Quantum pBac: An effective, high-capacity piggyBac-based gene integration vector system for unlocking gene therapy potential.

Recent advances in gene therapy have brought novel treatment options for cancer. However, the full potential of this approach has yet to be unlocked due to the limited payload capacity of commonly utilized viral vectors. Virus-free DNA transposons, including piggyBac, have the potential to obviate these shortcomings. In this study, we improved a previously modified piggyBac system with superior transposition efficiency. We demonstrated that the internal domain sequences (IDS) within the 3' terminal repeat domain of hyperactive piggyBac (hyPB) donor vector contain dominant enhancer elements. Plasmid-free donor vector devoid of IDS was used in conjunction with a helper plasmid expressing Quantum PBase™ v2 to generate an optimal piggyBac system, Quantum pBac™ (qPB), for use in T cells. qPB outperformed hyPB in CD20/CD19 CAR-T production in terms of performance as well as yield of the CAR-T cells produced. Furthermore, qPB also produced CAR-T cells with lower donor-associated variabilities compared to lentiviral vector. Importantly, qPB yielded mainly CD8+ CAR-TSCM cells, and the qPB-produced CAR-T cells effectively eliminated CD20/CD19-expressing tumor cells both in vitro and in vivo. Our findings confirm qPB as a promising virus-free vector system with an enhanced payload capacity to incorporate multiple genes. This highly efficient and potentially safe system will be expected to further advance gene therapy applications.

A hybrid model combining wavelet transform and recursive feature elimination for running state evaluation of heat-resistant steel using laser-induced breakdown spectroscopy.

Heat-resistant steel is widely used in various industries, and the running state is of great importance for equipment function and safety. In this work, laser-induced breakdown spectroscopy (LIBS) is applied to evaluate the running state of steel using indicators of micro and macro properties. The hybrid model based on wavelet threshold denoising (WTD) and K-fold-support vector machine-recursive feature elimination (K-SVM-RFE) is proposed to estimate the different indictors of various service conditions of steel. Fourteen T91 specimens, including 4 industrial specimens obtained from different service conditions in the power plant boiler, were used as the analytes. Firstly, the noise signal of the LIBS spectra of each specimen was analyzed and removed with WTD. Secondly, an improved approach K-SVM-RFE was applied to select the optimal feature subset and build the classification models of aging grade and hardness grade. The influence of denoising pretreatment on model performance was compared and discussed. Finally, the assessment matrix, established using the indicators from the aging grade and hardness grade, was used to evaluate the running state of steel. The results show that the test assessment matrix obtained with the hybrid model based on WTD and K-SVM-RFE is consistent with the reference matrix on the running state of steel.

Prognostic value of dynamic albumin-to-alkaline phosphatase ratio in limited stage small-cell lung cancer.

AIM: To dynamically investigate the prognostic value of albumin-to-alkaline phosphatase ratio (AAPR) in limited stage small-cell lung cancer. MATERIALS & METHODS:  The AAPR within 1 week before and after chemoradiation therapy (pre- and post-AAPR) was collected and analyzed. RESULTS: Patients with low pre- or post-AAPR had shorter overall survival and progression-free survival than the high groups (p-values all <0.05). Post-AAPR was an independent prognostic factor for progression-free survival (p = 0.007) and overall survival (p = 0.003). The integration of pre- or post-AAPR improved the prognostic ability of Tumor, Node, Metastasis stage alone (0.55-0.64 and 0.68, respectively). CONCLUSION:  Post-AAPR is a reliable prognostic factor for limited stage small-cell lung cancer patients. The complementary value of AAPR to Tumor, Node, Metastasis stage is worth further validation in the future.

Non-invasive plasma testing for CD274 UTR structural variations by next-generation sequencing in cancer.

