Adoptive T cell therapy (ACT) using ex vivo-expanded autologous tumor-infiltrating lymphocytes (TILs) can mediate complete regression of certain human cancers. The impact of TIL phenotypes on ...clinical success of TIL-ACT is currently unclear. Using high-dimensional analysis of human ACT products, we identified a memory-progenitor CD39-negative stem-like phenotype (CD39
CD69
) associated with complete cancer regression and TIL persistence and a terminally differentiated CD39-positive state (CD39
CD69
) associated with poor TIL persistence. Most antitumor neoantigen-reactive TILs were found in the differentiated CD39
state. However, ACT responders retained a pool of CD39
stem-like neoantigen-specific TILs that was lacking in ACT nonresponders. Tumor-reactive stem-like TILs were capable of self-renewal, expansion, persistence, and superior antitumor response in vivo. These data suggest that TIL subsets mediating ACT response are distinct from TIL subsets enriched for antitumor reactivity.
The accurate identification of antitumor T cell receptors (TCRs) represents a major challenge for the engineering of cell-based cancer immunotherapies. By mapping 55 neoantigen-specific TCR ...clonotypes (NeoTCRs) from 10 metastatic human tumors to their single-cell transcriptomes, we identified signatures of CD8
and CD4
neoantigen-reactive tumor-infiltrating lymphocytes (TILs). Neoantigen-specific TILs exhibited tumor-specific expansion with dysfunctional phenotypes, distinct from blood-emigrant bystanders and regulatory TILs. Prospective prediction and testing of 73 NeoTCR signature-derived clonotypes demonstrated that half of the tested TCRs recognized tumor antigens or autologous tumors. NeoTCR signatures identified TCRs that target driver neoantigens and nonmutated viral or tumor-associated antigens, suggesting a common metastatic TIL exhaustion program. NeoTCR signatures delineate the landscape of TILs across metastatic tumors, enabling successful TCR prediction based purely on TIL transcriptomic states for use in cancer immunotherapy.
A common theme across multiple successful immunotherapies for cancer is the recognition of tumor-specific mutations (neoantigens) by T cells. The rapid discovery of such antigen responses could lead ...to improved therapies through the adoptive transfer of T cells engineered to express neoantigen-reactive T cell receptors (TCRs). Here, through CITE-seq (cellular indexing of transcriptomes and epitopes by sequencing) and TCR-seq of non-small cell lung cancer (NSCLC) tumor-infiltrating lymphocytes (TILs), we develop a neoantigen-reactive T cell signature based on clonotype frequency and CD39 protein and CXCL13 mRNA expression. Screening of TCRs selected by the signature allows us to identify neoantigen-reactive TCRs with a success rate of 45% for CD8+ and 66% for CD4+ T cells. Because of the small number of samples analyzed (4 patients), generalizability remains to be tested. However, this approach can enable the quick identification of neoantigen-reactive TCRs and expedite the engineering of personalized neoantigen-reactive T cells for therapy.
Display omitted
•T cells in fresh NSCLC tumor were analyzed by CITE-seq with TCR-seq•Neoantigen-reactive T cells are CD39 protein+, CXCL13+, and high-frequency clonotype•The signature can identify both CD4 and CD8 neoantigen-reactive T cells•This method can expedite the personalized neoantigen-reactive T cell therapy
The discovery of T cell receptors (TCRs) that recognize tumor-specific neoantigens can lead to improved therapies through the adoptive transfer of T cells engineered to express these TCRs. Here, Hanada et al. design a signature of neoantigen-reactive T cells that enables the rapid identification of both CD4 and CD8 neoantigen-reactive TCRs.
Abstract
While traditional techniques have long allowed forensic investigators to positively identify fingermarks on documents of interest, understanding the chronological sequence of events that led ...to their deposition is still seen as a ‘holy grail’ for forensic examinations. By way of example, the question of whether a mark is above or below printed text is crucial. The work herein reveals that a novel application of a recently established fingermark development technique readily allows such differentiation. The process in question allies forensic gelatin lifters with RECOVER, a development system that hinges on the polymerisation of disulfur dinitride. While the latter was specifically developed in its current form for the retrieval of prints from metal surfaces exposed to extreme conditions or washing, its ability to target surface effects allows for visualisation of surface interactions on forensic gelatin lifts. Crucially, in doing so the order in which the lifted material was originally deposited is also revealed. This, therefore, permits clear elucidation of the order of deposition of printed text and fingermarks—and does so both rapidly and in a non-invasive way. This long sought-after capability has the potential to revolutionise forensic document examinations.
Rationale
Small amounts of biofluid samples are frequently found at crime scenes; however, existing gold standard methods such as LC–MS frequently require destructive extraction of the sample before ...a time‐consuming analysis which puts strain on forensic analysis providers and can preclude further sample analysis. This study presents the application of sheath‐flow probe electrospray ionization‐mass spectrometry (sfPESI–MS) to the direct analysis of drug metabolites in dried blood spots (DBS) as a high throughput, minimally destructive alternative.
