Mutation landscape in Chinese nodal diffuse large B-cell lymphoma by targeted next generation sequencing and their relationship with clinicopathological characteristics.

BACKGROUND: Diffuse large B-cell lymphoma (DLBCL), an aggressive and heterogenic malignant entity, is still a challenging clinical problem, since around one-third of patients are not cured with primary treatment. Next-generation sequencing (NGS) technologies have revealed common genetic mutations in DLBCL. We devised an NGS multi-gene panel to discover genetic features of Chinese nodal DLBCL patients and provide reference information for panel-based NGS detection in clinical laboratories. METHODS: A panel of 116 DLBCL genes was designed based on the literature and related databases. We analyzed 96 Chinese nodal DLBCL biopsy specimens through targeted sequencing. RESULTS: The most frequently mutated genes were KMT2D (30%), PIM1 (26%), SOCS1 (24%), MYD88 (21%), BTG1 (20%), HIST1H1E (18%), CD79B (18%), SPEN (17%), and KMT2C (16%). SPEN (17%) and DDX3X (6%) mutations were highly prevalent in our study than in Western studies. Thirty-three patients (34%) were assigned as genetic classification by the LymphGen algorithm, including 12 cases MCD, five BN2, seven EZB, seven ST2, and two EZB/ST2 complex. MYD88 L265P mutation, TP53 and BCL2 pathogenic mutations were unfavorable prognostic biomarkers in DLBCL. CONCLUSIONS: This study presents the mutation landscape in Chinese nodal DLBCL, highlights the genetic heterogeneity of DLBCL and shows the role of panel-based NGS to prediction of prognosis and potential molecular targeted therapy in DLBCL. More precise genetic classification needs further investigations.

Gastrointestinal pH-Sensitive Pickering Emulsions Stabilized by Zein Nanoparticles Coated with Bioactive Glycyrrhizic Acid for Improving Oral Bioaccessibility of Curcumin.

Pickering emulsions have received considerable attention for their stability and functionality. Environmentally responsive Pickering emulsions could be used as vehicles for oral administration. However, challenges still exist, such as nonbiocompatibility of emulsifier and mismatched response behavior in the gastrointestinal environment. In this study, a strategy was proposed that bioactive saponin glycyrrhizic acid (GA) was used as a pH-responsive substance to functionalize zein nanoparticles, and tannic acid (TA) was used as a primer for cross-linking GA and zein nanoparticles. The Pickering emulsions fabricated by zein/TA/GA nanoparticles (ZTGs) exhibited excellent stability at acid conditions while slowly demulsifying at neutral conditions, which can be further used as an intestine-targeted delivery system. Curcumin was encapsulated into ZTG-stabilized Pickering emulsions, and the encapsulation efficiency results suggested that the presence of GA coating remarkably facilitated the encapsulation of curcumin. An in vitro digestion study suggested that ZTGs provided protection for emulsions from pepsin hydrolysis and exhibited higher free fatty acid release as well as higher bioaccessibility of curcumin during simulated intestine digestion. This study provides an effective strategy to prepare pH-responsive Pickering emulsions for improving the oral bioaccessibility of hydrophobic nutraceuticals.

Functional System Based on Glycyrrhizic Acid Supramolecular Hydrogel: Toward Polymorph Control, Stabilization, and Controlled Release.

Developments of a drug delivery system (DDS) based on a natural supramolecular hydrogel have been of wide interest due to its biocompatibility, efficacy, and adjustable performance. However, a simple and efficient design of functional hydrogel DDS based on the templated interplay of gelator and model drug is still a challenge. In this work, natural glycyrrhetinic acid (GA) gel was selected as a carrier to encapsulate the model drug pyrazinamide (PZA). It was found that the carboxyl-amide interaction at the interface of gel-drug achieved polymorph control, stabilization, and pH-responsive release. Powder X-ray diffraction confirmed that the metastable γ form of PZA was obtained from the GA gel. Spectral analysis and molecular dynamics simulation showed that the protonation at the amide-O promoted the discretization of PZA molecules in solution, resulting in the polymorphism. Furthermore, the gel-drug interplay increased the stability of the γ form significantly from 2 days to 3 months by in situ encapsulation in the GA gel. In vitro release study indicated that the GA gel achieved targeted control release of PZA due to the pH-responsiveness property of GA. This work provides a promising option for hydrogel-based DDS design combined with polymorph control and stabilization.

SpyStapler-mediated assembly of nanoparticle vaccines.

The COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has wreaked havoc around the globe, with no end in sight. The rapid emergence of viral mutants, marked by rapid transmission and effective immune evasion, has also posed unprecedented challenges for vaccine development, not least in its speed, mass production, and distribution. Here we report a versatile "plug-and-display" strategy for creating protein vaccines, including those against malaria parasites and SARS-CoV-2, through the combined use of the intrinsically disordered protein ligase SpyStapler and computationally designed viral-like particles. The resulting protein nanoparticles harboring multiple antigens induce potent neutralizing antibody responses in mice, substantially stronger than those induced by the corresponding free antigens. This modular vaccine design enabled by SpyStapler furnishes us with a new weapon for combatting infectious diseases. Electronic Supplementary Material: Supplementary material (further details of the protein sequences, cloning procedures, TEM imaging, ELISA details, and reaction controls) is available in the online version of this article at 10.1007/s12274-022-4951-9.

Diversity and Professional Advancement in Medical Physics.

Purpose: While disparities in the inclusion and advancement of women and minorities in science, technology, engineering, mathematics, and medical fields have been well documented, less work has focused on medical physics specifically. In this study, we evaluate historical and current diversity within the medical physics workforce, in cohorts representative of professional advancement (PA) in the field, and within National Institutes of Health (NIH)-funded medical physics research activities. Methods and Materials: The 2020 American Association of Physicists in Medicine (AAPM) membership was queried as surrogate for the medical physics workforce. Select subsets of the AAPM membership were queried as surrogate for PA and early career professional advancement (ECPA) in medical physics. Self-reported AAPM-member demographics data representative of study analysis groups were identified and analyzed. Demographic characteristics of the 2020 AAPM membership were compared with those of the PA and ECPA cohorts and United States (US) population. The AAPM-NIH Research Database was appended with principal investigator (PI) demographics data and analyzed to evaluate trends in grant allocation by PI demographic characteristics. Results: Women, Hispanic/Latinx/Spanish individuals, and individuals reporting a race other than White or Asian alone comprised 50.8%, 18.7%, and 32.4% of the US population, respectively, but only 23.9%, 9.1%, and 7.9% of the 2020 AAPM membership, respectively. In general, representation of women and minorities was further decreased in the PA cohort; however, significantly higher proportions of women (P < .001) and Hispanic/Latinx/Spanish members (P < .05) were observed in the ECPA cohort than the 2020 AAPM membership. Analysis of historical data revealed modest increases in diversity within the AAPM membership since 2002. Across NIH grants awarded to AAPM members between 1985 and 2020, only 9.4%, 5.3%, and 1.7% were awarded to women, Hispanic/Latinx/Spanish, and non-White, non-Asian PIs, respectively. Conclusions: Diversity within medical physics is limited. Proactive policy should be implemented to ensure diverse, equitable, and inclusive representation within research activities, roles representative of PA, and the profession at large.

PD-L1 over-expression is driven by B-cell receptor signaling in diffuse large B-cell lymphoma.

Targeting the programmed death 1 (PD-1)/programmed death ligand 1 (PD-L1) pathway represents a milestone in cancer therapy. However, the biologic features of diffuse large B-cell lymphoma (DLBCL) with PD-L1 expression remains unknown. We evaluated the correlation between pSYK and PD-L1 mRNA levels with RNAscope in situ hybridization and protein levels with immunohistochemistry in 108 cases of DLBCL, 25 of which featured loss of B-cell receptor (BCR), and investigated the effects of BCR signaling and MYC on PD-L1 mRNA and protein level with qPCR, immunoblotting and flow cytometery in DLBCL cell lines. PD-L1 amplification was detected with fluorescent in situ hybridization. Animal studies were applied to validate the in vitro findings. pSYK and MYC correlated with both PD-L1 mRNA and protein level. Genetic aberrations involving PD-L1 were rare in DLBCL. BCR signaling and MYC increased PD-L1 mRNA and protein expression. Inhibition of BCR signaling and BCR knockdown down-regulated PD-L1. DLBCL with a loss of loss of BCR showed low levels of PD-L1 mRNA and protein. PD-L1 was down-regulated by ibrutinib in a xenograft mouse model and correlated with slower tumor growth. In conclusion, this study demonstrates that DLBCL with PD-L1 expression features an activated B-cell receptor signal pathway, and that BCR inhibition and PD-L1 blockage may potentially synergize to targeting DLBCL.

Expression of PHLPP2 correlates with clinicopathologic characteristics and prognosis in colorectal cancer.

