Artificial intelligence-driven design of the assembled major cat allergen Fel d 1 to improve its spatial folding and IgE-reactivity.

BACKGROUND: Cat-derived allergens are considered as one of the most common causes of allergic diseases worldwide. Fel d 1 is a major cat allergen and plays an important role in immunoglobulin E (IgE)-reaction diagnosis. However, the two separate chains of Fel d 1 exhibited lower IgE-reactivity than its complete molecule of an assembled form, which makes it difficult to efficiently prepare and limits the application of Fel d 1 in molecular diagnosis of cat allergy. METHODS: We first applied artificial intelligence (AI) based tool AlphaFold2 to build the 3-dimensional structures of Fel d 1 with different connection modes between two chains, which were evaluated by ERRAT program and were expressed in Escherichia coli. We then calculated the expression ratios of soluble form/inclusion bodies form of optimized Fel d 1. The Circular Dichroism (CD), High Performance Liquid Chromatography-Size Exclusion Chromatography (HPLC-SEC) and reducing/non-reducing SDS-PAGE were performed to characterize the folding status and dimerization of the optimized fusion Fel d 1. The improvement of specific-IgE reactivity to optimized fusion Fel d 1 was investigated by enzyme linked immunosorbent assay (ELISA). RESULTS: Among several linkers, 2 × GGGGS got the highest scores, with an overall quality factor of 100. The error value of the residues around the junction of 2 × GGGGS was lower than others. It exhibited highest proportion of soluble protein than other Fel d 1 constructs with ERRAT (GGGGS, KK as well as direct fusion Fel d 1). The results of CD and HPLC-SEC showed the consistent folding and dimerization of two fused subunits between the optimized fusion Fel d 1 and previously well-defined direct fusion Fel d 1. The overall IgE-binding absorbance of optimized fusion Fel d 1 tested by ELISA was improved compared with that of the direct fusion Fel d 1. CONCLUSION: We firstly provided an AI-design strategy to optimize the Fel d 1, which could spontaneously fold into its native-like structure without additional refolding process or eukaryotic folding factors. The improved IgE-binding activity and simplified preparation method could greatly facilitate it to be a robust allergen material for molecular diagnosis of cat allergy.

IGF2BP2 Drives Cell Cycle Progression in Triple-Negative Breast Cancer by Recruiting EIF4A1 to Promote the m6A-Modified CDK6 Translation Initiation Process.

IGF2BP2 is a new identified N6-methyladenosine (m6A) reader and associated with poor prognosis in many tumors. However, its role and related mechanism in breast cancer, especially in triple-negative breast cancer (TNBC), remains unclear. In this study, it is found that IGF2BP2 is highly expressed in TNBC due to the lower methylation level in its promoter region. Functional and mechanical studies displayed that IGF2BP2 could promote TNBC proliferation and the G1/S phase transition of the cell cycle via directly regulating CDK6 in an m6A-dependent manner. Surprising, the regulation of protein levels of CDK6 by IGF2BP2 is related to the changes in translation rate during translation initiation, rather than mRNA stability. Moreover, EIF4A1 is found to be recruited by IGF2BP2 to promote the translation output of CDK6, providing new evidence for a regulatory role of IGF2BP2 between m6A methylation and translation. Downregulation of IGF2BP2 in TNBC cell could enhance the sensitivity to abemaciclib, a CDK4/6 inhibitor. To sum up, our study revealed IGF2BP2 could facilitate the translation output of CDK6 via recruiting EIF4A1 and also provided a potential therapeutic target for the diagnosis and treatment of TNBC, as well as a new strategy for broadening the drug indications for CDK4/6 inhibitors.

Identification of Pla a 7 as a novel pollen allergen group in Platanus acerifolia pollen.

