Immune-checkpoint blockade (ICB) has demonstrated efficacy in many tumor types, but predictors of responsiveness to anti-PD1 ICB are incompletely characterized. In this study, we analyzed a clinically annotated cohort of patients with melanoma (n = 144) treated with anti-PD1 ICB, with whole-exome and whole-transcriptome sequencing of pre-treatment tumors. We found that tumor mutational burden as a predictor of response was confounded by melanoma subtype, whereas multiple novel genomic and transcriptomic features predicted selective response, including features associated with MHC-I and MHC-II antigen presentation. Furthermore, previous anti-CTLA4 ICB exposure was associated with different predictors of response compared to tumors that were naive to ICB, suggesting selective immune effects of previous exposure to anti-CTLA4 ICB. Finally, we developed parsimonious models integrating clinical, genomic and transcriptomic features to predict intrinsic resistance to anti-PD1 ICB in individual tumors, with validation in smaller independent cohorts limited by the availability of comprehensive data. Broadly, we present a framework to discover predictive features and build models of ICB therapeutic response.

Dysregulation of B-catenin is of importance to the development of diverse human malignancies. The MUC1 oncoprotein is aberrantly overexpressed by most human carcinomas and associates with B-catenin. However, the functional significance of the MUC1-B-catenin interaction is not known. Here, we show that MUC1 increases B-catenin levels in the cytoplasm and nucleus of carcinoma cells. Previous studies have shown that glycogen synthase kinase 3B (GSK3B) phosphorylates B-catenin and thereby targets it for proteosomal degradation. Consistent with the up-regulation of B-catenin levels, our results show that MUC1 blocks GSK3B-mediated phosphorylation and degradation of B-catenin. To further define the interaction between MUC1 and B-catenin, we identified a serine-rich motif (SRM) in the MUC1 cytoplasmic domain that binds directly to B-catenin Armadillo repeats. Mutation of the SRM attenuated binding of MUC1 to B-catenin and MUC1-mediated inhibition of B-catenin degradation. Importantly, disruption of the MUC1-B-catenin interaction with the SRM mutant also attenuated MUC1-induced anchorage-dependent and -independent growth and delayed MUC1-mediated tumorigenicity. These findings indicate that MUC1 promotes transformation, at least in part, by blocking GSK3B-mediated phosphorylation and thereby degradation of B-catenin. (Cancer Res 2005; 65(22): 10413-22)

The human DF3/MUC1 mucin-like glycoprotein is aberrantly overexpressed by most carcinomas of the breast and other epithelia. The contribution of MUC1 overexpression to the malignant phenotype is, however, not known. In the present studies, we have stably expressed MUC1 in rat 3Y1 fibroblasts. MUC1-positive cells were selected from independent transfections. The results demonstrate that, as found in human carcinomas, MUC1 is expressed on the cell surface and as a complex with b-catenin in the nucleus of the transfectants. Colony formation in soft agar demonstrates that cells expressing MUC1, but not the empty vector, exhibit anchorageindependent growth. The results also show that MUC1 expression confers tumor formation in nude mice. These findings provide the first evidence that MUC1 induces cellular transformation.

Objective: To determine the intra-, inter-and test-retest variability of CT-based texture analysis (CTTA) metrics. Materials and methods: In this study, we conducted a series of CT imaging experiments using a texture phantom to evaluate the performance of a CTTA panel on routine abdominal imaging protocols. The phantom comprises of three different regions with various textures found in tumors. The phantom was scanned on two CT scanners viz. the Philips Brilliance 64 CT and Toshiba Aquilion Prime 160 CT scanners. The intra-scanner variability of the CTTA metrics was evaluated across imaging parameters such as slice thickness, field of view, post-reconstruction filtering, tube voltage, and tube current. For each scanner and scanning parameter combination, we evaluated the performance of eight different types of texture quantification techniques on a predetermined region of interest (ROI) within the phantom image using 235 different texture metrics. We conducted the repeatability (test-retest) and robustness (intra-scanner) test on both the scanners and the reproducibility test was conducted by comparing the inter-scanner differences in the repeatability and robustness to identify reliable CTTA metrics. Reliable metrics are those metrics that are repeatable, reproducible and robust. Results: As expected, the robustness, repeatability and reproducibility of CTTA metrics are variably sensitive to various scanner and scanning parameters. Entropy of Fast Fourier Transform-based texture metrics was overall most reliable across the two scanners and scanning conditions. Post-processing techniques that reduce image noise while preserving the underlying edges associated with true anatomy or pathology bring about significant differences in radiomic reliability compared to when they were not used. Conclusion: Following large-scale validation, identification of reliable CTTA metrics can aid in conducting large-scale multicenter CTTA analysis using sample sets acquired using different imaging protocols, scanners etc.