Methods
A rapid direct analysis method using a sfPESI ionisation source coupled to an Orbitrap Exactive mass spectrometer was applied to detect cocaine metabolites (benzoylecgonine, BZE, cocaethylene, CE, and ecgonine methyl ester, EME) from DBS. An optimisation study exploring the use of different chemical modifiers (formic acid and sodium acetate) in the sfPESI probe extraction solvent was conducted to enhance the sensitivity and reproducibility of the sfPESI–MS method.
Results
Optimisation of the extraction solvent significantly enhanced the sensitivity and reproducibility of the sfPESI–MS method. A quantitative response over a five‐point calibration range 0.5 to 10 μg/ml was obtained for BZE (R2 = 0.9979) and CE (R2 = 0.9948). Limits of detection (LOD) of 1.31, 0.29 and 0.15 μg/ml were achieved for EME, BZE and CE, respectively, from 48 h aged DBSs with % RSD (relative standard deviation) across the calibration range ranging between 19%–28% for BZE + H+, 13%–21% for CE + H+ and 12%–29% for EME + H+.
Conclusions
A rapid (< 20 s) quantitative method for the direct analysis of cocaine metabolites from DBS which requires no prior sample preparation was developed. Although the LOD achieved for BZE (LOD: 0.29 μg/ml) was above the UK threshold limit of exposure for drug driving (0.05 μg/ml), the method may be suitable for use in identifying overdose in forensic analysis.
Using revised Atlanta classification defined outcomes, we compare absolute values in C-reactive protein (CRP), with interval changes in CRP, for severity stratification in acute pancreatitis (AP).
A ...retrospective study of all first incidence AP was conducted over a 5-year period. Interval change in CRP values from admission to day 1, 2 and 3 was compared against the absolute values. Receiver-operator characteristic (ROC) curve and likelihood ratios (LRs) were used to compare ability to predict severe and mild disease.
337 cases of first incidence AP were included in our analysis. ROC curve analysis demonstrated the second day as the most useful time for repeat CRP measurement. A CRP interval change >90 mg/dL at 48 h (+LR 2.15, −LR 0.26) was equivalent to an absolute value of >150 mg/dL within 48 h (+LR 2.32, −LR 0.25). The optimal cut-off for absolute CRP based on new, more stringent definition of severity was >190 mg/dL (+LR 2.72, −LR 0.24).
Interval change in CRP is a comparable measure to absolute CRP in the prognostication of AP severity. This study suggests a rise of >90 mg/dL from admission or an absolute value of >190 mg/dL at 48 h predicts severe disease with the greatest accuracy.
The identification and recovery of suspected human biofluid evidence can present a bottleneck in the crime scene investigation workflow. Crime Scene Investigators typically deploy one of a number of ...presumptive enhancement reagents, depending on what they perceive an analyte to be; the selection of this reagent is largely based on the context of suspected evidence and their professional experience. Positively identified samples are then recovered to a forensic laboratory where confirmatory testing is carried out by large lab-based instruments, such as through mass-spectrometry-based techniques. This work proposes a proof-of-concept study into the use of a small, robust and portable ion mobility spectrometry device that can analyse samples in situ, detecting, identifying and discriminating commonly encountered body fluids from interferences. This analysis exploits the detection and identification of characteristic volatile organic compounds generated by gentle heating, at ambient temperature and pressure, and categorises samples using machine learning, providing investigators with instant identification. The device is shown to be capable of producing characteristic mobility spectra using a dual micro disc pump configuration which separates blood and urine from three visually similar interferences using an unsupervised PCA model with no misclassified samples. The device has the potential to reduce the need for potentially contaminating and destructive presumptive tests, and address the bottleneck created by the time-consuming and laborious detection, recovery and analysis workflow currently employed.
Abstract Background The Prostate, Lung, Colorectal and Ovarian (PLCO) Cancer Screening Trial originally reported no mortality benefit of ovarian cancer screening after a median of 12.4 years of ...follow-up. The UKCTOCS screening trial failed to show a statistically significant mortality reduction in the primary analysis but reported an apparent increased mortality benefit in trial years 7–14 compared to 0–7. Here we report an updated analysis of PLCO with extended mortality follow-up. Methods Participants were randomized from 1993 to 2001 at ten U.S. centers to an intervention or usual care arm. Intervention arm women were screened for ovarian cancer with annual trans-vaginal ultrasound (TVU) (4 years) and CA-125 (6 years), with a fixed cutoff at 35 U/mL for CA-125. The original follow-up period was for up to 13 years (median follow-up 12.4 years); in this analysis follow-up for mortality was extended by up to 6 years. Results 39,105 (intervention) and 39,111 (usual care) women were randomized, of which 34,253 and 34,304, respectively, had at least one ovary at baseline. Median follow-up was 14.7 years in each arm and maximum follow-up 19.2 years in each arm. A total of 187 (intervention) and 176 (usual care) deaths from ovarian cancer were observed, for a risk-ratio of 1.06 (95% CI: 0.87–1.30). Risk-ratios were similar for study years 0–7 (RR = 1.04), 7–14 (RR = 1.06) and 14 + (RR = 1.09). The risk ratio for all-cause mortality was 1.01 (95% CI: 0.97–1.05). Ovarian cancer specific survival was not significantly different across trial arms (p = 0.16). Conclusion Extended follow-up of PLCO indicated no mortality benefit from screening for ovarian cancer with CA-125 and TVU.
PDF
