Regulating Tumorigenicity and Cancer Metastasis through TRKA Signaling.

Tropomyosin receptor kinase (TRK) A, TRKA, is a specific binding receptor of nerve growth factor (NGF), which plays an essential role in the occurrence and progression of human cancers. TRKA overexpression has been proven to be a powerful carcinogenic driver and has been verified in many tumors. The TRKA receptor kinase domain is over-activated in an NGF-dependent manner, accompanied by activation of downstream signal pathways, such as RAS-MAPK, PI3K-AKT, JAK2-STAT3 pathway, PLC γ pathway, and Hippo pathway, which participate in tumor cell proliferation, invasion, epithelial-mesenchymal transition (EMT), perineural invasion (PNI), drug resistance, and cancer pain. In addition, chimeric oncogenes produced by the fusion of NTRK1 and other genes are also the direct cause of tumorigenesis and cancer development. The newly developed TRK inhibitors can improve symptoms and tumor regression in cancer patients with overexpression of TRKA or NTRK1 fusion gene. With the emergence of drug resistance, next generation of TRK inhibitors can still maintain strong clinical efficacy in the case of TRK kinase domain mutations, and these inhibitors are in clinical trials. This review summarizes the characteristics and research progress of TRKA, focusing on the regulatory role of the TRKA signal pathway in different tumors. In addition, we have summarized the clinical significance of TRKA and the TRK inhibitors. This review may provide a new reference for the study of the mechanism of TRKA in different tumors, and also provide a new perspective for the in-depth understanding of the role of TRKA as a biomarker and therapeutic target in human cancer.

Long bone shaft metastasis: a comparative study between cement filling and intercalary prosthesis.

BACKGROUND: Metastatic bone lesions in the extremities can cause severe pain and pathological fractures, significantly affecting patients' quality of life. Timely intervention and effective management of long bone metastases can positively influence patient outcomes, including survival rates and subsequent treatment options. OBJECTIVE: The objective of this study is to compare the efficacy and associated complications of two surgical reconstruction techniques and propose a more effective limb reconstruction approach for long bone metastases. METHODS: A retrospective study was conducted on 28 patients with complete clinical data who underwent a surgical procedure for long bone metastases of the extremities in our department between January 2017 and June 2022. The patients were divided into two groups based on their surgical methods. In group 1, the affected bones were curetted and filled with cement, then secured with plates or intramedullary nails. In group 2, the affected bone segments were completely removed and replaced with custom intercalary prostheses. Various factors, including general patient information, surgical details, surgical effectiveness, and common complications, were compared and analyzed. RESULTS: There were no significant differences in general patient information between the two groups, including age, gender, surgical site, and primary tumor type. The operative times were 115.37 min for group 1 and 108.90 min for group 2, respectively (p > 0.05). However, intraoperative blood loss differed significantly between the groups, with 769 ml in group 1 and 521 ml in group 2 (p < 0.05). The postoperative MSTS scores were 91% for group 1 and 92% for group 2 (p > 0.05). Postoperative complications included two cases of internal fixation failure and three cases of tumor recurrence in group 1, resulting in a 33% incidence rate, while group 2 experienced a 15% incidence rate with two cases of internal fixation failure. CONCLUSION: The results of this study suggest that both surgical techniques are effective for the treatment of long bone metastases of the extremities. However, the custom intercalary prostheses technique in group 2 showed a lower incidence of complications and less intraoperative blood loss. Therefore, it may be a more effective limb reconstruction approach for long bone metastases. Further studies with larger sample sizes are needed to confirm these findings.

DLX2 promotes osteosarcoma epithelial-mesenchymal transition and doxorubicin resistance by enhancing HOXC8-CDH2 axis.

Metastasis and doxorubicin resistance are challenges in the clinical diagnosis and treatment of osteosarcoma, the mechanisms underlying these phenomena remain unclear. In this study, we found that DLX2 is highly expressed in metastatic osteosarcoma and is closely related to clinical prognosis. Knockdown of DLX2 inhibited tumor proliferation and migration in vitro and inhibited tumor growth in vivo. Mechanistically, we found that DLX2 enhanced the repression of CDH2 transcription by binding to HOXC8, thereby promoting the epithelial-mesenchymal transition in osteosarcoma cells. Through subsequent exploration, we found that targeting DLX2/HOXC8 signaling significantly restores the sensitivity of osteosarcoma cells to doxorubicin. In conclusion, our findings demonstrate that DLX2 may enhance the transcriptional regulation of CDH2 through interacting with HOXC8, which in turn promotes epithelial-mesenchymal transition and doxorubicin resistance in osteosarcoma. These findings hold great potential for clinical application and may guide the development of novel targeted therapies for osteosarcoma.