Immunotherapy is now the main choice of systemic therapy for many cancer patients, while current biomarkers for tumor immunotherapy may be limited by the accessibility of patient tumor tissue or tumor neoplastic content. Rare mutation in the 5' and 3'-untranslated region (UTR) of CD274 gene (Protein name: PD-L1) has been recently reported in hematologic and solid tumors as a potential biomarker for assessing efficacy during immunotherapy. However, multi-omics analysis for CD274 UTR region, especially circulating tumor DNA (ctDNA), have been little explored in the pan-cancer perspective. We developed a cSMART2.0 technology featured with higher capture efficiency and homogeneity to detect this rare structural variant in 2249 Chinese patients' cohort with multiple cancers. An incidence of 0.36% was detected in this cohort, consistent with TCGA (The Cancer Genome Atlas), while the prevalence of SV in CD274 UTR region in liver and breast cancer were significantly higher than TCGA. The liquid biopsy result from ctDNA was 100% concordance with gDNA result getting from tumor tissue detection, and further validated by immunohistochemistry (IHC) and multiplex immunofluorescence (mIF) experiments. Patients carrying this SV in CD274 UTR region without driver gene mutation responded to immune checkpoint inhibitors (ICIs). This study proves that rare structural variants in CD274 UTR region exist in various cancer in Chinese population for the first time, which can induce immune escape and be used for prediction of response to ICIs. Liquid biopsy based cSMART 2.0 technology could offer more sensitive and accurate detection to navigate potential ICIs patients and to benefit patients with advanced disease when tissue samples are not available.

The predictive value of neutrophil-to-lymphocyte ratio for overall survival and pathological complete response in breast cancer patients receiving neoadjuvant chemotherapy.

Purpose: Previous studies have reported that neutrophil-to-lymphocyte ratio (NLR) at pre-treatment was predictive for overall survival (OS) and pathologic complete response (pCR) in breast cancer (BC) patients receiving neoadjuvant chemotherapy (NAC). This study aims to explore the predictive role of both pre- and post-NLR for OS as well as longitudinal NLR kinetics towards pCR in BC patients undergoing NAC. Methods: We retrospectively included 501 BC patients who received NAC from 2009 to 2018. NLR at pre-, mid (every two cycles of NAC)-, and post-treatment were collected. Overall, 421 patients were included in the survival analysis. These patients were randomly divided into a training cohort (n = 224) and a validation cohort (n = 197). A multivariable Cox model was built using all significant factors in the multivariable analysis from the training cohort. The performance of the model was verified in the validation cohort by the concordance index (C-index). Longitudinal analysis for pCR prediction of NLR was performed using a mixed-effects regression model among 176 patients who finished eight cycles of NAC. Results: The median follow-up time was 43.2 months for 421 patients. In the training cohort, multivariable analysis revealed that ER status, clinical node stage, pCR, pre-NLR, and post-NLR (all p < 0.05) were independent predictors of OS. The OS nomogram was established based on these parameters. The C-indexes of the nomogram were 0.764 and 0.605 in the training and validation cohorts, respectively. In the longitudinal analysis, patients who failed to achieve pCR experienced an augment of NLR during NAC while NLR remained stable among patients with pCR. Pre-NLR tended to be significantly associated with OS in patients of HER2 overexpressing and TNBC subtypes (all p < 0.05), but not in Luminal A and Luminal B subtypes. Conclusions: This study demonstrated the prognostic value of both pre-NLR and post-NLR on clinical outcomes in BC patients receiving NAC. A novel nomogram was established to predict OS. Non-pCR patients developed increased NLRs during NAC. Routine assessment of NLR may be a simple and affordable tool to predict prognosis for BC patients receiving NAC.

Impact of effective dose to immune cells (EDIC) on lymphocyte nadir and survival in limited-stage SCLC.

BACKGROUND: Effective dose to immune cell (EDIC), an estimated radiation dose to the circulating lymphocytes, is of significance for overall survival (OS) in non-small cell lung cancer. This study aimed to validate the EDIC's OS effect on limited-stage small cell lung cancer (LS-SCLC). METHOD AND MATERIALS: This study included LS-SCLC patients received definitive chemo-radiation in one single center from 2012 to 2017. All patients had multiple complete-blood-count tests including lymphocyte count at pre-, during- and end- radiotherapy. EDIC, computed according to doses of the lung, heart, and the total body, was assessed for its correlation with lymphocyte nadir, OS and progression free survival (PFS). RESULTS: Of 503 eligible patients, the mean EDIC was 7.34 Gy. The mean lymphocyte nadir was 0.48 × 109 cells/L, significantly lower than 1.65 × 109 cells/L at pre-radiotherapy (p < 0.001). EDIC was significantly correlated with lymphocyte nadir under both univariate (p < 0.001) and multivariable linear regression (p < 0.001). Multivariable analysis showed EDIC (HR = 0.1072, p = 0.005) and lymphocyte nadir (HR = 0.345, p = 0.003) were both significant for OS. EDIC was also significant for PFS (HR = 1.046, p = 0.026). The C-indexes of OS prediction were 0.593, 0.617, 0.676, and 0.684, for lymphocyte nadir alone, EDIC alone, combined lymphocyte nadir model, and combined EDIC model, respectively. CONCLUSIONS: This study demonstrated that EDIC is an independent predictor for lymphocyte nadir, PFS and OS. EDIC may serve as a predictor for lymphocyte nadir and a surrogate marker for OS in LS-SCLC. More attention should be paid to EDIC to decease the lymphocyte toxicity and improve survival.

