Programmed cell death protein-1 (PD-1) checkpoint immunotherapy efficacy remains unpredictable in glioblastoma (GBM) patients due to the genetic heterogeneity and immunosuppressive tumor ...microenvironments. Here, we report a microfluidics-based, patient-specific 'GBM-on-a-Chip' microphysiological system to dissect the heterogeneity of immunosuppressive tumor microenvironments and optimize anti-PD-1 immunotherapy for different GBM subtypes. Our clinical and experimental analyses demonstrated that molecularly distinct GBM subtypes have distinct epigenetic and immune signatures that may lead to different immunosuppressive mechanisms. The real-time analysis in GBM-on-a-Chip showed that mesenchymal GBM niche attracted low number of allogeneic CD154+CD8+ T-cells but abundant CD163+ tumor-associated macrophages (TAMs), and expressed elevated PD-1/PD-L1 immune checkpoints and TGF-β1, IL-10, and CSF-1 cytokines compared to proneural GBM. To enhance PD-1 inhibitor nivolumab efficacy, we co-administered a CSF-1R inhibitor BLZ945 to ablate CD163+ M2-TAMs and strengthened CD154+CD8+ T-cell functionality and GBM apoptosis on-chip. Our ex vivo patient-specific GBM-on-a-Chip provides an avenue for a personalized screening of immunotherapies for GBM patients.
Screening for therapeutic targets is standard of care in the management of advanced non-small cell lung cancer. However, most molecular assays utilize tumor tissue, which may not always be available. ..."Liquid biopsies" are plasma-based next generation sequencing (NGS) assays that use circulating tumor DNA to identify relevant targets. To compare the sensitivity, specificity, and accuracy of a plasma-based NGS assay to solid-tumor-based NGS we retrospectively analyzed sequencing results of 100 sequential patients with lung adenocarcinoma at our institution who had received concurrent testing with both a solid-tissue-based NGS assay and a commercially available plasma-based NGS assay. Patients represented both new diagnoses (79%) and disease progression on treatment (21%); the majority (83%) had stage IV disease. Tissue-NGS identified 74 clinically relevant mutations, including 52 therapeutic targets, a sensitivity of 94.8%, while plasma-NGS identified 41 clinically relevant mutations, a sensitivity of 52.6% (p < 0.001). Tissue-NGS showed significantly higher sensitivity and accuracy across multiple patient subgroups, both in newly diagnosed and treated patients, as well as in metastatic and nonmetastatic disease. Discrepant cases involved hotspot mutations and actionable fusions including those in EGFR, ALK, and NTRK1. In summary, tissue-NGS detects significantly more clinically relevant alterations and therapeutic targets compared to plasma-NGS, suggesting that tissue-NGS should be the preferred method for molecular testing of lung adenocarcinoma when tissue is available. Plasma-NGS can still play an important role when tissue testing is not possible. However, given its low sensitivity, a negative result should be confirmed with a tissue-based assay.
Isocitrate dehydrogenase (
)-mutant glioma is a distinct glioma molecular subtype for which no effective molecularly directed therapy exists. Low-grade gliomas, which are 80%-90%
-mutant, have high ...RNA levels of the cell surface Notch ligand DLL3. We sought to determine DLL3 expression by IHC in glioma molecular subtypes and the potential efficacy of an anti-DLL3 antibody-drug conjugate (ADC), rovalpituzumab tesirine (Rova-T), in
-mutant glioma.
We evaluated
expression by RNA using TCGA data and by IHC in a discovery set of 63 gliomas and 20 nontumor brain tissues and a validation set of 62 known
wild-type and mutant gliomas using a monoclonal anti-DLL3 antibody. Genotype was determined using a DNA methylation array classifier or by sequencing. The effect of Rova-T on patient-derived endogenous
-mutant glioma tumorspheres was determined by cell viability assay.
Compared to
wild-type glioblastoma,
-mutant gliomas have significantly higher
RNA (
< 1 × 10
) and protein by IHC (
= 0.0014 and
< 4.3 × 10
in the discovery and validation set, respectively). DLL3 immunostaining was intense and homogeneous in
-mutant gliomas, retained in all recurrent tumors, and detected in only 1 of 20 nontumor brains. Patient-derived
-mutant glioma tumorspheres overexpressed DLL3 and were potently sensitive to Rova-T in an antigen-dependent manner.
