It has been more than three decades since the first monoclonal antibody was approved by the United States Food and Drug Administration (US FDA) in 1986, and during this time, antibody engineering has dramatically evolved. Current antibody drugs have increasingly fewer adverse effects due to their high specificity. As a result, therapeutic antibodies have become the predominant class of new drugs developed in recent years. Over the past five years, antibodies have become the best-selling drugs in the pharmaceutical market, and in 2018, eight of the top ten bestselling drugs worldwide were biologics. The global therapeutic monoclonal antibody market was valued at approximately US$115.2 billion in 2018 and is expected to generate revenue of $150 billion by the end of 2019 and $300 billion by 2025. Thus, the market for therapeutic antibody drugs has experienced explosive growth as new drugs have been approved for treating various human diseases, including many cancers, autoimmune, metabolic and infectious diseases. As of December 2019, 79 therapeutic mAbs have been approved by the US FDA, but there is still significant growth potential. This review summarizes the latest market trends and outlines the preeminent antibody engineering technologies used in the development of therapeutic antibody drugs, such as humanization of monoclonal antibodies, phage display, the human antibody mouse, single B cell antibody technology, and affinity maturation. Finally, future applications and perspectives are also discussed.

Oncogenic activation of the Wnt/b-catenin signaling pathway is common in hepatocellular carcinoma (HCC). Our recent studies have demonstrated that SRY (sex determining region Y)-box 1 (SOX1) and secreted frizzled-related proteins are concomitantly promoter-hypermethylated, and this might lead to abnormal activation of the Wnt signaling pathway in HCC. SOX1 encodes a transcription factor involved in the regulation of embryonic development and cell fate determination. However, the expression and functional role of SOX1 in HCC remains unclear. In this study, we confirmed via quantitative methylation-specific polymerase chain reaction that SOX1 was frequently downregulated through promoter hypermethylation in HCC cells and tissues. Overexpression of SOX1 by a constitutive or inducible approach could suppress cell proliferation, colony formation, and invasion ability in HCC cell lines, as well as tumor growth in nonobese diabetic/severe combined immunodeficiency mice. Conversely, knockdown of SOX1 by withdrawal of doxycycline could partially restore cell proliferation and colony formation in HCC cells. We used a T cell factor (TCF)-responsive luciferase reporter assay and western blot analysis to prove that SOX1 could regulate TCF-responsive transcriptional activity and inhibit the expression of Wnt downstream genes. Furthermore, we used glutathione S-transferase pull-down, co-immunoprecipitation, and confocal microscopy to demonstrate that SOX1 could interact with b-catenin but not with the b-catenin/TCF complex. Moreover, restoration of the expression of SOX1 induces significant cellular senescence in Hep3B cells. Conclusion: Our data show that a developmental gene, SOX1, may function as a tumor suppressor by interfering with Wnt/b-catenin signaling in the development of HCC. (HEPATOLOGY 2012;56:2277-2287 T he incidence and mortality of hepatocellular carcinoma (HCC) have been increasing rapidly worldwide in recent decades.1 The risk factors associated with hepatocarcinogenesis are numerous and include chronic hepatitis B or C viral infection, alcohol, aflatoxin B1, and others. However, the molecular mechanisms involved in the development of HCC remain unclear. Recent studies have demonstrated that inactivation of tumor suppressor genes (TSGs) through promoter hypermethylation plays an essential role in carcinogenesis.2,3 Furthermore, methylation profiles have been used as potential biomarkers for early diagnosis, prognostic prediction, and screening in HCC. 4 Therefore, exploring the molecular mechanisms of the inactivation of TSGs involved in HCC development could improve the treatment of HCC in the future.