PH domain leucine-rich repeat protein phosphatase 2 (PHLPP2) belongs to the phosphokinase family, that has been reported to play an important role in several cancers. However, the expression of PHLPP2 and its correlation with clinicopathologic characteristics in colorectal cancer (CRC) have yet to be determined. The aim of this study is to investigate the expression of PHLPP2 and explore its role in CRC. The expression of PHLPP2, PTEN, PI3KCA, and PI3KCB in 130 cases of CRC and normal tissues was assessed by immunohistochemistry. In addition, the expression of PHLPP2, PTEN, PI3KCA, and PI3KCB in 32 pairs of CRC tissues and their corresponding normal tissues was determined by RT-PCR and western blotting, respectively. PHLPP2 expression in CRC was significantly lower than that of normal tissues. However, PHLPP2 mRNA shows no significant difference between CRC and normal tissue. PTEN expression in left colorectal cancer (LCC) was absent, while PI3KCA and PI3KCB in right colorectal cancer (RCC) were significantly higher than those in LCC. PHLPP2 was negatively correlated with p-Akt1 in CRC. The expression of p-Akt1 in PHLPP2 (+)/PTEN (+) in CRC tissues was significantly lower than that in other groups. PHLPP2 expression was correlated with differentiation, invasion, and lymph node metastasis. Kaplan-Meier analysis and multivariate analysis reveal that PHLPP2 is closely related to prognosis; more importantly, it is an independent prognostic factor for CRC. In conclusion, PHLPP2 may play a major role in the development, metastasis, and prognosis of CRC.

Integrative analysis of PD-L1 DNA status, mRNA status and protein status, and their clinicopathological correlation, in diffuse large B-cell lymphoma.

AIMS: The protein expression of programmed death-ligand 1 (PD-L1) has been recognised as being a biomarker for poor prognosis in diffuse large B-cell lymphoma (DLBCL). The aims of this study were to determine PD-L1 DNA status and mRNA status, and to explore whether they contribute to protein expression and their clinicopathological correlation in DLBCL. METHODS AND RESULTS: In this study, we used fluorescence in-situ hybridisation, RNA in-situ hybridisation and immunohistochemistry to determine PD-L1 status at three different levels in 287 DLBCL samples with follow-up. Their correlation and clinical pathological relevance were also analysed. Our results showed that 1.7% (3/175) of patients had PD-L1 DNA amplification, 19.9% (57/287) had high PD-L1 mRNA expression, and 11.8% (34/287) had high PD-L1 protein expression. Both mRNA and protein expression of PD-L1 were significantly higher in non-germinal centre B-cell-like (GCB) DLBCL than in GCB DLBCL (P < 0.05). In addition, the patients with high PD-L1 mRNA or high PD-L1 protein expression but no PD-L1 DNA amplification had significantly poorer overall survival (OS) than those with low PD-L1 expression (P < 0.05). Furthermore, we found that PD-L1 mRNA and PD-L1 protein expression were highly correlated (P = 0.012), which was observed in all three samples with DNA amplification. CONCLUSIONS: PD-L1 DNA amplification is a rare event, PD-L1 mRNA is the main contributor to the high PD-L1 protein expression, and the latter two will serve as important biomarkers for predicting the prognosis of DLBCL patients and selecting them for immunotherapy.

Primary breast diffuse large B-cell lymphoma: a population-based study from 1975 to 2014.

Primary breast diffuse large B-cell lymphoma (DLBCL) is a rare non-Hodgkin's lymphoma with limited data. In this study, a population-based study of primary breast DLBCL in the United States was performed to determine its incidence trends, prognostic factors, survival, the role of surgery as well as the comparison with nodal DLBCL. 1021 patients diagnosed with breast DLBCL were identified in the Surveillance, Epidemiology, and End Results (SEER) cancer registries from 1973-2014. The incidence of both breast and nodal DLBCL increased over time. Patients with breast DLBCL were older, mainly women, diagnosed at earlier stages and had lower prevalence in white and black races compared with nodal DLBCL. Multivariate analysis revealed older age (≥ 70 years old) and advanced stage as independent predictors of worse OS. Independent predictor of better DSS were younger age (< 70 years old), early stage and diagnosis after 2000. When analyzed according to age, stage, race, tumor laterality and year of diagnosis, the overall survival did not benefit from surgery except in patients diagnosed between 2001-2010 and the surgery rate decreased overtime. Compared with nodal DLBCL, breast DLBCL patients exhibited a better outcome. In conclusion, breast DLBCL is a rare tumor with increasing incidence and improved survival over the last four decades. The introduction of rituximab seems to improve the outcome of breast DLBCL. Further studies are needed to advance our understanding of breast DLBCL and optimize the treatment strategy.

Expression and clinical significances of Beclin1, LC3 and mTOR in colorectal cancer.

