Ballesteros J, Cooper ML, Berrien-Elliott M, Fehniger TA, Davidson-Moncada JK; 11P WU-NK-101: An enhanced NK cell therapy optimized for function in the tumor microenvironment (TME). Annals of Oncology 2022;33(S7):S549–S550.Leedom TA, Muz B, Vadakekolathu J, Muth JJ, Li X-H, Watson G, Magee K, Sullivan RP, Berrien-Elliott M, Fenigher T, Rutella S, Cooper ML, Kabakibi A, Davidson-Moncada JK; 6418: xWU-NK-101 as salvage therapy post immune checkpoint blockade (ICB). Cancer Res 1 April 2023;83(7_Supplement):6418. 332 Figure 1In vivo xenograft model of immune surveillance and escape. Engrafted human PBMCs initially control MDA-MB-231 tumor growth before immune evasion occurs (Day 18) and tumor growth resumes. Treatment initiation at the time of immune evasion resulted in synergistic anti-tumor activity when WUNK was combined with Pembrolizumab compared to each delivered as a monotherapy.

BackgroundCytokine-induced memory NK cells, like WU-NK-101, offer several advantages over conventional NK cells.1, 2 These advantages include enhanced functional persistence, efficacy, and metabolic fitness that improve their survival and activity in liquid and solid tumor microenvironments.3, 4 Additionally, unlike conventional NK (cNK) cells derived from iPSC, cord blood, or adult peripheral blood, WU-NK-101 does not require engineering to enable anti-tumor activity.The MonetaTM Platform is a feeder cell-free system of fusion protein complexes to generate, expand, phenotypically maintain, and cryopreserve memory NK cells for an ‘off-the-shelf’ allogeneic drug product. Here we define the molecular characteristics of WU-NK-101, through RNA and protein phenotyping, and elucidate pathways involved in memory NK cell expansion and activity.MethodsWU-NK-101 was generated from previously frozen NK cells derived from healthy donor whole blood using the MonetaTM platform. We evaluated pre- and post-expansion phenotype by flow cytometry, CyTOF, and RNA-Seq. We further assessed post-thaw functionality by cytokine secretion and cancer cell cytotoxicity in the absence or presence of monoclonal antibodies.ResultsCyTOF analysis revealed that WU-NK-101 have an expression profile distinct from cNK. Compared to cNK, WU-NK-101 have elevated expression of Memory NK markers (CD25, CD69, NKG2A), cytotoxic molecules (GzmB, TRAIL), activating receptors (NKp30, NKp44, NKG2D), and nutrient transporters (CD71, CD98), providing mechanistic rationale for their enhanced anti-tumor activity and metabolic flexibility. Flow cytometry and RNAseq analysis confirm this phenotype. When stimulated by cancer cells, WU-NK-101 have enhanced secretion of IFN-γ, MIP-1α, and TNFα; and also demonstrate improved cytotoxicity compared to cNK. WU-NK-101 is able to utilize monoclonal antibodies to effectively drive antibody-dependent cellular cytotoxicity against solid tumor cancer cell lines.ConclusionsThe MonetaTM platform expands memory NK cells while maintaining their cytokine-induced memory phenotype, as identified by RNA-Seq, flow cytometry, and CyTOF. As a result, WU-NK-101 demonstrates improved anti-tumor activity compared to cNK cells, even within the adverse tumor microenvironment. These data support the clinical development of WU-NK-101, an allogeneic Memory NK cell therapy in both liquid and in solid tumors, as a monotherapy and in combination with monoclonal antibodies, solid tumor engagers, or other anti-tumor modalities.ReferencesRomee R, et al. Cytokine-induced memory-like natural killer cells exhibit enhanced responses against myeloid leukemia. Science translational medicine 8.357 (2016): 357ra123–357ra123.Berrien-Elliott, M, et al. Multidimensional Analyses of Donor Memory-Like NK Cells Reveal New Associations with Response after Adoptive Immunotherapy for Leukemia Cancer discovery 10.12 (2020): 1854–1871.Terren I, et al. Metabolic changes of Interleukin-12/15/18-stimulated human NK cells. Sci Rep 2021Rutella S, et al. WU-NK-101: An Enhanced NK Cell Therapy Optimized for Function in the Tumor Microenvironment (TME). ESMO Congress (2022).