Pre-radiotherapy lymphocyte count and platelet-to-lymphocyte ratio may improve survival prediction beyond clinical factors in limited stage small cell lung cancer: model development and validation.

BACKGROUND: Few small sample size studies have reported lymphocyte count was prognostic for survival in small-cell lung cancer (SCLC). This study aimed to validate this finding, to build prediction model for overall survival (OS) and to study whether novel models that combine lymphocyte-related variables can predict OS more accurately than a conventional model using clinical factors alone in a large cohort of limited-stage SCLC patients. METHODS: This study enrolled 544 limited-stage SCLC patients receiving definitive chemo-radiation with pre-radiotherapy lymphocyte-related variables including absolute lymphocyte count (ALC), platelet-to-lymphocyte ratio (P/L ratio), neutrophil-to-lymphocyte ratio (N/L ratio), and lymphocyte-to-monocyte ratio (L/M ratio). The primary endpoint was OS. These patients were randomly divided into a training dataset (n=274) and a validation dataset (n=270). Multivariate survival models were built in the training dataset, and the performance of these models were further tested in the validation dataset using the concordance index (C-index). RESULTS: The median follow-up time was 36 months for all patients. In the training dataset, univariate analysis showed that ALC (P=0.020) and P/L ratio (P=0.023) were significantly correlated with OS, while L/M ratio (P=0.091) and N/L ratio (P=0.436) were not. Multivariate modeling demonstrated the significance of ALC (P=0.063) and P/L ratio (P=0.003), and the improvement for OS prediction in combined models with the addition of ALC (C-index =0.693) or P/L ratio (C-index =0.688) over the conventional model (C-index =0.679). The validation dataset analysis confirmed a modest improvement of C-index with the addition of ALC or P/L ratio. All these models showed reasonable discriminations and calibrations. CONCLUSIONS: This study validated the significant value of pre-radiotherapy ALC and P/L ratio on OS in limited-stage SCLC. The combined model with ALC or P/L ratio showed additional OS prediction values than the conventional model with clinical factors alone.

Delineating the pattern of treatment for elderly locally advanced NSCLC and predicting outcomes by a validated model: A SEER based analysis.

INTRODUCTION: Locally advanced nonsmall-cell lung cancer (LA-NSCLC) represented a highly heterogeneous group, with more than half of the patients aged ≥65 years at the time of diagnosis. However, the optimal treatment for elderly LA-NSCLC patients was still not defined. METHODS: A total of 33530 elderly patients (≥65 years) diagnosed with LA-NSCLC from 2004 to 2014 were identified from Surveillance, Epidemiology, and End Results (SEER) database. RESULTS: Locally advanced nonsmall-cell lung cancer patients aged 65-74 years were more frequently treated with chemoradiotherapy (CRT) (40%), while patients aged ≥75 years received more best supportive care (BSC) (36%). For age group of 65-74 years, patients who had surgery with or without (neo)adjuvant therapy had a median survival of 28 months, CRT 15 months, radiotherapy (RT) alone 6 months, chemotherapy alone 11 months, and BSC 3 months; while for patients aged ≥ 75 years, the median OS was 20, 13, 7, 9, and 2, respectively. Besides, independent clinicopathological factors were integrated into nomograms for OS and CSS prediction, C-indexes achieved 0.692 and 0.698, respectively. Importantly, the discrimination of nomogram was superior to that of the American Joint Committee on Cancer TNM classification (0.742 vs 0.572 for training set and 0.731 vs 0.565 for validation set). CONCLUSION: For elderly patients with LA-NSCLC, the curative-intent treatment (surgery or CRT) conferred better survival compared to chemotherapy alone, RT alone and BSC. The proposed nomograms based on independent clinicopathological variables may be practical and helpful for precise evaluation of patient prognosis, and guiding the individualized treatment for elderly LA-NSCLC.

[Value of functional magnetic resonance imaging in predicting outcomes of neoadjuvant chemoradiotherapy in rectal cancer].

