Mechanistic regulation of HERV activation in tumors and implications for translational research in oncology.

Transcription of distinct loci of human endogenous retroviruses (HERVs) and in some cases, translation of these transcripts have been consistently observed in many types of cancer. It is still debated whether HERV activation serves as a trigger for carcinogenesis or rather occurs as a consequence of epigenetic alterations and other molecular sequelae that characterize cellular transformation. Here we review the known molecular and epigenetic mechanisms of HERV activation in cancer cells as well as its potential contribution to carcinogenesis. Further, we describe the use of HERV expression in cancer diagnostic and characterize the potential of HERV-derived antigens to serve as novel targets for cancer immunotherapy. We believe this review, which summarizes both what is known as well as unknown in this rapidly developing field, will boost interest in research on the therapeutic potential of targeting HERV elements in tumors and the impact of HERV activation in oncogenesis.

Granulocyte transfusions in severe aplastic anemia.

Patients with severe aplastic anemia (SAA) are at high risk for morbidity and mortality due to severe infections. We aimed to characterize the role of granulocyte transfusion (GT) in SAA. Primary outcomes were survival from first GT, including overall survival (OS) at last follow up, survival to discharge, and receipt of HSCT. Secondary outcomes included evaluation of clinical response at 7 and 30 days after GT initiation based on a clinical scoring system incorporating microbiological and radiographic response. Twenty-eight SAA patients underwent 30 GT courses with a per-dose median of 1.28 x 109 granulocyte cells/kilogram (range 0.45-4.52 x 109). OS from initial GT to median last follow up (551 days) was 50%, with 39% (11/28) alive at last follow up. Sixty-four percent (18/28) of all patients survived to hospital discharge. Patients with complete, partial, or stable response at 30 days had significantly improved OS compared to non-responders (p=0.0004). Eighty-six percent (18/21) of patients awaiting HSCT during GT underwent transplant and 62% (13/21) survived to post-HSCT discharge. Sex, type of infection, or percentage of days with absolute neutrophil count > 0.2x109/L during GT course were not predictive of survival (p=0.52, p=0.7, p=0.28). Nine of 28 (32%) patients developed new or increased human leukocyte antigen (HLA) alloimmunization during their GT course. GTs in SAA may impact survival in those with improvement or stabilization of their underlying infection. Alloimmunization can occur and OS in this population remains poor, but GTs may be a useful tool to bridge patients to curative treatment with HSCT.

Forging international collaboration and alliances to establish the largest transplant center in the north of Vietnam.

Through collaboration with international experts, our institution established a highly active and successful hematopoietic stem cell transplant program, providing access to this potentially curative treatment modality for patients with a variety of benign and malignant hematological diseases. The initial development of an autologous stem cell transplant program provided our institution with the infrastructure, equipment, and expertise needed for the subsequent development of an allogeneic stem cell transplant program. Key transplant staff received training from international transplant experts at the NHLBI/NIH, the Mayo Clinic, the Johns Hopkins Hospital, and Nagoya Japan, providing them with the expertise to conduct a variety of different transplant approaches, including PBSC transplants from HLA-matched relatives, unrelated cord blood transplants, haploidentical transplants, and CD34 selected stem cell transplants. Patient characteristics were varied among all groups. The number of allogeneic and autologous transplants performed at the NIHBT has increased steadily every year since the initiation of our transplant program. By 2022, 547 transplant procedures had been performed, including 268 autologous and 279 allogeneic transplants. Allogeneic transplants were performed for both malignant and nonmalignant hematological diseases, with acute leukemia (AL) being the most common indication for allogeneic HCT. The majority of recipients undergoing allogeneic transplantation received G-CSF mobilized PBSC allografts from either HLA identical or haplo-identical relatives, with a smaller percentage of patients receiving a UCB transplant or a PBSC allograft that had been CD34+ selected. Amongst the 279 recipients of an allogeneic transplant, mortality rates within day 100 and beyond day 100 were 12.6% and 26.2% respectively. Overall survival (OS) and event-free survival at 5 years in benign and malignant subgroups were 81% and 73% vs 52% and 48% respectively. Through collaboration with international transplant experts, the National Institute of Hematology and Blood Transfusion in Hanoi has stood up the most active transplant center in the northern region of Vietnam. Patients coming from low-income financial backgrounds are now able to receive a variety of different state-of-the-art transplant approaches that are affordable and have been associated with excellent long-term outcomes.