BACKGROUND: Platanus acerifolia is recognized as a source of allergenic pollen worldwide. Currently, five Platanus acerifolia pollen allergens belonging to different protein families have been identified, in which profilin and enolase were characterized by our group recently. Besides, we also screened and identified a novel allergen candidate as triosephosphate isomerase, which was different from already known types of pollen allergens. However, the role of this novel allergen group in Platanus acerifolia pollen allergy was unclear. Therefore, we further investigated the allergenicity and clarify its clinical relevance in this study. METHODS: The natural triosephosphate isomerase from Platanus acerifolia pollen was purified by three steps of chromatography and identified by mass spectrometry. The cDNA sequence of this protein was matched from in-house transcripts based on internal peptide sequences, which was further confirmed by PCR cloning. The recombinant triosephosphate isomerase was expressed and purified from E. coli. Allergenicity analysis of this protein was carried out by enzyme linked immunosorbent assay, immunoblot, and basophil activation test. RESULTS: A novel allergen group belonging to triosephosphate isomerase was firstly identified in Platanus acerifolia pollen and named as Pla a 7. The cDNA of Pla a 7 contained an open reading frame of 762 bp encoding 253 amino acids. The natural Pla a 7 displayed 41.4% IgE reactivity with the patients' sera by ELISA, in which the absorbance value showed correlation to the serum sIgE against Platanus acerifolia pollen extract. Inhibition of IgE-binding to pollen extracts reached 26%-94% in different Pla a 7-positive sera. The recombinant Pla a 7 exhibited weaker IgE-reactivity in ELISA than its natural form, but showed comparable activity in immunoblot. The allergenicity was further confirmed by basophil activation test. CONCLUSIONS: Triosephosphate isomerase (Pla a 7) was first recognized as pollen allergen in Platanus acerifolia pollen, which is a completely different type of pollen allergen from those previously reported. This finding is essential to enrich information on allergen components and pave the way for molecular diagnosis or treatment strategies for Platanus acerifolia pollen allergy.

IGF2BP3 mediates the mRNA degradation of NF1 to promote triple-negative breast cancer progression via an m6A-dependent manner.

BACKGROUND: N6-methyladenosine (m6A) is an abundant reversible modification in eukaryotic mRNAs. Emerging evidences indicate that m6A modification plays a vital role in tumourigenesis. As a crucial reader of m6A, IGF2BP3 usually mediates the stabilisation of mRNAs via an m6A-dependent manner. But the underlying mechanism of IGF2BP3 in the tumourigenesis of triple-negative breast cancer (TNBC) is unclear. METHODS: TCGA cohorts were analysed for IGF2BP3 expression and IGF2BP3 promoter methylation levels in different breast cancer subtypes. Colony formation, flow cytometry assays and subcutaneous xenograft were performed to identify the phenotype of IGF2BP3 in TNBC. RNA/RNA immunoprecipitation (RIP)/methylated RNA immunoprecipitation (MeRIP) sequencing and luciferase assays were used to certify the target of IGF2BP3 in TNBC cells. RESULTS: IGF2BP3 was highly expressed in TNBC cell lines and tissues. TET3-mediated IGF2BP3 promoter hypomethylation led to the upregulation of IGF2BP3. Knocking down IGF2BP3 markedly reduced the proliferation of TNBC in vitro and in vivo. Intersection co-assays revealed that IGF2BP3 decreased neurofibromin 1 (NF1) stabilisation via an m6A-dependent manner. NF1 knockdown could rescue the phenotypes of IGF2BP3 knockdown cells partially. CONCLUSION: TET3-mediated IGF2BP3 accelerated the proliferation of TNBC by destabilising NF1 mRNA via an m6A-dependent manner. This suggests that IGF2BP3 could be a potential therapeutic target for TNBC.

Aiming to IgE: Drug development in allergic diseases.

The incidence of allergic disease significantly increases in recent decades, causing it become a major public health problem all over the world. The common allergic diseases such as allergic dermatitis, allergy rhinitis, allergic asthma and food allergy are mediated, at least in part, by immunoglobulin E (IgE), and so IgE acts as a central role in allergic diseases. IgE can interact with its high-affinity receptor (FcεRⅠ) which is primarily expressed on tissue-resident mast cells and circulating basophils, initiating intracellular signal transduction and then causing the activation and degranulation of mast cells and basophils. On the other hand, IgE interaction with its low-affinity receptor (CD23), can regulate various IgE-mediated immune responses including IgE-allergen complex presentation, IgE synthesis, the growth and differentiation of both B and T cells, and the secretion of pro-inflammatory mediators. With the deeper mechanism research for allergic diseases, new therapeutic strategies for interfering IgE are developed and receive a great attention. In this review, we summarize a current profile of therapeutic strategies for interfering IgE in allergic diseases. Besides, we suggest that targeting memory B cells (including long-lived plasma cells and (or) IgE+ memory B cells) may help to completely control allergic diseases, and highlight that the development of drugs synergistically aiming to multiple targets can be a better choice for improving treatment efficacy which results from allergic diseases as the systemic disorders caused by an impaired immune system.