Experience in utilizing a novel 3D digital model with CT and MRI fusion data in sarcoma evaluation and surgical planning.

OBJECTIVE: To assess sarcoma margins with more accuracy and aid surgical planning, we constructed three-dimensional (3D) digital models with computed tomography(CT) and magnetic resonance imaging (MRI) image fusion data and validated the preciseness of the models by comparing them with 3D models constructed with CT only data. MATERIALS AND METHODS: We retrospectively reviewed a consecutive set of patients treated in our center who were preoperatively evaluated with the fusion image model. Models based on fusion images or CT-only data were constructed. Volumes of both tumors were calculated and the tumors were overlapped to see the location of differences between the two models. RESULTS: A consecutive 12 cases (4 male vs. 8 female) were included in this study. Most of the tumors were located in the pelvic bone or spine. The volume of the two tumor models was different and the differences were mainly in the peripheral region of the tumor. CONCLUSION: CT and MRI fusion image 3D models are more accurate than models with CT-only data and can be very helpful in preoperative planning of sarcoma patients.

Taurine Protects Retinal Cells and Improves Synaptic Connections in Early Diabetic Rats.

Purpose: Taurine has long been thought to be involved in retinal protection from retinal degenerative diseases, but the underlying molecular mechanisms remain unclear. Retinal neurodegeneration is an early event in the pathogenesis of diabetic retinopathy (DR) that precedes and participates in the microcirculatory abnormalities that occur in DR. Our objective was to investigate the role and mechanisms of taurine in early diabetic retinas.Methods: Eight-week-old STZ-induced diabetic rats and control animals were randomly assigned to receive taurine or vehicle by intraperitoneal injection or by intragastric administration. The retinal function and retinal cell counts were evaluated using an electroretinography (ERG) and immunofluorescence microscopy. Plasma amino acids were measured by ion-exchange chromatography (IEC). The expression levels of retinal taurine transporter (Tau-T), mitochondria-dependent apoptosis-associated genes and reactive gliosis markers were studied by western blotting and immunofluorescence. Pre- and post-synaptic markers (PSD95 and mGluR6) in outer plexiform layer (OPL), and the bipolar cell marker protein kinase C alpha (PKCα) were localized by immunofluorescence. Levels of PSD95 and mGluR6 were determined by quantitative western blot.Results: Taurine significantly prevented the reduction of photopic b-wave amplitude and retinal cone cells and ganglion cells loss and maintained the Bcl-2/Bax ratio balance in diabetic rats. Taurine also prevented the upregulation of glial fibrillary acidic protein (GFAP) and reduced retinal reactive gliosis. Taurine reduced plasma glutamate and tyrosine levels, which were elevated in diabetic rats. Moreover, mGluR6 levels reduction detected by western blot and immunofluorescence in diabetic retinas was inhibited and the displacement of mGluR6 in OPL into the inner nuclear layer (INL) detected by immunofluorescence was reduced by Taurine treatment.Conclusion: Taurine may protect retinal cells from diabetic attacks by activating Tau-T, reducing retinal reactive gliosis, improving retinal synaptic connections and decreasing retinal cell apoptosis. Thus, taurine treatment may be a novel approach for early DR.

Tumor Suppression of Ras GTPase-Activating Protein RASA5 through Antagonizing Ras Signaling Perturbation in Carcinomas.

Aberrant RAS signaling activation is common in cancers with even few Ras mutations, indicating alternative dysregulation other than genetic mutations. We identified a Ras GTPase-activating gene RASA5/SYNGAP1, at the common 6p21.3 deletion, methylated/downregulated in multiple carcinomas and different from other RASA family members (RASA1-RASA4), indicating its special functions in tumorigenesis. RASA5 mutations are rare, unlike other RASA members, whereas its promoter CpG methylation is frequent in multiple cancer cell lines and primary carcinomas and associated with patient's poor survival. RASA5 expression inhibited tumor cell migration/invasion and growth in mouse model, functioning as a tumor suppressor. RASA5 suppressed RAS signaling, depending on its Ras GTPase-activating protein catalytic activity, which could be counteracted by oncogenic HRas Q61L mutant. RASA5 knockdown enhanced Ras signaling to promote tumor cell growth. RASA5 also inhibited epithelial-mesenchymal transition (EMT) through regulating actin reorganization. Thus, epigenetic inactivation of RASA5 contributing to hyperactive RAS signaling is involved in Ras-driven human oncogenesis.

