Allogeneic natural killer cell therapy

Interest in adoptive cell therapy for treating cancer is exploding owing to early clinical successes of autologous chimeric antigen receptor (CAR) T lymphocyte therapy. However, limitations using T cells and autologous cell products are apparent as they (1) take weeks to generate, (2) utilize a 1:1 donor-to-patient model, (3) are expensive, and (4) are prone to heterogeneity and manufacturing failures. CAR T cells are also associated with significant toxicities, including cytokine release syndrome, immune effector cell-associated neurotoxicity syndrome, and prolonged cytopenias. To overcome these issues, natural killer (NK) cells are being explored as an alternative cell source for allogeneic cell therapies. NK cells have an inherent ability to recognize cancers, mediate immune functions of killing and communication, and do not induce graft-versus-host disease, cytokine release syndrome, or immune effector cell-associated neurotoxicity syndrome. NK cells can be obtained from blood or cord blood or be derived from hematopoietic stem and progenitor cells or induced pluripotent stem cells, and can be expanded and cryopreserved for off-the-shelf availability. The first wave of point-of-care NK cell therapies led to the current allogeneic NK cell products being investigated in clinical trials with promising preliminary results. Basic advances in NK cell biology and cellular engineering have led to new translational strategies to block inhibition, enhance and broaden target cell recognition, optimize functional persistence, and provide stealth from patients\u27 immunity. This review details NK cell biology, as well as NK cell product manufacturing, engineering, and combination therapies explored in the clinic leading to the next generation of potent, off-the-shelf cellular therapies for blood cancers

Bridging therapy with axicabtagene ciloleucel for large B-cell lymphoma: Results from the US Lymphoma CAR-T Consortium

During the manufacturing period of autologous chimeric antigen receptor (CAR) T-cell therapy, patients may experience a decline in their condition due to cancer progression. In this study, we investigated the impact of bridging therapy (BT) on the outcome of patients with relapsed/refractory large B-cell lymphoma who received antilymphoma treatment between leukapheresis and axicabtagene ciloleucel (axi-cel) infusion. We conducted our analysis using data from the multicenter US Lymphoma CAR-T Consortium, with a median follow-up of 33 months (range, 4.3-42.1). Out of the 298 patients who underwent leukapheresis, 275 patients received axi-cel. A total 52% of patients (n = 143) who received BT had a higher baseline risk profile than patients who did not receive BT, and these patients, as a group, had inferior outcomes compared with those who did not receive BT. However, after propensity score matching between the 2 groups, there were no statistically significant differences in overall response rate (77% vs 87%; P = .13), complete response rate (58% vs 70%; P = .1), progression-free survival (hazard ratio [HR], 1.25; P = .23), and overall survival (HR, 1.39; P=.09) between the BT group and the no-BT group, respectively. Analyzing the effects of BT in the whole cohort that underwent leukapheresis regardless of receiving axi-cel (intention-to-treat analysis) showed similar results. Radiation BT resulted in outcomes similar to those observed with nonradiation BT. Our findings suggest that BT may be safe without a significant impact on long-term survival for patients who require disease stabilization during the manufacturing period. Moreover, our results suggest that there is no clear advantage to using radiation-based BT over nonradiation-based BT

CD7-deleted hematopoietic stem cells can restore immunity after CAR T cell therapy

Targeting T cell malignancies with universal CD7-targeting chimeric antigen receptor T cells (UCART7) can lead to profound immune deficiency due to loss of normal T and NK cells. While a small population of endogenous CD7- T cells exists, these cells are unlikely to be able to repopulate the entire immune repertoire after UCART7 treatment, as they are limited in number and proliferative capacity. To rescue T and NK cells after UCART7, we created hematopoietic stem cells genetically deleted for CD7 (CD7-KO HSCs). CD7-KO HSCs were able to engraft immunodeficient mice and differentiate into T and NK cells lacking CD7 expression. CD7-KO T and NK cells could perform effector functions as robustly as control T and NK cells. Furthermore, CD7-KO T cells were phenotypically and functionally distinct from endogenous CD7- T cells, indicating that CD7-KO T cells can supplement immune functions lacking in CD7- T cells. Mice engrafted with CD7-KO HSCs maintained T and NK cell numbers after UCART7 treatment, while these were significantly decreased in control mice. These studies support the development of CD7-KO HSCs to augment host immunity in patients with T cell malignancies after UCART7 treatment

