Achieving the full potential of zinc-finger nucleases (ZFNs) for genome engineering in human cells requires their efficient delivery to the relevant cell types. Here we exploited the infectivity of ...integrase-defective lentiviral vectors (IDLV) to express ZFNs and provide the template DNA for gene correction in different cell types. IDLV-mediated delivery supported high rates (13-39%) of editing at the IL-2 receptor common gamma-chain gene (IL2RG) across different cell types. IDLVs also mediated site-specific gene addition by a process that required ZFN cleavage and homologous template DNA, thus establishing a platform that can target the insertion of transgenes into a predetermined genomic site. Using IDLV delivery and ZFNs targeting distinct loci, we observed high levels of gene addition (up to 50%) in a panel of human cell lines, as well as human embryonic stem cells (5%), allowing rapid, selection-free isolation of clonogenic cells with the desired genetic modification.
Background
Genomic fusions of the anaplastic lymphoma kinase gene (ALK) are a well‐established therapy target in non‐small cell lung cancer (NSCLC). From a survey of 114,200 clinical cases, we ...determined the prevalence of ALK rearrangements (rALK) in non‐NSCLC tumors and report their responsiveness to therapies targeting ALK.
Materials and Methods
Comprehensive genomic profiling of 114,200 relapsed and metastatic malignancies, including both solid tumors and hematolymphoid cancers, was performed using a hybrid‐capture, adaptor ligation‐based next‐generation sequencing assay.
Results
Of 114,200 clinical samples, 21,522 (18.8%) were NSCLC and 92,678 (81.2%) were other tumor types. Of the 876 (0.8%) cases with ALK fusions (fALK) or rALK, 675 (77.1%) were NSCLC and 201 (22.9%) were other tumor types. ALK fusions were significantly more frequent in NSCLC (3.1%) than non‐NSCLC (0.2%; p < .0001). Patients with non‐NSCLC tumors harboring fALK were significantly younger (p < .0001) and more often female (p < .0001) than patients with fALK‐positive NSCLC. EML4 was more often the fusion partner in NSCLC (83.5%) versus non‐NSCLC tumors (30.9%; p < .0001).
Conclusion
ALK rearrangements can be identified in a wide variety of epithelial and mesenchymal malignancies beyond NSCLC. Anti‐ALK therapies can be effective in non‐NSCLC tumors driven by fALK, and further study of therapies targeting ALK in clinical trials involving a wider variety of cancer types appears warranted.
Implications for Practice
Rearrangements involving the ALK gene have been detected in dozens of cancer types using next‐generation sequencing. Patients whose tumors harbor ALK rearrangements or fusions respond to treatment with crizotinib and alectinib, including tumors not normally associated with ALK mutations, such as non‐Langerhans cell histiocytosis or renal cell carcinoma. Comprehensive genomic profiling using next‐generation sequencing can detect targetable ALK fusions irrespective of tumor type or fusions partner.
摘要
背景.间变性淋巴瘤激酶基因（ALK）融合是非小细胞肺癌（NSCLC）治疗的确定靶点。根据对114 200例临床病例展开的调查, 我们确定了非NSCLC肿瘤患者中ALK重排（rALK）的发生率, 并报告了他们对靶向ALK的疗法的反应。
材料与方法.通过基于杂交捕获和适配器连接的下一代测序法对114 200例复发及转移性恶性肿瘤患者（包括实体瘤和淋巴造血系统癌）进行了全面的基因组测序。
结果.在这114 200例临床病例中, 21 522例（18.8%）罹患NSCLC, 92 678例（81.2%）罹患其他类型肿瘤。876例（0.8%）患者出现ALK融合（fALK）或rALK, 其中675例（77.1%）罹患NSCLC, 201例（22.9%）罹患其他类型肿瘤。NSCLC患者（3.1%）中ALK融合的发生率明显高于非NSCLC患者（0.2%）（p < 0.0001）。与fALK‐阳性 NSCLC患者相比, 负荷fALK的非NSCLC患者的年龄明显更小（p < 0.0001）, 并且女性患者的比例通常更高（p < 0.0001）。NSCLC患者（83.5%）中出现融合伴侣EML4的比例通常高于非NSCLC患者（30.9%; p < 0.0001）。
结论.除NSCLC外, 多种上皮和间质恶性肿瘤中均存在ALK重排。抗ALK疗法可有效治疗fALK驱动的非NSCLC肿瘤, 因此, 有必要在涉及各类癌症的临床试验中对靶向ALK的疗法进行进一步研究。
对临床实践的启示：下一代测序技术已被使用对多种癌症类型进行了ALK基因重排检测。负荷ALK重排或融合的肿瘤患者对克唑替尼和alectinib治疗产生反应, 其中包括通常无ALK突变的肿瘤, 例如, 非朗格罕组织细胞增多症或肾细胞癌。使用下一代测序技术进行的全面基因组测序可检测靶向ALK融合, 而这与肿瘤类型或融合伴侣无关。
This article reports a series of 114,200 consecutive clinical cases with comprehensive genomic profiles available to identify ALK fusions that could facilitate the use of precision medicine in both non‐small cell lung cancer (NSCLC) and non‐NSCLC malignancies.