Anticancer therapies are often compromised by nonspecific effects and challenged by tumour environments’ inherent physicochemical and biological characteristics. Often, therapeutic effect can be increased by addressing multiple parameters simultaneously. Here we report on exploiting extravasation due to inherent vascular leakiness for the delivery of a pH-sensitive polymer carrier. Tumours’ acidic microenvironment instigates a charge reversal that promotes cellular internalization where endosomes destabilize and gene delivery is achieved. We assess our carrier with an aggressive non-small cell lung carcinoma (NSCLC) in vivo model and achieve >30% transfection efficiency via systemic delivery. Rejuvenation of the p53 apoptotic pathway as well as expression of KillerRed protein for sensitization in photodynamic therapy (PDT) is accomplished. A single administration greatly suppresses tumour growth and extends median animal survival from 28 days in control subjects to 68 days. The carrier has capacity for multiple payloads for greater therapeutic response where inter-individual variability can compromise efficacy.

Solid tumors characteristically display higher levels of lactate production due to anaerobic metabolism of glucose. Meanwhile, the U.S. Food and Drug Administration (FDA) has approved virotherapy for use in cancer treatment; however systemic administration remains as a particular challenge. Here we report exploitation of tumor lactate production in designing a hypoxia-responsive carrier, self-assembled from hyaluronic acid (HA) conjugated with 6-(2-nitroimidazole)­hexylamine, for localized release of recombinant adeno-associated virus serotype 2 (AAV2). The carrier is loaded with lactate oxidase (LOX) and is permeable to small molecules such as the lactate that accumulates in the tumor. Subsequently, LOX oxidizes the lactate to pyruvate inside the carrier, accompanied by internal lowering of oxygen partial pressure. Bioreduction of the 2-nitroimidazole of the HA conjugated with 6-(2-nitroimidazole)­hexylamine converts it into a hydrophilic moiety and electrostatically dissociates the carrier and virus. Efficacious and specific delivery was proven by transduction of a photosensitive protein (KillerRed), enabling significant limitation in tumor growth in vivo with photodynamic therapy. An approximate 2.44-fold reduction in tumor weight was achieved after a 2-week course, compared with control groups. Furthermore, conjugation of the AAV2 with iron oxide nanoparticles (“magnetized” AAV2) facilitated magnetic resonance imaging tracking of the virus in vivo. Taken together, the solid tumor microenvironment promotes bioreduction of the lactate-responsive carrier, providing rapid and specific delivery of AAV2 for light-triggered virotherapy via systemic administration.

Autoimmune thyroid disease (AITD), including Graves disease (GD) and Hashimoto disease (HD), is an organ-specific autoimmune disease with a strong genetic component. Although the cytotoxic T-lymphocyte-associated protein 4 (CTLA4) polymorphism has been reported to be associated with AITD in adults, few studies have focused on children. The aim of our study was to investigate whether the CTLA4 polymorphisms, including -318C/T (rs5742909), +49A/G (rs231775), and CT60 (rs3087243), were associated with GD and HD in Han Chinese adults and children. We studied 289 adult GD, 265 pediatric GD, 229 pediatric HD patients, and 1058 healthy controls and then compared genotype, allele, carrier, and haplotype frequencies between patients and controls. We found that CTLA4 SNPs +49A/G and CT60 were associated with GD in adults and children. Allele G of +49A/G was significantly associated with GD in adults (odds ratio [OR], 1.50; 95% confidence interval [CI], 1.21–1.84; corrected P value [Pc] < 0.001) and children (OR, 1.42; 95% CI, 1.15–1.77; Pc = 0.002). Allele G of CT60 also significantly increased risk of GD in adults (OR, 1.63; 95% CI, 1.27–2.09; Pc < 0.001) and GD in children (OR, 1.58; 95% CI, 1.22–2.04; Pc < 0.001). Significant linkage disequilibrium was found between +49A/G and CT60 in GD and control subjects (D’ = 0.92). Our results showed that CTLA4 was associated with both GD and HD and played an equivalent role in both adult and pediatric GD in Han Chinese population.