Social Determinants modulate NK cell activity via obesity, LDL, and DUSP1 signaling.

Adverse social determinants of health (aSDoH) are associated with obesity and related comorbidities like diabetes, cardiovascular disease, and cancer. Obesity is also associated with natural killer cell (NK) dysregulation, suggesting a potential mechanistic link. Therefore, we measured NK phenotypes and function in a cohort of African-American (AA) women from resource-limited neighborhoods. Obesity was associated with reduced NK cytotoxicity and a shift towards a regulatory phenotype. In vitro, LDL promoted NK dysfunction, implicating hyperlipidemia as a mediator of obesity-related immune dysregulation. Dual specific phosphatase 1 (DUSP1) was induced by LDL and was upregulated in NK cells from subjects with obesity, implicating DUSP1 in obesity-mediated NK dysfunction. In vitro, DUSP1 repressed LAMP1/CD107a, depleting NK cells of functional lysosomes to prevent degranulation and cytokine secretion. Together, these data provide novel mechanistic links between aSDoH, obesity, and immune dysregulation that could be leveraged to improve outcomes in marginalized populations.

Spleen tyrosine kinase inhibition restores myeloid homeostasis in COVID-19.

Spleen tyrosine kinase (SYK) is a previously unidentified therapeutic target that inhibits neutrophil and macrophage activation in coronavirus disease 2019 (COVID-19). Fostamatinib, a SYK inhibitor, was studied in a phase 2 placebo-controlled randomized clinical trial and was associated with improvements in many secondary end points related to efficacy. Here, we used a multiomic approach to evaluate cellular and soluble immune mediator responses of patients enrolled in this trial. We demonstrated that SYK inhibition was associated with reduced neutrophil activation, increased circulation of mature neutrophils (CD10+CD33-), and decreased circulation of low-density granulocytes and polymorphonuclear myeloid-derived suppressor cells (HLA-DR-CD33+CD11b-). SYK inhibition was also associated with normalization of transcriptional activity in circulating monocytes relative to healthy controls, an increase in frequency of circulating nonclassical and HLA-DRhi classical monocyte populations, and restoration of interferon responses. Together, these data suggest that SYK inhibition may mitigate proinflammatory myeloid cellular and soluble mediator responses thought to contribute to immunopathogenesis of severe COVID-19.

Expanded NK cells used for adoptive cell therapy maintain diverse clonality and contain long-lived memory-like NK cell populations.

Multiple clinical trials exploring the potential of adoptive natural killer (NK) cell therapy for cancer have employed ex vivo expansion using feeder cells to obtain large numbers of NK cells. We have previously utilized the rhesus macaque model to clonally track the NK cell progeny of barcode-transduced CD34+ stem and progenitor cells after transplant. In this study, NK cells from barcoded rhesus macaques were used to study the changes in NK cell clonal patterns that occurred during ex vivo expansion using culture protocols similar to those employed in clinical preparation of human NK cells including irradiated lymphoblastoid cell line (LCL) feeder cells or K562 cells expressing 4-1BBL and membrane-bound interleukin-21 (IL-21). NK expansion cultures resulted in the proliferation of clonally diverse NK cells, which, at day 14 harvest, contained greater than 50% of the starting barcode repertoire. Diversity as measured by Shannon index was maintained after culture. With both LCL and K562 feeders, proliferation of long-lived putative memory-like NK cell clones was observed, with these clones continuing to constitute a mean of 31% of the total repertoire of expanded cells. These experiments provide insight into the clonal makeup of expanded NK cell clinical products.

Healthcare worker safety program in a coronavirus disease 2019 (COVID-19) alternate care site: The Javits New York Medical Station experience.

OBJECTIVE: In March 2020, New York City became the epicenter of the coronavirus disease 2019 (COVID-19) pandemic in the United States. Because healthcare facilities were overwhelmed with patients, the Jacob K. Javits Convention Center was transformed into the nation's largest alternate care site: Javits New York Medical Station (hereafter termed Javits). Protecting healthcare workers (HCWs) during a global shortage of personal protective equipment (PPE) in a nontraditional healthcare setting posed unique challenges. We describe components of the HCW safety program implemented at Javits. SETTING: Javits, a large convention center transformed into a field hospital, with clinical staff from the US Public Health Service Commissioned Corps and the US Department of Defense. METHODS: Key strategies to ensure HCW safety included ensuring 1-way flow of traffic on and off the patient floor, developing a matrix detailing PPE required for each work activity and location, PPE extended use and reuse protocols, personnel training, and monitoring adherence to PPE donning/doffing protocols when entering or exiting the patient floor. Javits staff who reported COVID-19 symptoms were immediately isolated, monitored, and offered a severe acute respiratory coronavirus virus 2 (SARS-CoV-2) reverse-transcriptase polymerase chain reaction (RT-PCR) test. CONCLUSIONS: A well-designed and implemented HCW safety plan can minimize the risk of SARS-CoV-2 infection for HCWs. The lessons learned from operating the nation's largest COVID-19 alternate care site can be adapted to other environments during public health emergencies.