Autophagy is related to cancer and other diseases, and compromised autophagy could promote chromosome instability associated with carcinogenesis and tumor progression. The role of autophagy in the growth and metastasis of colorectal cancer (CRC) remains poorly understood. Beclin1 mediates autophagic initiation, and LC3 is a specific marker for autophagy. Inactivation of mTOR caused by cellular hypoxia or energy deficiency induces autophagic activity. This study aims to examine the expression and clinical significance of these proteins in CRC. Immunohistochemistry results showed that the positive expression rates of Beclin1, LC3, and mTOR in cancer tissues were 90.50%, 87.19%, and 46.28%, respectively, which were higher than those in adjacent tissues (P < 0.05). Differentiation degree and lymph node metastasis were associated with LC3 overexpression (P < 0.05) but not with Beclin1 (P > 0.05). Lymph node metastasis was also related to mTOR. Spearman analysis results showed that LC3 expression was positively correlated with Beclin1 but negatively correlated with mTOR (r = 0.593 and -0.165, respectively; P < 0.01). Beclin1 expression was also not associated with mTOR (P > 0.05). Survival analysis further indicated that LC3, mTOR, and lymph node metastasis were independent prognostic factors in CRC. Real-time PCR results and Western blot indicated that Beclin1, LC3, and mTOR expression in CRC was significantly higher than that in adjacent tissues (P < 0.01). The aberrant protein expression may be associated with the development and progression of CRC. The LC3 and mTOR genes must be simultaneously detected to evaluate progression and prognosis of CRC.

Expression of PI3Kp110α and PI3Kp110ß in the colorectal conventional adenoma, serrated lesions and adenoma with canceration and their significance.

AIMS: To evaluate the expression and clinical significance of PI3Kp110α and PI3Kp110ß in colorectal conventional adenoma, serrated lesions and adenoma with canceration. METHODS AND RESULTS: Immunohistochemistry and Western blot analysis were conducted to detect the expression of p110α and p110ß in normal colorectal tissues, conventional adenoma, serrated lesions and adenoma canceration. Results revealed that the expression of P110α and P110ß in the adenoma canceration was significantly higher than that in normal tissues, tubular adenoma (low grade) and tubular-villous adenoma (low grade) of conventional adenoma, hyperplastic polyps of serrated lesions (P<0.05). But there was no significant difference between the adenoma canceration and the high grade adenoma of conventional adenoma, all grade of villous adenoma and serrated adenoma (P>0.05). The expression of p110α and p110ß was correlated with different clinicopathologic factors in conventional adenoma, serrated adenoma and adenoma canceration (P<0.05). CONCLUSIONS: p110α and p110ß were highly expressed in villous adenoma, serrated adenoma and adenoma with canceration. Its high expression may be the risk factor of the progress of adenoma to adenocarcinoma, and may be an important cause of what canceration rate of villous adenoma and serrated adenoma was higher than that of other adenomas. Combined detection of p110α and p110ß is helpful to determine the canceration potential of colorectal villous adenoma and serrated adenoma.

PIK3CA and PIK3CB expression and relationship with multidrug resistance in colorectal carcinoma.

The phosphoinositide 3-kinases (PI3Ks) are a critical family of signaling enzymes that participate in many cellular processes that promote the transformation of a normal cell into a cancer cell. These processes include cancer cell proliferation, migration, and invasion. However, the correlation between PI3Ks and multidrug resistance (MDR) remains unclear. The prognostic value of PI3Ks has not been previously evaluated. Thus, this study aims to evaluate the association between PIK3CA and PIK3CB expression and the MDR gene in colorectal cancer (CRC) patients. Immunohistochemistry was employed to detect the expressions of PIK3CA, PIK3CB, MDR-1, LRP, GST-π, and Topo II in 316 CRC specimens. Patients were followed-up annually by telephone or at an outpatient clinic. Results revealed that PIK3CA and PIK3CB expression was correlated with the degree of tumor differentiation and lymph node metastasis (P < 0.05). The overexpression of MDR-1, LRP, Topo II, and GST-π was found to be 72.78%, 70.89%, 77.53%, and 76.58% of CRC, respectively. Correlation analysis showed that PIK3CA and PIK3CB expression exhibits a positive correlation with MDR-1, LRP, and GST-π with correlation coefficients of 0.288, 0.128, and 0.197, respectively (P < 0.05). Kaplan-Meier analysis revealed that the five-year survival rate of patients without lymph node metastasis, positive expression of PIK3CA and PIK3CB, and negative expression of GST-π and MDR-1 was higher than those with these characteristics. Multivariate analysis revealed that GST-π, MDR-1 expression, and lymph node metastasis could serve as independent predictive factors of overall survival. The expression of both PIK3CA and PIK3CB is increased and related to the development and progress of colorectal carcinoma and MDR. The combined detection of PIK3CA andPIK3CB is important for patients with colorectal carcinoma in prognosis and optimal therapy.