ENO1 (α-enolase) expression is significantly correlated with reduced survival and poor prognosis in many cancer types, including lung cancer. However, the function of ENO1 in carcinogenesis remains elusive. In this study, we found that high expression of ENO1 is present in metastatic lung cancer cell lines and malignant tumors and is associated with poor overall survival of patients with lung cancer. Knockdown of ENO1 decreased cancer cell proliferation and invasiveness, whereas overexpression of ENO1 enhanced these processes. Moreover, ENO1 expression promoted tumor growth in orthotopic models and enhanced lung tumor metastasis in tail-vein injection models. These effects were mediated by upregulation of mesenchymal markers N-cadherin and vimentin and the epithelial-to-mesenchymal transition regulator SLUG, along with concurrent downregulation of E-cadherin. Mechanistically, ENO1 interacted with hepatocyte growth factor receptor (HGFR) and activated HGFR and Wnt signaling via increased phosphorylation of HGFR and the Wnt coreceptor LRP5/6. Activation of these signaling axes decreased GSK3β activity via Src–PI3K–AKT signaling and inactivation of the β-catenin destruction complex to ultimately upregulate SLUG and β-catenin. In addition, we generated a chimeric anti-ENO1 mAb (chENO1-22) that can decrease cancer cell proliferation and invasion. chENO1-22 attenuated cancer cell invasion by inhibiting ENO1-mediated GSK3β inactivation to promote SLUG protein ubiquitination and degradation. Moreover, chENO1-22 prevented lung tumor metastasis and prolonged survival in animal models. Taken together, these findings illuminate the molecular mechanisms underlying the function of ENO1 in lung cancer metastasis and support the therapeutic potential of a novel antibody targeting ENO1 for treating lung cancer. Significance: This study shows that ENO1 promotes lung cancer metastasis via HGFR and WNT signaling and introduces a novel anti-ENO1 antibody for potential therapeutic use in lung cancer.

We have reported previously that LIM homeobox transcription factor 1α (LMX1A) is hypermethylated and functions as a metastasis suppressor in cervical cancer cells. However, the regulation of LMX1A in carcinogenesis has not been reported. We aim to clarify whether specificity protein 1 (Sp1) and enhancer of zeste homolog 2 (EZH2) are involved in the regulation of LMX1A in cervical cancer. First we characterized the LMX1A promoter and used overexpression, knockdown, and reporter assays to show that Sp1 increased LMX1A promoter activity. Next, we used site-directed mutagenesis and electrophoresis mobility shift assays (EMSAs) to demonstrate that Sp1-binding sites were important for Sp1-mediated activation of the LMX1A promoter. Chromatin immunoprecipitation data demonstrated that Sp1 could bind directly to the LMX1A promoter and activate endogenous LMX1A expression in cells pretreated with 5-aza-2'-deoxycytidine (5-aza-dC). Knockdown of EZH2 decreased H3K27me3 histone modification but was insufficient to restore LMX1A expression. To explore the effect of EZH2 on the endogenous LMX1A promoter, we treated EZH2-knockdown cells with 5-aza-dC and trichostatin A (TSA) and then depleted the cells of drugs for 3days. H3K14ac was enriched at the LMX1A promoter in EZH2-knockdown cells and LMX1A mRNA was still expressed. Taken together, these data imply that Sp1 may activate LMX1A expression upon oncogenic stress during cervical cancer development. Moreover, suppression of EZH2 may delay resilencing of LMX1A after the removal of 5-aza-dC and TSA.

Background: Graves disease (GD) is the most common cause of thyrotoxicosis in children and adolescents, accounting for 15% of all thyroid diseases during childhood. Anti-thyroid drugs (ATD) are recommended as the first-line treatment in children and adolescents. However, the remission rate is lower in children than in adults, and the optimal treatment duration and favorable factors associated with remission remain unknown. We aimed to investigate long-term outcomes of pediatric GD patients receiving ATD. Methods: We retrospectively reviewed medical charts of 396 GD subjects from 1985 to 2017 at MacKay Children's Hospital. Ninetyesix patients were excluded from the analyses, including 71 patients followed for less than one year, 6 patients who received radioactive therapy and 19 patients who received surgery. The remaining 300 patients initially treated with ATD and