Concurrent transposon engineering and CRISPR/Cas9 genome editing of primary CLL-1 chimeric antigen receptor-natural killer cells.

BACKGROUND: Natural killer (NK) cell genome editing promises to enhance the innate and alloreactive anti-tumor potential of NK cell adoptive transfer. DNA transposons are versatile non-viral gene vectors now being adapted to primary NK cells, representing important tools for research and clinical product development. AIMS AND METHODS: We set out to generate donor-derived, primary chimeric antigen receptor (CAR)-NK cells by combining the TcBuster transposon system with Epstein-Barr virus-transformed lymphoblastoid feeder cell-mediated activation and expansion. RESULTS: This approach allowed for clinically relevant NK-cell expansion capability and CAR expression, which was further enhanced by immunomagnetic selection based on binding to the CAR target protein.The resulting CAR-NK cells targeting the myeloid associated antigen CLL-1 efficiently targeted CLL-1-positive AML cell lines and primary AML populations, including a population enriched for leukemia stem cells. Subsequently, concurrent delivery of CRISPR/Cas9 cargo was applied to knockout the NK cell cytokine checkpoint cytokine-inducible SH2-containing protein (CIS, product of the CISH gene), resulting in enhanced cytotoxicity and an altered NK cell phenotype. CONCLUSIONS: This report contributes a promising application of transposon engineering to donor-derived NK cells and emphasizes the importance of feeder mediated NK cell activation and expansion to current protocols.

Predictors of clonal evolution and myeloid neoplasia following immunosuppressive therapy in severe aplastic anemia.

Predictors, genetic characteristics, and long-term outcomes of patients with SAA who clonally evolved after immunosuppressive therapy (IST) were assessed. SAA patients were treated with IST from 1989-2020. Clonal evolution was categorized as "high-risk" (overt myeloid neoplasm [meeting WHO criteria for dysplasia, MPN or acute leukemia] or isolated chromosome-7 abnormality/complex karyotype without dysplasia or overt myeloid neoplasia) or "low-risk" (non-7 or non-complex chromosome abnormalities without morphological evidence of dysplasia or myeloid neoplasia). Univariate and multivariate analysis using Fine-Gray competing risk regression model determined predictors. Long-term outcomes included relapse, overall survival (OS) and hematopoietic stem cell transplant (HSCT). Somatic mutations in myeloid cancer genes were assessed in evolvers and in 407 patients 6 months after IST. Of 663 SAA patients, 95 developed clonal evolution. Pre-treatment age >48 years and ANC > 0.87 × 109/L were strong predictors of high-risk evolution. OS was 37% in high-risk clonal evolution by 5 years compared to 94% in low-risk. High-risk patients who underwent HSCT had improved OS. Eltrombopag did not increase high-risk evolution. Splicing factors and RUNX1 somatic variants were detected exclusively at high-risk evolution; DNMT3A, BCOR/L1 and ASXL1 were present in both. RUNX1, splicing factors and ASXL1 somatic mutations detected at 6 months after IST predicted high-risk evolution.

Graft-Versus-Solid-Tumor Effect: From Hematopoietic Stem Cell Transplantation to Adoptive Cell Therapies.

After allogeneic hematopoietic stem cell transplantation (HSCT), donor lymphocytes may contribute to the regression of hematological malignancies and select solid tumors, a phenomenon referred to as the graft-versus-tumor effect (GVT). However, this immunologic reaction is frequently limited by either poor specificity resulting in graft-versus-host disease or the frequency of tumor-specific T cells being too low to induce a complete and sustained anti-tumor response. Over the past 2 decades, it has become clear that the driver of GVT following allogeneic HSCT is T-cell-mediated recognition of antigens presented on tumor cells. With that regard, even though the excitement for using HSCT in solid tumors has declined, clinical trials of HSCT in solid tumors provided proof of concept and valuable insights leading to the discovery of tumor antigens and the development of targeted adoptive cell therapies for cancer. In this article, we review the results of clinical trials of allogeneic HSCT in solid tumors. We focus on lessons learned from correlative studies of these trials that hold the potential for the creation of tumor-specific immunotherapies with greater efficacy and safety for the treatment of malignancies.