Gastrointestinal stromal tumors (GIST) are rare neoplasms arising from the interstitial cell of Cajal in the gastrointestinal tract. Two thirds of GIST in adult patients have
c-Kit
mutation and ...smaller fractions have platelet derived growth factor receptor alpha (
PDGFRA
) mutation. Surgery is the only curative treatment for localized disease. Imatinib improves survival when used adjuvantly and in advanced disease. Several targeted therapies have also improved survival in GIST patients after progression on imatinib including sunitinib and regorafenib. Recently, United States Federal and Drug Administration (FDA) approved two new tyrosine kinase inhibitors for the treatment of heavily pretreated advanced/unresectable GIST including avapritinib (a selective inhibitor for PDGFRA exon 18 mutation including D842V mutations) and ripretinib (a broad-spectrum kinase inhibitor of c-Kit and PDGFRA). In this article, we will provide a comprehensive review of GIST including the current standard of care treatment and exploring future paradigm shifts in therapy.
The growth of high-quality single crystals of graphene by chemical vapor deposition on copper (Cu) has not always achieved control over domain size and morphology, and the results vary from lab to ...lab under presumably similar growth conditions. We discovered that oxygen (O) on the Cu surface substantially decreased the graphene nucleation density by passivating Cu surface active sites. Control of surface O enabled repeatable growth of centimeter-scale single-crystal graphene domains. Oxygen also accelerated graphene domain growth and shifted the growth kinetics from edge-attachment-limited to diffusion-limited. Correspondingly, the compact graphene domain shapes became dendritic. The electrical quality of the graphene films was equivalent to that of mechanically exfoliated graphene, in spite of being grown in the presence of O.
A measurement of the mass of the Higgs boson in the diphoton decay channel is presented. This analysis is based on 35.9fb−1 of proton-proton collision data collected during the 2016 LHC running ...period, with the CMS detector at a centre-of-mass energy of 13 TeV. A refined detector calibration and new analysis techniques have been used to improve the precision of this measurement. The Higgs boson mass is measured to be mH=125.78±0.26GeV. This is combined with a measurement of mH already performed in the H→ZZ→4ℓ decay channel using the same data set, giving mH=125.46±0.16GeV. This result, when further combined with an earlier measurement of mH using data collected in 2011 and 2012 with the CMS detector, gives a value for the Higgs boson mass of mH=125.38±0.14GeV. This is currently the most precise measurement of the mass of the Higgs boson.
Adoptive transfer of peripheral blood-derived, melanoma-reactive CD8(+) cytotoxic T lymphocytes (CTLs) alone is generally insufficient to eliminate bulky tumors. Similarly, monotherapy with ...anti-CTLA4 infrequently yields sustained remissions in patients with metastatic melanoma. We postulated that a bolus of enhanced IL-21-primed polyclonal antigen-specific CTL combined with CTLA4 blockade might boost antitumor efficacy. In this first-in-human case study, the combination successfully led to a durable complete remission (CR) in a patient whose disease was refractory to both monoclonal CTL and anti-CTLA4. Long-term persistence and sustained anti-tumor activity of transferred CTL, as well as responses to nontargeted antigens, confirmed mutually beneficial effects of the combined treatment. In this first-in-human study, Chapuis et al. demonstrate that the combination of adoptive cellular therapy with CTLA4 blockade induces long-term remission in a melanoma patient resistant to both modalities administered serially and individually.
CD8+ T cells are master effectors of antitumor immunity, and their presence at tumor sites correlates with favorable outcomes. However, metabolic constraints imposed by the tumor microenvironment ...(TME) can dampen their ability to control tumor progression. We describe lipid accumulation in the TME areas of pancreatic ductal adenocarcinoma (PDA) populated by CD8+ T cells infiltrating both murine and human tumors. In this lipid-rich but otherwise nutrient-poor TME, access to using lipid metabolism becomes particularly valuable for sustaining cell functions. Here, we found that intrapancreatic CD8+ T cells progressively accumulate specific long-chain fatty acids (LCFAs), which, rather than provide a fuel source, impair their mitochondrial function and trigger major transcriptional reprogramming of pathways involved in lipid metabolism, with the subsequent reduction of fatty acid catabolism. In particular, intrapancreatic CD8+ T cells specifically exhibit down-regulation of the very-long-chain acyl-CoA dehydrogenase (VLCAD) enzyme, which exacerbates accumulation of LCFAs and very-long-chain fatty acids (VLCFAs) that mediate lipotoxicity. Metabolic reprogramming of tumor-specific T cells through enforced expression of ACADVL enabled enhanced intratumoral T cell survival and persistence in an engineered mouse model of PDA, overcoming one of the major hurdles to immunotherapy for PDA.
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