Low-energy x-rays are known to have a higher relative biological effectiveness (RBE) than higher energy photons such as the gamma rays from 192Ir and 60Co. In this work the initial yield of single- and double-strand DNA breaks (SSB and DSB) and the RBE was estimated for a novel electronic brachytherapy source (EBS), emitting 40-50 kVp photons. An EGSnrc Monte Carlo model of the source was used in combination with the 'Monte Carlo damage simulation' program (Semenenko and Stewart 2004 Radiat. Res. 161 451-57; 2006 Phys. Med. Biol. 51 1693-706). The results indicate a substantially reduced SSB yield and increased DSB yield for the EBS compared to 60Co or 192Ir, leading to an enhanced RBE by 40-50%. The RBE estimate for the low-energy x-ray EBS was found to be very similar to the low-energy gamma ray brachytherapy isotope 125I. Biological damage was estimated in several human tissues: muscle, breast, calcified breast and cortical bone. SSB and DSB yields were similar in all media, except in bone. These findings should be taken into account if the EBS is intended to replace brachytherapy with the commonly used 192Ir isotope.

T-cell immunoglobulin mucin-3 (Tim-3) is expressed on pathogenic T cells, and its ligand galectin-9 (gal-9) is up-regulated in inflamed tissues. When Tim-3 ؉ T cells encounter high gal-9 levels, they are deleted. Tim-3 is up-regulated on activated T cells during GVHD. Inhibition of Tim-3/ gal-9 binding by infusion of a Tim-3-Ig fusion protein or Tim-3 ؊/؊ donor T cells increased T-cell proliferation and GVHD lethality. When the Tim-3/gal-9 pathway engagement was augmented using gal-9 transgenic recipients, GVHD lethality was slowed. Together, these data indicate a potential for modulating this pathway to reduce disease by increasing Tim-3 or gal-9 engagement. Paradoxically, when Tim-3/gal-9 was inhibited in the absence of donor T-regulatory cells (Tregs), GVHD was inhibited. GVHD reduction was associated with decreased colonic inflammatory cytokines as well as epithelial barrier destruction. CD25-depleted Tim-3 ؊/؊ donor T cells underwent increased activationinduced cell death because of increased IFN-␥ production. To our knowledge, these studies are the first to show that although the absence of Tim-3/gal-9 pathway interactions augments systemic GVHD, concurrent donor Treg depletion paradoxically and surprisingly inhibits GVHD. Thus, although donor Tregs typically inhibit GVHD, under some conditions, such Tregs actually may contribute to GVHD by reducing activation-induced T-cell death. (Blood. 2012;120(3):682-690) IntroductionGVHD remains the leading cause of morbidity and mortality after bone marrow transplantation (BMT). Patients are given immune suppressive therapy to prevent or diminish the severity of GVHD after allogeneic BMT that in turn increases the risk of infection and disease recurrence. Novel GVHD strategies remain a high priority.The T-cell immunoglobulin mucin (TIM) family consists of 3 proteins (TIM-1, -3, and -4), homologous in mouse and human. 1 Tim-3 was the first described member 2 and has been the most well studied. Differentiated T-effector cells (Teffs) express Tim-3 with the highest density on T-helper (Th)1, lower density on Th17, and no expression on Th2 cells. 3,4 The expression of galectin-9 (gal-9), identified as a ligand for Tim-3, is up-regulated in inflamed tissues. [5][6][7][8] When Tim-3 ϩ Teffs encounter high gal-9 levels, they are deleted. 5,9-11 A major function of the Tim-3/gal-9 pathway is to limit immune responses under conditions of tissue inflammation and injury. In vivo blockade of Tim-3/gal-9 interaction or the use of Tim-3 knockout (Ϫ/Ϫ) mice increases Th1 cells within inflamed tissues. 2,12,13 When Tim-3 binds with gal-9, Th1 responses are inhibited and peripheral tolerance is induced. 5,12,13 In vivo blocking strategies relying on monoclonal anti-Tim-3 antibody and Tim-3-Ig fusion protein showed exacerbation of experimental autoimmune encephalomyelitis and autoimmune diabetes. 2,12 Transplant tolerance induced by donor-specific transfusion and anti-CD154 treatment was impaired. 13 Thus, Tim-3/gal-9 signaling acts to dampen a Th1 immune response, whereas signaling ...

The Axxent developed by Xoft Inc. is a novel electronic brachytherapy system capable of generating x-rays up to 50 keV. These low energy photon-emitting sources merit attention not only because of their ability to vary the dosimetric properties of the radiation, but also because of the radiobiological effects of low energy x-rays. The objective of this study is to characterize the x-ray source and to model it using the Geant4 Monte Carlo code. Spectral and attenuation curve measurements are performed at various peak voltages and angles and the source is characterized in terms of spectrum and half-value layers (HVLs). Also, the effects of source variation and source aging are quantified. Bremsstrahlung splitting, phase-space scoring and particle-tagging features are implemented in the Geant4 code, which is bench-marked against BEAMnrc simulations. HVLs from spectral measurements, attenuation curve measurements and Geant4 simulations mostly agree within uncertainty. However, there are discrepancies between measurements and simulations for photons emitted on the source transverse plane (90 degrees).

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