Harnessing natural killer cells for the treatment of multiple myeloma.

Multiple myeloma (MM) is hematologic malignancy that is associated with profound immune alterations. Myeloma cells are susceptible to killing by natural killer (NK) cells but acquire the ability to elude NK cell surveillance by avoiding recognition and suppressing NK cell function. Major advances in the treatment of multiple myeloma have been achieved by effective new drugs that redirect NK cells and enhance their function. Despite significant progress, myeloma remains incurable and novel treatment approaches are required. Strategies to take advantage of the intrinsic antitumor properties of NK cells to treat MM represent a novel immunotherapeutic approach. One such strategy is adoptive NK cell therapy that consist of infusions of NK cells that have been propagated ex vivo. Adoptive NK cell therapy encompasses contemporary genetic engineering strategies such as chimeric antigen receptor (CAR)-engineered NK cells. An alternative approach involves the use of pharmacologic agents to enhance NK cell activity against myeloma. NK cell-modulating therapies can be used to bolster the function of endogenous NK cells or to enhance the efficacy of adoptively infused NK cells. Here, we review the mechanistic complexities influencing NK cell activity in MM and highlight a variety of innovative NK cell-based strategies being developed for the treatment of MM.

Fostamatinib for the Treatment of Hospitalized Adults With Coronavirus Disease 2019: A Randomized Trial.

BACKGROUND: Coronavirus disease 2019 (COVID-19) requiring hospitalization is characterized by robust antibody production, dysregulated immune response, and immunothrombosis. Fostamatinib is a novel spleen tyrosine kinase inhibitor that we hypothesize will ameliorate Fc activation and attenuate harmful effects of the anti-COVID-19 immune response. METHODS: We conducted a double-blind, randomized, placebo-controlled trial in hospitalized adults requiring oxygen with COVID-19 where patients receiving standard of care were randomized to receive fostamatinib or placebo. The primary outcome was serious adverse events by day 29. RESULTS: A total of 59 patients underwent randomization (30 to fostamatinib and 29 to placebo). Serious adverse events occurred in 10.5% of patients in the fostamatinib group compared with 22% in placebo (P = .2). Three deaths occurred by day 29, all receiving placebo. The mean change in ordinal score at day 15 was greater in the fostamatinib group (-3.6 ±â€…0.3 vs -2.6 ±â€…0.4, P = .035) and the median length in the intensive care unit was 3 days in the fostamatinib group vs 7 days in placebo (P = .07). Differences in clinical improvement were most evident in patients with severe or critical disease (median days on oxygen, 10 vs 28, P = .027). There were trends toward more rapid reductions in C-reactive protein, D-dimer, fibrinogen, and ferritin levels in the fostamatinib group. CONCLUSION: For COVID-19 requiring hospitalization, the addition of fostamatinib to standard of care was safe and patients were observed to have improved clinical outcomes compared with placebo. These results warrant further validation in larger confirmatory trials. CLINICAL TRIALS REGISTRATION: Clinicaltrials.gov, NCT04579393.

LIR-1 educates expanded human NK cells and defines a unique antitumor NK cell subset with potent antibody-dependent cellular cytotoxicity.

OBJECTIVE: KIR and NKG2A receptors educate human NK cells to stay responsive to cells with diminished HLA class I. Here, we addressed whether the HLA class I-binding receptor LIR-1 (LILRB1/ILT2/CD85j), which is widely expressed on human NK cells, can mediate education and contribute to antitumor functions of NK cells. METHODS: Healthy donor NK cells either unstimulated, overnight cytokine-activated or ex vivo-expanded were used to target human cell lines. Phenotype and function were analysed using flow cytometry and 51Cr-release assays. RESULTS: We found that the inhibitory receptor LIR-1 can mediate NK cell education under specific conditions. This novel finding was exclusive to expanded NK cells and further characterisation of the cells revealed high expression of granzyme B and DNAM-1, which both previously have been linked to NK cell education. Corroborating the rheostat education model, LIR-1 co-expression with an educating KIR further increased the responsiveness of expanded NK cells. Inversely, antibody masking of LIR-1 decreased the responsiveness. LIR-1+ expanded NK cells displayed high intrinsic ADCC that, in contrast to KIR and NKG2A, was not inhibited by HLA class I. CONCLUSION: These findings identify a unique NK cell subset attractive for adoptive cell therapy to treat cancer. Given that LIR-1 binds most HLA class I molecules, this subset may be explored in both autologous and allogeneic settings to innately reject HLA class I- tumor cells as well as HLA class I+ target cells when combined with antitumor antibodies. Further studies are warranted to address the potential of this subset in vivo.