DLL3 is selectively and homogeneously expressed in
-mutant gliomas and can be targeted with Rova-T in patient-derived
-mutant glioma tumorspheres. Our findings are potentially immediately translatable and have implications for therapeutic strategies that exploit cell surface tumor-associated antigens.
Tenosynovial giant cell tumor (TGCT) is a benign neoplasm characterized by recurrent fusions involving the colony-stimulating factor 1 (
) gene and translocation partners including collagen type VI ...alpha 3 chain (
) or S100 calcium-binding protein A10 (
). Herein, we report three atypical TGCT cases with very unusual morphology comprising areas with increased cellular atypia, mitotic activity, and worrisome features that harbor unique non-
gene fusions. Anchored multiplex PCR (AMP) for next-generation sequencing utilizing a customized panel targeting 86 cancer-related genes was performed, and it identified novel non-
-driven gene fusions:
,
, and
. Screening of three control TGCTs with conventional morphology found translocations involving
, with partner genes
,
, and newly identified
. All novel fusions were further validated by reverse transcriptase-PCR (RT-PCR) and Sanger sequencing. Late and multiple local recurrences occurred in the atypical TGCTs, while no recurrences were reported in the conventional TGCTs. Our findings reveal that atypical TGCTs harbor gene fusions not implicating
and suggest that non-
fusions potentially confer greater propensity to recurrences and local aggressiveness while indicating the presence of alternate pathogenic mechanisms that warrant further investigation.
Soft tissue tumors can be categorized molecularly into two categories: tumors which are known to have recurrent molecular alterations and tumors which do not have consistent recurrent molecular ...alterations or translocations. These “nontranslocation” associated sarcomas are clinically more aggressive than their more stable counterparts. However, recent advances in RNA sequencing have discovered recurrent novel fusions within the latter group, namely TERT‐TRIO fusions. Furthermore, a recent report discovered this fusion in a spindle cell liposarcoma. Our case describes a novel fusion of CTNND2, a neighbor gene of TRIO, and TERT in a spindle cell liposarcoma, and provides further evidence that spindle cell liposarcoma should be a distinct entity from dedifferentiated liposarcoma.
HIV-1 envelope glycoproteins (Env) are the only viral antigens present on the virus surface and serve as the key targets for virus-neutralizing antibodies. However, HIV-1 deploys multiple strategies ...to shield the vulnerable sites on its Env from neutralizing antibodies. The V1V2 domain located at the apex of the HIV-1 Env spike is known to encompass highly variable loops, but V1V2 also contains immunogenic conserved elements recognized by cross-reactive antibodies. This study evaluates human monoclonal antibodies (mAbs) against V2 epitopes which overlap with the conserved integrin α4β7-binding LDV/I motif, designated as the V2i (integrin) epitopes. We postulate that the V2i Abs have weak or no neutralizing activities because the V2i epitopes are often occluded from antibody recognition. To gain insights into the mechanisms of the V2i occlusion, we evaluated three elements at the distal end of the V1V2 domain shown in the structure of V2i epitope complexed with mAb 830A to be important for antibody recognition of the V2i epitope. Amino-acid substitutions at position 179 that restore the LDV/I motif had minimal effects on virus sensitivity to neutralization by most V2i mAbs. However, a charge change at position 153 in the V1 region significantly increased sensitivity of subtype C virus ZM109 to most V2i mAbs. Separately, a disulfide bond introduced to stabilize the hypervariable region of V2 loop also enhanced virus neutralization by some V2i mAbs, but the effects varied depending on the virus. These data demonstrate that multiple elements within the V1V2 domain act independently and in a virus-dependent fashion to govern the antibody recognition and accessibility of V2i epitopes, suggesting the need for multi-pronged strategies to counter the escape and the shielding mechanisms obstructing the V2i Abs from neutralizing HIV-1.
Of 52 patients with recurrent tuberculosis in Shanghai, People's Republic of China, 32 (61.5%) had isolates in which genotype patterns of
Mycobacterium tuberculosis
differed between first and second ...episodes. This result indicates that exogenous reinfection is common in an area with a high incidence of tuberculosis.
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