Holocene warming of alpine rockwalls decreased rockwall erosion rates

Alpine rockwall erosion studies suggest that deglaciated rockwalls in the European Alps are eroding slower today than at earlier times in the Holocene. Explanations for this have included a waned glacial debuttressing effect since the retreat of Last Glacial ice loads and the establishment of more moderate climates, but seldom have such explanations been robustly tested. We combine field data with modelling to reconstruct changes in rockwall erosion and associated climate drivers, in an alpine valley of the European Alps since the beginning of the Holocene. Paleo (Holocene to decadal-scale) erosion rates were calculated from talus accumulation below rockwalls, and compared with recent rates (2016–2019) measured from repeat laserscan surveys of the same rockwalls. We reconstructed the glacial retreat history in the valley, and modelled Holocene changes in permafrost distribution and frost cracking using calibrated rockwall temperature reconstructions. We found that rockwalls that have been free of glacier ice since ∼10 ka experienced higher Holocene-averaged erosion rates compared to recent erosion rates. Our modelling suggests this relates to periods of higher intensities of frost cracking and cycles of permafrost aggradation and degradation in the Holocene, relative to today. For a recently deglaciated high-elevation rockwall, erosion rates were 1–2 orders of magnitude greater than the lower-elevation sites, but decayed rapidly over time since deglaciation. A high, but rapidly decaying rate results from short-lived paraglacial adjustment, permafrost thaw and high frost cracking activity. Our findings suggest that periglacial activity strongly influences the rates and patterns of erosion of deglaciated alpine rockwalls. This helps to explain why paleo Holocene deglaciated rockwall erosion rates tend to exceed recent rates in the European Alps

Induced CD8α identifies human NK cells with enhanced proliferative fitness and modulates NK cell activation

The surface receptor CD8α is present on 20%-80% of human (but not mouse) NK cells, yet its function on NK cells remains poorly understood. CD8α expression on donor NK cells was associated with a lack of therapeutic responses in patients with leukemia in prior studies, thus, we hypothesized that CD8α may affect critical NK cell functions. Here, we discovered that CD8α- NK cells had improved control of leukemia in xenograft models compared with CD8α+ NK cells, likely due to an enhanced capacity for proliferation. Unexpectedly, we found that CD8α expression was induced on approximately 30% of previously CD8α- NK cells following IL-15 stimulation. These induced CD8α+ (iCD8α+) NK cells had the greatest proliferation, responses to IL-15 signaling, and metabolic activity compared with those that sustained existing CD8α expression (sustained CD8α+) or those that remained CD8α- (persistent CD8α-). These iCD8α+ cells originated from an IL-15Rβhi NK cell population, with CD8α expression dependent on the transcription factor RUNX3. Moreover, CD8A CRISPR/Cas9 deletion resulted in enhanced responses through the activating receptor NKp30, possibly by modulating KIR inhibitory function. Thus, CD8α status identified human NK cell capacity for IL-15-induced proliferation and metabolism in a time-dependent fashion, and its presence had a suppressive effect on NK cell-activating receptors

Memory-like differentiation enhances NK cell responses against colorectal cancer

Metastatic (m) colorectal cancer (CRC) is an incurable disease with a poor prognosis and thus remains an unmet clinical need. Immune checkpoint blockade (ICB)-based immunotherapy is effective for mismatch repair-deficient (dMMR)/microsatellite instability-high (MSI-H) mCRC patients, but it does not benefit the majority of mCRC patients. NK cells are innate lymphoid cells with potent effector responses against a variety of tumor cells but are frequently dysfunctional in cancer patients. Memory-like (ML) NK cells differentiated after IL-12/IL-15/IL-18 activation overcome many challenges to effective NK cell anti-tumor responses, exhibiting enhanced recognition, function, and in vivo persistence. We hypothesized that ML differentiation enhances the NK cell responses to CRC. Compared to conventional (c) NK cells, ML NK cells displayed increased IFN-γ production against both CRC cell lines and primary patient-derived CRC spheroids. ML NK cells also exhibited improved killing of CRC target cells in vitro in short-term and sustained cytotoxicity assays, as well as in vivo in NSG mice. Mechanistically, enhanced ML NK cell responses were dependent on the activating receptor NKG2D as its blockade significantly decreased ML NK cell functions. Compared to cNK cells, ML NK cells exhibited greater antibody-dependent cytotoxicity when targeted against CRC by cetuximab. ML NK cells from healthy donors and mCRC patients exhibited increased anti-CRC responses. Collectively, our findings demonstrate that ML NK cells exhibit enhanced responses against CRC targets, warranting further investigation in clinical trials for mCRC patients, including those who have failed ICB