Pre-vaccination and early B cell signatures predict antibody response to SARS-CoV-2 mRNA vaccine.

SARS-CoV-2 mRNA vaccines are highly effective, although weak antibody responses are seen in some individuals with correlates of immunity that remain poorly understood. Here we longitudinally dissected antibody, plasmablast, and memory B cell (MBC) responses to the two-dose Moderna mRNA vaccine in SARS-CoV-2-uninfected adults. Robust, coordinated IgA and IgG antibody responses were preceded by bursts of spike-specific plasmablasts after both doses, but earlier and more intensely after dose two. Distinct antigen-specific MBC populations also emerged post-vaccination with varying kinetics. We identified antigen non-specific pre-vaccination MBC and post-vaccination plasmablasts after dose one and their spike-specific counterparts early after dose two that correlated with subsequent antibody levels. These baseline and response signatures can thus provide early indicators of serological efficacy and explain response variability in the population.

Combined haploidentical and cord blood transplantation for refractory severe aplastic anaemia and hypoplastic myelodysplastic syndrome.

Umbilical cord blood (UCB) transplantation is a potentially curative treatment for patients with refractory severe aplastic anaemia (SAA), but has historically been associated with delayed engraftment and high graft failure and mortality rates. We conducted a prospective phase 2 trial to assess outcome of an allogeneic transplant regimen that co-infused a single UCB unit with CD34+ -selected cells from a haploidentical relative. Among 29 SAA patients [including 10 evolved to myelodysplastic syndrome (MDS)] who underwent the haplo cord transplantation (median age 20 years), 97% had neutrophil recovery (median 10 days), and 93% had platelet recovery (median 32 days). Early myeloid engraftment was from the haplo donor and was gradually replaced by durable engraftment from UCB in most patients. The cumulative incidences of grade II-IV acute and chronic graft-versus-host disease (GVHD) were 21% and 41%, respectively. With a median follow-up of 7·5 years, overall survival was 83% and GVHD/relapse-free survival was 69%. Patient- and transplant-related factors had no impact on engraftment and survival although transplants with haplo-versus-cord killer-cell immunoglobulin-like receptor (KIR) ligand incompatibility had delayed cord engraftment. Our study shows haplo cord transplantation is associated with excellent engraftment and long-term outcome, providing an alternative option for patients with refractory SAA and hypoplastic MDS who lack human leucocyte antigen (HLA)-matched donors.

Systematic improvements in lentiviral transduction of primary human natural killer cells undergoing ex vivo expansion.

Transduction of primary human natural killer (NK) cells with lentiviral vectors has historically been challenging. We sought to evaluate multiple parameters to optimize lentiviral transduction of human peripheral blood NK cells being expanded to large numbers using a good manufacturing practice (GMP)-compliant protocol that utilizes irradiated lymphoblastoid (LCL) feeder cells. Although prestimulation of NK cells with interleukin (IL)-2 for 2 or more days facilitated transduction with vesicular stomatitis virus glycoprotein (VSVG)-pseudotyped lentivirus, there was a subsequent impairment in the capacity of transduced NK cells to proliferate when stimulated with LCL feeder cells. In contrast, incubation of human NK cells with LCL feeder cells plus IL-2 before transduction in the presence of the TBK1 inhibitor BX795 resulted in efficient lentiviral integration (mean of 23% transgene+ NK cells) and successful subsequent proliferation of the transduced cells. Investigation of multiple internal promoter sequences within the same lentiviral vector revealed differences in percentage and level of transgene expression per NK cell. Bicistronic lentiviral vectors encoding both GFP and proteins suitable for the isolation of transduced cells with magnetic beads led to efficient transgene expression in NK cells. The optimized approaches described herein provide a template for protocols that generate large numbers of fully functional and highly purified lentivirus-transduced NK cells for clinical trials.