Rectal cancer is one of the common cancers which poses a threat to the health of mankind. In recent years. Multi-modality treatment strategies for locally advanced rectal cancer improve the treatment efficiency. Accurate prediction of the treatment response after the neoadjuvant chemoradiotherapy (CRT) can guide more suitable treatment strategy. MERCURY study proved the prognostic value of post-CRT standard morphologic MRI(T2-weighted) assessment of tumor regression grade(TRG), and MRI assessment of circumferential resection margin can guide the definitive surgery. Compared with standard morphologic MRI (T2-weighted), functional MRI, including diffusion weighted imaging (DWI) and dynamic contrast enhanced (DCE) MRI, has shown more promising results for the prediction of therapeutic response in rectal cancer. The addition of diffusion-weighted images to T2-weighted images improves the accuracy of restaging examinations for determination of complete pathologic responders. DCE can reflect the tumor micro-vascular environment, and the change of perfusion in response to treatment. These images have the potential to improve the accuracy of therapeutic response in rectal cancer.

Dnmt3a haploinsufficiency cooperates with oncogenic Kras to promote an early-onset T-cell acute lymphoblastic leukemia.

Mutations in DNA methyltransferase 3A (DNMT3A) are prevalent in various myeloid and lymphoid malignancies. The most common DNMT3A R882 mutations inhibit methyltransferase activity of the remaining wild-type DNMT3A proteins at a heterozygous state due to their dominant-negative activity. Reports and COSMIC database analysis reveal significantly different frequencies of R882 mutations in myeloid versus T-cell malignancies, inspiring us to investigate whether downregulation of DNMT3A regulates malignancies of different lineages in a dose-dependent manner. In a competitive transplant setting, the survival of recipients with KrasG12D/+ ; Dnmt3a+/- bone marrow (BM) cells was significantly shortened than that of recipients with KrasG12D/+ cells. Moreover, all of the recipients with KrasG12D/+ ; Dnmt3a+/- cells developed a lethal T-cell acute lymphoblastic leukemia (T-ALL) without significant myeloproliferative neoplasm (MPN) phenotypes, while ~20% of recipients with KrasG12D/+ cells developed MPN with or without T-ALL. This is in sharp contrast to the recipients with KrasG12D/+ ; Dnmt3a-/- cells, in which ~60% developed a lethal myeloid malignancy (MPN or acute myeloid leukemia [AML]). Our data suggest that in the context of oncogenic Kras, loss of Dnmt3a promotes myeloid malignancies, while Dnmt3a haploinsufficiency induces T-ALL. This dose-dependent phenotype is highly consistent with the prevalence of DNMT3A R882 mutations in AML versus T-ALL in human.

Internal mammary lymph nodes radiotherapy of breast cancer in the era of individualized medicine.

Inclusion internal mammary lymph nodes as a part of regional nodal irradiation have a potential to reduce local recurrence, distant recurrence, and improve survival in breast cancer. However, the increased risk of cardiac toxicity and lungs injure associated with internal mammary lymph nodes irradiation has drew more and more attention. Estimating risk of metastasis in internal mammary lymph nodes based on axillary lymph nodes metastasis status is not always reliable: low-risk do not always mean negative in internal mammary lymph nodes and high-risk do not always indicate positive in internal mammary lymph nodes. Inaccurate prediction of in internal mammary lymph nodes metastasis might lead to over- or under-treatment of in internal mammary lymph node. Internal mammary sentinel lymph node biopsy is a minimally invasive technique which has a high potential to accurately evaluate the metastasis status in in internal mammary lymph nodes and improve accuracy of nodal staging. This technique might be a useful tool to guide individualized internal mammary lymph nodes irradiation.

False-positive diagnosis of disease progression by magnetic resonance imaging for response assessment in prostate cancer with bone metastases: A case report and review of the pitfalls of images in the literature.

Bone metastases are common in prostate cancer. However, differentiating neoplastic from non-neoplastic alterations of bone on images is challenging. In the present report, a rare case of bone marrow reconversion on magnetic resonance imaging (MRI) assessment, which may lead to a false-positive diagnosis of disease progression of bone metastases in hormone-resistant prostate cancer, is presented. Furthermore, a review of the literature regarding the pitfalls of images for response assessment, including the 'flare' phenomenon on bone scintigraphy, computed tomography (CT), positron emission tomography/CT and marrow reconversion on MRI is also provided. These inaccuracies, which may lead to a premature termination of an efficacious treatment, should be carefully considered by the radiologists and oncologists involved in clinical trials. The case reported in the present study showed how to assess the early therapeutic response and select the appropriate treatment for the patient when these pitfalls are encountered on clinical images.