The new 6q27 tumor suppressor DACT2, frequently silenced by CpG methylation, sensitizes nasopharyngeal cancer cells to paclitaxel and 5-FU toxicity via ß-catenin/Cdc25c signaling and G2/M arrest.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is prevalent in South China, including Hong Kong and Southeast Asia, constantly associated with Epstein-Barr virus (EBV) infection. Epigenetic etiology attributed to EBV plays a critical role in NPC pathogenesis. Through previous CpG methylome study, we identified Disheveled-associated binding antagonist of beta-catenin 2 (DACT2) as a methylated target in NPC. Although DACT2 was shown to regulate Wnt signaling in some carcinomas, its functions in NPC pathogenesis remain unclear. METHODS: RT-PCR, qPCR, MSP, and BGS were applied to measure expression levels and promoter methylation of DACT2 in NPC. Transwell, flow cytometric analysis, colony formation, and BrdU-ELISA assay were used to assess different biological functions affected by DACT2. Immunofluorescence, Western blot, and dual-luciferase reporter assay were used to explore the mechanisms of DACT2 functions. Chemosensitivity assay was used to measure the impact of DACT2 on chemotherapy drugs. RESULTS: We found that DACT2 is readily expressed in multiple normal adult tissues including upper respiratory tissues. However, it is frequently downregulated in NPC and correlated with promoter methylation. DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine restored its expression in NPC cells. DACT2 methylation was further detected in 29/32 (91%) NPC tumors but not in any (0/8) normal nasopharyngeal tissue samples. Ectopic expression of DACT2 in NPC cells suppressed their proliferation, migration, and invasion through downregulating matrix metalloproteinases. DACT2 expression also induced G2/M arrest in NPC cells through directly suppressing ß-catenin/Cdc25c signaling, which sensitized NPC cells to paclitaxel and 5-FU, but not cisplatin. CONCLUSION: Our results demonstrate that DACT2 is frequently inactivated epigenetically by CpG methylation in NPC, while it inhibits NPC cell proliferation and metastasis via suppressing ß-catenin/Cdc25c signaling. Our study suggests that DACT2 promoter methylation is a potential epigenetic biomarker for the detection and chemotherapy guidance of NPC.

FXR1 regulates transcription and is required for growth of human cancer cells with TP53/FXR2 homozygous deletion.

Tumor suppressor p53 prevents cell transformation by inducing apoptosis and other responses. Homozygous TP53 deletion occurs in various types of human cancers for which no therapeutic strategies have yet been reported. TCGA database analysis shows that the TP53 homozygous deletion locus mostly exhibits co-deletion of the neighboring gene FXR2, which belongs to the Fragile X gene family. Here, we demonstrate that inhibition of the remaining family member FXR1 selectively blocks cell proliferation in human cancer cells containing homozygous deletion of both TP53 and FXR2 in a collateral lethality manner. Mechanistically, in addition to its RNA-binding function, FXR1 recruits transcription factor STAT1 or STAT3 to gene promoters at the chromatin interface and regulates transcription thus, at least partially, mediating cell proliferation. Our study anticipates that inhibition of FXR1 is a potential therapeutic approach to targeting human cancers harboring TP53 homozygous deletion.

DACT2 silencing by promoter CpG methylation disrupts its regulation of epithelial-to-mesenchymal transition and cytoskeleton reorganization in breast cancer cells.

Wnt signaling plays an important role in breast carcinogenesis. DAPPER2 (DACT2) functions as an inhibitor of canonical Wnt signaling and plays distinct roles in different cell contexts, with its role in breast tumorigenesis unclear. We investigated DACT2 expression in breast cancer cell lines and primary tumors, as well as its functions and molecular mechanisms. Results showed that DACT2 expression was silenced in 9/9 of cell lines. Promoter CpG methylation of DACT2 was detected in 89% (8/9) of cell lines, as well as in 73% (107/147) of primary tumors, but only in 20% (1/5) of surgical margin tissues and in none of normal breast tissues. Demethylation of BT549 and T47D cell lines with 5-aza-2'-deoxycytidine restored DACT2 expression along with promoter demethylation, suggesting that its downregulation in breast cancer is dependent on promoter methylation. Furthermore, ectopic expression of DACT2 induced breast cell apoptosis in vitro, and further inhibited breast tumor cell proliferation, migration and EMT, through antagonizing Wnt/ß-catenin and Akt/GSK-3 signaling. Thus, these results demonstrate that DACT2 functions as a tumor suppressor for breast cancer but was frequently disrupted epigenetically in this cancer.