Abstract The European Parliament identifies virgin olive oil (VOO) as one of the foods which are often subject to fraudulent activities. Possibilities of adulteration are the application of illegal soft deodorization of extra virgin olive oil (EVOO) or the commercialization of blends of EVOO with soft‐deodorized EVOO or refined vegetable oils. Despite the search for possibilities to prove the illegal soft deodorization of EVOO or the addition of cheaper vegetable oils to EVOO, suitable methods are still missing. Therefore, the aim of the study is to develop a new analytical and statistical approach addressing detection of mild deodorization or addition of refined foreign oils. For this purpose, VOOs are treated in lab‐scale for 1 h up to 28 days at different temperatures (20, 50, 60, 80,100, 110, and 170 °C) in order to simulate and study the effect of heat treatment on known analytical parameters by near infrared spectroscopy (NIR). A logit regression model enabling the calculation of the probability for a heat treatment is developed. This new methodology allows detecting both soft deodorized olive oils and blends of EVOO with cheaper full refined vegetable oils. Adding only 10% of full refined oil could be detected in extra VOO.Practical Applications: NIR methods combined with chemometrics have become one of the most attractive analytical tools to control quality of food. It is a simple, precise, and rapid method. All relevant analytical parameters of oxidative and thermal fat degradation can be determined in a single run and be used to detect adulterated virgin olive oils (VOOs). The use of a simple equation developed from the logistic regression using peroxide value, K‐values, p‐anisidine value, pyropheophytine, 1,2‐diacylglycerols, total polar compounds and monomeric oxidized triacylglycerols, and other well‐known parameters allows to detect mild deodorized olive oils or also blends of VOO with soft‐deodorized ones or the addition of low amounts of foreign vegetable oils. This technique has potential to be used as a screening method for the detection of adulterated olive oils using both the traditional laboratory methods and the corresponding NIR‐methods.

Abstract Hepatocellular Carcinoma (HCC) and Endometrioid Endometrial Carcinoma (EEC) are two lethal diseases of public health importance worldwide. Using QIAGEN’s RNA sequencing solution, we were able to highlight key molecular and biological processes that indicate similarities in the tumor progression toward metastasis between one EEC patient and the three HCC patients. We have identified ITGB1-001 isoform as a potential common biomarker between these two complex cancers. We have also shown these two cancers share activated pathways such as actin cytoskeleton signaling, and RAF1 as an upstream regulator indicating similar transcriptional program between these patients. Furthermore, we have identified as a potential therapeutic target the master regulator CTGF that connects a network of 286 downstream targets to invasion of tumor cells. We have filtered a set of unique variants in the EEC patient that affect a network of upstream regulators common with HCC. This approach may be useful in the context of precision or personalized medicine. Examining the gene expression in tumors from groups of patients with each disease revealed that at a molecular level, early stages of EEC resemble established HBV-positive, HCV-negative, liver cirrhosis positive HCC. Citation Format: Jean-Noel Billaud, Stuart Tugendreich, Debra Toburen. Integrative approach to biomarker discovery by performing comparative analysis of two cancers, hepatocellular carcinoma and endometrioid endometrial carcinoma, using genetics and transcriptomics from RNA sequencing data [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2840. doi:10.1158/1538-7445.AM2017-2840

Immunization against tumor-associated antigens is a promising approach to cancer therapy and prevention, but it faces several challenges and limitations, such as tolerance mechanisms associated with self-antigens expressed by the tumor cells. Costimulatory molecules B7.1 (CD80) and B7.2 (CD86) have improved the efficacy of gene-based and cell-based vaccines in animal models and are under investigation in clinical trials. However, their efficacy as vaccine adjuvants is likely limited by the fact that they mediate both stimulatory and inhibitory signals to T cells via CD28 and CTLA-4, respectively. To overcome these limitations, we have generated a B7.1-like, chimeric costimulatory molecule with preferential binding to CD28, named CD28-binding protein (CD28BP), which we combined with a modified, nonself tumor antigen variant of epithelial cell adhesion molecule (EpCAM), named TAg25. TAg25 induced a cross-reactive immune response against human wild-type EpCAM upon DNA vaccination in cynomolgus monkeys. However, TAg25 DNA immunization alone or in combination with human (h) B7.1 induced no detectable antigen-specific T cells in the peripheral blood of the animals. In contrast, TAg25 combined with CD28BP induced both CD4 and CD8 T cells specific for EpCAM. Moreover, TAg25 combined with CD28BP induced significantly higher levels of EpCAM-specific antibodies than TAg25 plus hB7.1. These improved adjuvant properties of CD28BP, when compared with hB7.1, illustrate the importance of CD28 costimulation in vaccine responses in nonhuman primates and warrant further studies on the potential of CD28BP in improving the efficacy of cancer vaccines.