Identification and characterization of natural PR-1 protein as major allergen from Humulus japonicus pollen.

BACKGROUND: The Humulus japonicus pollen is one of the most common allergenic pollens in China. However, little is unveiled regarding the allergenic components in Humulus japonicus pollen. Our study aimed to purify and identify the pathogenesis-related 1 (PR-1) protein from Humulus japonicus pollen, and to characterize the molecular and immunochemical properties of this novel allergen. METHODS: The natural PR-1 protein (named as Hum j PR-1) was purified from Humulus japonicus pollen extracts with a combined strategy of chromatography, and identified by mass spectrometry. The coding sequence of Hum j PR-1 was confirmed by cDNA cloning. The recombinant Hum j PR-1 was expressed and purified from Escherichia coli. The allergenicity was assessed by immunoblot, enzyme-linked immunosorbent assay (ELISA), inhibition ELISA, and basophil activation test using Humulus japonicus allergic patients' whole blood. The physicochemical properties and 3-dimensional structure of it were comprehensively characterized by in silico methods. RESULTS: The allergenicity analysis revealed that 76.6 % (23/30) of the Humulus japonicus pollen allergic patients displayed specific IgE recognition of the natural Hum j PR-1. The cDNA sequence of Hum j PR-1 had a 516-bp open reading frame encoding 171 amino acids. Physicochemical analysis indicated that Hum j PR-1 was a stable and relatively thermostable protein. Hum j PR-1 shared a similar 3-dimensional folding pattern with other homologous allergens, which was a unique αßα sandwich structure containing 4 α-helices and 6 antiparallel ß-sheets, encompassing 4 conserved CAP domain. CONCLUSION: The natural PR-1 was firstly purified and characterized as a major allergenic allergen in Humulus japonicus pollen. These findings would contribute to developing diagnostic and therapeutic strategies for Humulus japonicus pollinosis.

Biomedical Indicators of Patients with Non-Puerperal Mastitis: A Retrospective Study.

BACKGROUND: Non-puerperal mastitis (NPM), a recurrent chronic inflammation of non-lactating breast, often proves tremendous difficulty in treatment, and it may give rise to its complicated symptoms and unclear etiology. Furthermore, the clinical morbidity rate of NPM has been increasing in recent years. METHODS: Overall, 284 patients diagnosed with NPM were consecutively recruited as cases in this study, and patients with benign breast disease (n = 1128) were enrolled as control. The clinical, biomedical, and pathological indicators were analyzed. Univariate and multivariate logistic analysis were used to distinguish risks between NPM and benign breast mass patients. Furthermore, according to the pathological characteristics, the patients of NPM were classified into two subgroups: mammary duct ectasia (MDE) and granulomatous lobular mastitis (GLM). The differences of biomedical indicators between MDE and GLM groups were also analyzed. RESULTS: Compared with benign breast mass group, the level of high-density lipoprotein (HDL-C) significantly decreased, while lipoprotein(a) (Lp(a)) and blood glucose (GLU) both increased in NPM group. According to univariate and multivariate logistic analysis, the onset age and HDL-C were generally decreased, while Lp(a) and GLU were increased in NPM group. The onset age, HDL-C, Lp(a), and GLU were modeled to distinguish NPM and benign breast mass. Significant differences were also observed between MDE and GLM patients in biomedical indicators, such as lipoprotein(a) (Lp(a)), lactate dehydrogenase (LDH), creatine kinase (CK), total cholesterol (TC), and so on. CONCLUSIONS: Our results indicated for the first time that biomarkers were associated with NPM. The biomedical indicators involved in lipid metabolism might be important factors in the development and treatment of NPM. In addition, MDE and GLM are two diseases with different inflammatory states of NPM. These findings would be helpful for a better understanding of NPM and give us some insights to develop new diagnostic and therapeutic strategies.