Guillain-Barré syndrome and adjuvanted pandemic influenza A (H1N1) 2009 vaccines: A multinational self-controlled case series in Europe

Background: The risk of Guillain-Barré syndrome (GBS) following the United States' 1976 swine flu vaccination campaign in the USA led to enhanced active surveillance during the pandemic influenza (A(H1N1)pdm09) immunization campaign. This study aimed to estimate the risk

T-BET and EOMES sustain mature human NK cell identity and antitumor function

Since the T-box transcription factors (TFs) T-BET and EOMES are necessary for initiation of NK cell development, their ongoing requirement for mature NK cell homeostasis, function, and molecular programming remains unclear. To address this, T-BET and EOMES were deleted in unexpanded primary human NK cells using CRISPR/Cas9. Deleting these TFs compromised in vivo antitumor response of human NK cells. Mechanistically, T-BET and EOMES were required for normal NK cell proliferation and persistence in vivo. NK cells lacking T-BET and EOMES also exhibited defective responses to cytokine stimulation. Single-cell RNA-Seq revealed a specific T-box transcriptional program in human NK cells, which was rapidly lost following T-BET and EOMES deletion. Further, T-BET- and EOMES-deleted CD56bright NK cells acquired an innate lymphoid cell precursor-like (ILCP-like) profile with increased expression of the ILC-3-associated TFs RORC and AHR, revealing a role for T-box TFs in maintaining mature NK cell phenotypes and an unexpected role of suppressing alternative ILC lineages. Our study reveals the critical importance of sustained EOMES and T-BET expression to orchestrate mature NK cell function and identity

t4 Workshop Report: Integrated Testing Strategies (ITS) for Safety Assessment

Integrated testing strategies (ITS), as opposed to single definitive tests or fixed batteries of tests, are expected to efficiently combine different information sources in a quantifiable fashion to satisfy an information need, in this case for regulatory safety assessments. With increasing awareness of the limitations of each individual tool and the development of highly targeted tests and predictions, the need for combining pieces of evidence increases. The discussions that took place during this workshop, which brought together a group of experts coming from different related areas, illustrate the current state of the art of ITS, as well as promising developments and identifiable challenges. The case of skin sensitization was taken as an example to understand how possible ITS can be constructed, optimized and validated. This will require embracing and developing new concepts such as adverse outcome pathways (AOP), advanced statistical learning algorithms and machine learning, mechanistic validation and “Good ITS Practices”.JRC.I.5-Systems Toxicolog

Human epididymis protein 4 reference limits and natural variation in a Nordic reference population

The objectives of this study are to establish reference limits for human epididymis protein 4, HE4, and investigate factors influencing HE4 levels in healthy subjects. HE4 was measured in 1,591 samples from the Nordic Reference Interval Project Bio-bank and Database biobank, using the manual HE4 EIA (Fujirebio) for 802 samples and the Architect HE4 (Abbott) for 792 samples. Reference limits were calculated using the statistical software R. The influence of donor characteristics such as age, sex, body mass index, smoking habits, and creatinine on HE4 levels was investigated using a multivariate model. The study showed that age is the main determinant of HE4 in healthy subjects, corresponding to 2% higher HE4 levels at 30 years (compared to 20 years), 9% at 40 years, 20% at 50 years, 37% at 60 years, 63% at 70 years, and 101% at 80 years. HE4 levels are 29% higher in smokers than in nonsmokers. In conclusion, HE4 levels in healthy subjects are associated with age and smoking status. Age-dependent reference limits are suggested