Purification and characterization of enolase as a novel allergen in Platanus acerifolia pollen.

BACKGROUND: The pollen from Platanus acerifolia (P. acerifolia) is one of the main causes of allergic disorders. To date, only 4 allergens have been identified from this pollen. But previous studies showed that there still exist under-recognized allergens in it. The aim of this study was to comprehensively investigate the newly identified enolase (Pla a 6) as a novel allergen in the P. acerifolia pollen. METHODS: The natural (n) Pla a 6 was purified by combined chromatographic strategies. According to the identified internal peptides, the cDNA sequence encoding this allergen was matched from the mRNA-sequencing results of P. acerifolia pollen, which was further amplified and cloned. The recombinant (r) Pla a 6 was expressed and purified from E. coli. The allergenicity of this novel allergen was characterized by enzyme linked immunosorbent assay (ELISA), Western blot, inhibition ELISA, and basophil activation test (BAT). RESULTS: A novel allergen from P. acerifolia pollen, named as Pla a 6 was thoroughly studied, which contained an open reading frame of 1338 bp encoding 445 amino acids. The IgE-binding activity of nPla a 6 was initially proved by Western-blot, and a similar IgE-binding pattern to rPla a 6 was also exhibited. Moreover, the positivity for specific IgE against rPla a 6 was tested as 45.95% (17/37) by ELISA, and IgE binding to pollen extract could be inhibited up to 45.77% by 10 µg/ml of rPla a 6. The protein was also confirmed to activate patients' basophils. CONCLUSIONS: In this study, a novel allergen belonging to enolase family was comprehensively investigated and characterized through its natural and recombinant forms in P. acerifolia pollen. The study will contribute to the development of novel molecular-based diagnostic and therapeutic approaches for P. acerifolia pollen allergy.

Current Therapeutic Strategies for Metastatic Triple-Negative Breast Cancer: From Pharmacists' Perspective.

Triple-negative breast cancer (TNBC) is characterized by its high invasiveness, high metastasis and poor prognosis. More than one-third of patients with TNBC will present with recurrence or distant metastasis. Chemotherapy based on anthracyclines and taxanes is the standard treatment strategy for metastatic TNBC (mTNBC). Due to the lack of expression of progesterone receptor, estrogen receptor, and human epidermal growth factor receptor 2, therapies targeting these receptors are ineffective for mTNBC, thus special treatment strategies are required. In recent years, the development of new chemotherapy drugs, targeted drugs and immunotherapy drugs offers good prospects for the treatment of mTNBC. However, as these drugs are still in their infancy, several problems regarding the optimization and management of the clinical application of these new options should be considered. Pharmacists can play an important role in drug selection, drug therapy management, the management of adverse drug reactions and pharmacoeconomic evaluation. In this review, we summarized traditional treatment strategies, and discussed the efficacy and safety of novel agents approved in the last ten years and combination regimens for mTNBC, with the aim of providing management strategies for the clinical management of mTNBC from pharmacists' perspective.

Nano-silica particles synergistically IgE-mediated mast cell activation exacerbating allergic inflammation in mice.

Background: Allergic respiratory diseases have increased dramatically due to air pollution over the past few decades. However, studies are limited on the effects of inorganic components and particulate matter with different particle sizes in smog on allergic diseases, and the possible molecular mechanism of inducing allergies has not been thoroughly studied. Methods: Four common mineral elements with different particle sizes in smog particles were selected, including Al2O3, TiO2, Fe2O3, and SiO2. We studied the relationship and molecular mechanism of smog particle composition, particle size, and allergic reactions using mast cells, immunoglobulin E (IgE)-mediated passive cutaneous anaphylaxis (PCA) model, and an ovalbumin (OVA)-induced asthmatic mouse model in vitro and in vivo, combined with transmission electron microscopy, scanning transmission X-ray microscopy analysis, and transcriptome sequencing. Results: Only 20 nm SiO2 particles significantly increased ß-hexosaminidase release, based on dinitrophenol (DNP)-human serum albumin (HSA) stimulation, from IgE-sensitized mast cells, while other particles did not. Meanwhile, the PCA model showed that Evan's blue extravasation in mice was increased after treatment with nano-SiO2 particles. Nano-SiO2 particles exposure in the asthmatic mouse model caused an enhancement of allergic airway inflammation as manifested by OVA-specific serum IgE, airway hyperresponsiveness, lung inflammation injury, mucous cell metaplasia, cytokine expression, mast cell activation, and histamine secretion, which were significantly increased. Nano-SiO2 particles exposure did not affect the expression of FcϵRI or the ability of mast cells to bind IgE but synergistically activated mast cells by enhancing the mitogen-activated protein kinase (MAPK) signaling pathway, especially the phosphorylation levels of the extracellular signal-regulated kinase (ERK)1/2. The ERK inhibitors showed a significant inhibitory effect in reducing ß-hexosaminidase release. Conclusion: Our results indicated that nano-SiO2 particles stimulation might synergistically activate IgE-sensitized mast cells by enhancing the MAPK signaling pathway and that nano-SiO2 particles exposure could exacerbate allergic inflammation. Our experimental results provide useful information for preventing and treating allergic diseases.

The bacteria inside human cancer cells: Mainly as cancer promoters.

The roles of the microbiome in human beings have become clearer with the development of next-generation sequencing techniques. Several pieces of evidence showed strong correlations between the microbiome and human health and disease, such as metabolic disorders, infectious diseases, digestive system diseases, and cancers. Among these diverse microbiomes, the role of bacteria in human cancers, especially in cancer cells, has received extensive attention. Latest studies found that bacteria widely existed in cancers, mainly in cancer cells and immune cells. In this review, we summarize the latest advances in understanding the role of bacteria in human cancer cells. We also discuss how bacteria are transported into cancer cells and their physiological significance in cancer progression. Finally, we present the prospect of bacterial therapy in cancer treatment.

Colorimetric visualization of histamine secreted by basophils based on DSP-functionalized gold nanoparticles.

Histamine released by activated basophils has become an important biomarker and therapeutic target in the development of allergic diseases. To date, several gold nanoparticle (AuNP)-based nanosensors have been reported for histamine detection in foods. However, rapid, highly sensitive and direct detection of histamine in allergic diseases is still lacking due to the complexity of the physical environment. Herein, we developed a novel nanosensor for colorimetric visualization of histamine in activated basophils by simply coupling dithiobis(succinimidylpropionate) (DSP) on the surface of AuNPs (DSP-AuNPs). The DSP moiety serves as a linker and can react with the aliphatic amino group of histamine, and the imidazole ring of histamine can selectively bind with Au by means of p-p conjugation, thus inducing the aggregation of AuNPs. In this study, we experimentally proved that DSP-AuNPs showed good sensitivity and selectivity to histamine among various amino acids, including histidine. Additionally, this nanosensor displayed a rapid response to histamine with a linear range of 0.8-2.5 µM, and the limit of detection (LOD) was 0.014 µM, which is a relatively low LOD in comparison with those of other AuNP-based nanosensors. Finally, DSP-AuNPs are used, for the first time, to successfully detect endogenous histamine changes in activated basophils. Therefore, our work may provide a promising strategy to monitor histamine levels in the basophil activation test.

Biosimilar monoclonal antibodies in China: A patent review.

Biosimilars play an important role in reducing the burden on patients and increasing the market competition. Biosimilar monoclonal antibodies are currently one of the hotspots of research and development in China with policies support. With the continuous improvement of policies, the enthusiasm for the research and development of biosimilars has increased year by year. The policy requirements in different periods have different degrees of impact on the patent applications of pharmaceutical companies. This review introduces the biosimilar monoclonal antibodies market status and approval process in China, analyzes the patents in this field, and helps pharmaceutical companies protect their intellectual property rights.

Kinesin family member 23, regulated by FOXM1, promotes triple negative breast cancer progression via activating Wnt/ß-catenin pathway.

BACKGROUND: Triple negative breast cancer (TNBC) is highly malignant and has a worse prognosis, compared with other subtypes of breast cancer due to the absence of therapeutic targets. KIF23 plays a crucial role in the tumorigenesis and cancer progression. However, the role of KIF23 in development of TNBC and the underlying mechanism remain unknown. The study aimed to elucidate the biological function and regulatory mechanism of KIF23 in TNBC. METHODS: Quantitative real-time PCR and Western blot were used to determine the KIF23 expression in breast cancer tissues and cell lines. Then, functional experiments in vitro and in vivo were performed to investigate the effects of KIF23 on tumor growth and metastasis in TNBC. Chromatin immunoprecipitation assay was conducted to illustrate the potential regulatory mechanisms of KIF23 in TNBC. RESULTS: We found that KIF23 was significantly up-regulated and associated with poor prognosis in TNBC. KIF23 could promote TNBC proliferation, migration and invasion in vitro and in vivo. KIF23 could activate Wnt/ß-catenin pathway and promote EMT progression in TNBC. In addition, FOXM1, upregulated by WDR5 via H3K4me3 modification, directly bound to the promoter of KIF23 gene to promote its transcription and accelerated TNBC progression via Wnt/ß-catenin pathway. Both of small inhibitor of FOXM1 and WDR5 could inhibit TNBC progression. CONCLUSIONS: Our findings elucidate WDR5/FOXM1/KIF23/Wnt/ß-catenin axis is associated with TNBC progression and may provide a novel and promising therapeutic target for TNBC treatment.

A new cysteine protease allergen from Ambrosia trifida pollen: proforms and mature forms.

Giant ragweed (Ambrosia trifida) pollen is closely associated with respiratory allergy in late summer and autumn, and the prevalence of giant ragweed pollen allergy progressively increases. Compared with short ragweed (Ambrosia artemisiifolia), allergenic components from giant ragweed pollen are poorly investigated. To promote component-resolved diagnosis and treatment for giant ragweed pollen allergy, it becomes necessary to identify and characterize unknown allergens from giant ragweed pollen. In the present study, we identified and characterized a new cysteine-protease (CP) allergen from giant ragweed pollen, named as Amb t CP. The cloned Amb t CP gene encoded 387 amino acids. Recombinant Amb t CP (rAmb t CP) and natural Amb t CP (nAmb t CP) were purified by high-affinity Ni2+ resin and immunoaffinity chromatography respectively. During refolding, purified rAmb t CP could autocatalytically converted to its mature forms displaying a higher enzymatic activity. Moreover, the autocatalytic conversion of proforms to mature forms of nAmb t CP could cause their amount to change in giant ragweed pollen extracts. Then, the allergenicity of Amb t CP was characterized: 23 (33.8%) of 68 Chinese patients with ragweed pollen allergy showed positive IgE binding to nAmb t CP by enzyme-linked immunosorbent assay (ELISA); the result of subsequent ELISA showed that IgE-binding activity of proforms and mature forms of rAmb t CP was different, with positive rate of 39.1% (9/23) and 47.8% (11/23) respectively; Amb t CP showed IgE cross-reactivity with the CP components from short ragweed, Artemisia annua and Artemisia sieversiana pollen. Our findings will help to promote component-resolved diagnosis and treatment for giant ragweed pollen allergy, standardize allergen products and individualize allergen-specific immunotherapy.

Prediction of clinical efficacy of subcutaneous immunotherapy for Artemisia sieversiana pollen allergic rhinitis by serum metabolomics.

BACKGROUND/PURPOSE: Specific immunotherapy is the only effective etiological treatment for allergic rhinitis, but subcutaneous immunotherapy has a slow onset and poor compliance. Predicting the clinical efficacy of subcutaneous immunotherapy in advance can reduce unnecessary medical costs and resource waste. This study aimed to identify metabolites that could predict the efficacy of subcutaneous immunotherapy on seasonal allergic rhinitis by serum metabolomics. METHODS: Patients (n = 43) with Artemisia sieversiana pollen allergic rhinitis were enrolled and treated with subcutaneous immunotherapy for one year. Patients were divided into the ineffective group (n = 10) and effective group (n = 33) according to the therapeutic index. Serum samples were collected before treatment. Metabolomics was determined by liquid chromatography-mass spectrometry combined with gas chromatography-mass spectrometry and analyzed differential compounds and related metabolic pathways. RESULTS: A total of 129 differential metabolites (P < 0.05) were identified and 4 metabolic pathways, namely taurine and hypotaurine metabolism, pentose and glucuronate interconversions, pentose phosphate pathway, and alanine, aspartate, and glutamate metabolism, were involved. CONCLUSION: Some metabolites, such as hypotaurine, taurine, and l-alanine, have the potential to become predictive biomarkers for effective subcutaneous immunotherapy.

The development of a candidate of desensitization vaccines against Der f 2 nearly without IgE-binding activity.

Considering the important role of Der f 2 in house dust mites mediating allergic diseases and allergic adverse effects during allergen-specific immunotherapy (AIT), we intend to develop a candidate of desensitization vaccines against Der f 2 without allergenicity. According to the reported immunoglobulin E (IgE)-binding B and T cell epitopes of Der f 2, four candidates of desensitization vaccines against Der f 2 were developed. Recombinant wild-type Der f 2 (rWt-Der f 2) preserved conformational and linear IgE-binding B epitopes. rWt-Der f 2 linearized by reduction and alkylation reactions (rWt-Der f 2 (red/alk)) and recombinant modified-type Der f 2 (rMt-Der f 2) were developed via destroying conformational and linear IgE-binding B epitopes respectively. rMt-Der f 2 linearized by reduction and alkylation reactions (rMt-Der f 2 (red/alk)) was developed by destroying conformational and linear IgE-binding B epitopes. T cell epitopes of 4 candidates were preserved. The change of their IgE-binding activity was determined by enzyme linked immunosorbent assay (ELISA), western blot and inhibition ELISA. Compared with rWt-Der f 2, the IgE-binding activity of rWt-Der f 2 (red/alk), rMt-Der f 2 and rMt-Der f 2 (red/alk) all decreased, which was consistent with the result of western blot. The IgE-binding activity of rMt-Der f 2 and rMt-Der f 2 (red/alk) was not significantly different (P = 0.0863 > 0.05), which was comparable to that of their corresponding negative controls (P = 0.3488 and 0.4459, both > 0.05). The result of inhibition ELISA also showed that their IgE-binding activity decreased, and rMt-Der f 2 (red/alk) was the lowest. Conclusively, we developed the candidate of desensitization vaccines against Der f 2, rMt-Der f 2 or rMt-Der f 2 (red/alk), nearly without allergenicity, which would potentially prevent HDM allergic patients from allergic adverse effects caused by AIT.

CAR-T Cell Therapy for Breast Cancer: From Basic Research to Clinical Application.

Breast cancer rises as the most commonly diagnosed cancer in 2020. Among women, breast cancer ranks first in both cancer incidence rate and mortality. Treatment resistance developed from the current clinical therapies limits the efficacy of therapeutic outcomes, thus new treatment approaches are urgently needed. Chimeric antigen receptor (CAR) T cell therapy is a type of immunotherapy developed from adoptive T cell transfer, which typically uses patients' own immune cells to combat cancer. CAR-T cells are armed with specific antibodies to recognize antigens in self-tumor cells thus eliciting cytotoxic effects. In recent years, CAR-T cell therapy has achieved remarkable successes in treating hematologic malignancies; however, the therapeutic effects in solid tumors are not up to expectations including breast cancer. This review aims to discuss the development of CAR-T cell therapy in breast cancer from preclinical studies to ongoing clinical trials. Specifically, we summarize tumor-associated antigens in breast cancer, ongoing clinical trials, obstacles interfering with the therapeutic effects of CAR-T cell therapy, and discuss potential strategies to improve treatment efficacy. Overall, we hope our review provides a landscape view of recent progress for CAR-T cell therapy in breast cancer and ignites interest for further research directions.