Experimental Evaluation of Three Designs of Electrodynamic Flexural Transducers.

Three designs for electrodynamic flexural transducers (EDFT) for air-coupled ultrasonics are presented and compared. An all-metal housing was used for robustness, which makes the designs more suitable for industrial applications. The housing is designed such that there is a thin metal plate at the front, with a fundamental flexural vibration mode at ∼50 kHz. By using a flexural resonance mode, good coupling to the load medium was achieved without the use of matching layers. The front radiating plate is actuated electrodynamically by a spiral coil inside the transducer, which produces an induced magnetic field when an AC current is applied to it. The transducers operate without the use of piezoelectric materials, which can simplify manufacturing and prolong the lifetime of the transducers, as well as open up possibilities for high-temperature applications. The results show that different designs perform best for the generation and reception of ultrasound. All three designs produced large acoustic pressure outputs, with a recorded sound pressure level (SPL) above 120 dB at a 40 cm distance from the highest output transducer. The sensitivity of the transducers was low, however, with single shot signal-to-noise ratio ( SNR ) ≃ 15 dB in transmit-receive mode, with transmitter and receiver 40 cm apart.

Characterization of the nasopharyngeal carcinoma methylome identifies aberrant disruption of key signaling pathways and methylated tumor suppressor genes.

AIMS: Nasopharyngeal carcinoma (NPC) is a common tumor consistently associated with Epstein-Barr virus infection and prevalent in South China, including Hong Kong, and southeast Asia. Current genomic sequencing studies found only rare mutations in NPC, indicating its critical epigenetic etiology, while no epigenome exists for NPC as yet. MATERIALS & METHODS: We profiled the methylomes of NPC cell lines and primary tumors, together with normal nasopharyngeal epithelial cells, using methylated DNA immunoprecipitation (MeDIP). RESULTS: We observed extensive, genome-wide methylation of cellular genes. Epigenetic disruption of Wnt, MAPK, TGF-ß and Hedgehog signaling pathways was detected. Methylation of Wnt signaling regulators (SFRP1, 2, 4 and 5, DACT2, DKK2 and DKK3) was frequently detected in tumor and nasal swab samples from NPC patients. Functional studies showed that these genes are bona fide tumor-suppressor genes for NPC. CONCLUSION: The NPC methylome shows a special high-degree CpG methylation epigenotype, similar to the Epstein-Barr virus-infected gastric cancer, indicating a critical epigenetic etiology for NPC pathogenesis.

Exendin-4 protects retinal cells from early diabetes in Goto-Kakizaki rats by increasing the Bcl-2/Bax and Bcl-xL/Bax ratios and reducing reactive gliosis.

PURPOSE: Exendin-4 (E4), a long-acting agonist of the hormone glucagon-like peptide 1 receptor (GLP-1R), is administered to treat type II diabetes in the clinical setting and also shows a neuroprotective effect. Our previous studies demonstrated its protective effect in early experimental diabetic retinopathy (DR), but the molecular and cellular mechanisms are largely unknown. This study aimed to investigate the protective mechanism of a GLP-1R agonist E4 against early DR in Goto-Kakizaki (GK) rats. METHODS: Diabetic GK rats and control animals were randomly assigned to receive E4 or vehicle by intravitreal injection. The retinal function and retinal cell counts were evaluated using an electroretinogram and light microscopy. The expressions of retinal GLP-1R, mitochondria-dependent apoptosis-associated genes, reactive gliosis markers, and endoplasmic reticulum stress-related pathway genes were studied by western blotting and immunohistochemistry in vivo and in vitro. RESULTS: E4 significantly prevented the reduction of the b-wave and oscillatory potential amplitudes and retinal cell loss and maintained the Bcl-2/Bax and Bcl-xL/Bax ratio balances in GK rats. It also downregulated the expression of glial fibrillary acidic protein and reduced retinal reactive gliosis. Similar results were found in primary rat Müller cells under high glucose culture in vitro. CONCLUSIONS: E4 may protect retinal cells from diabetic attacks by activating GLP-1R, decreasing retinal cell apoptosis, and reducing retinal reactive gliosis. Thus, E4 treatment may be a novel approach for early DR.

Exendin-4 alleviates retinal vascular leakage by protecting the blood-retinal barrier and reducing retinal vascular permeability in diabetic Goto-Kakizaki rats.

The breakdown of the inner endothelial blood-retinal barrier (BRB) and subsequent retinal vascular leakage are the main causes of vision loss due to diabetic retinopathy (DR). Exendin-4 (E4) is a long-acting agonist of the glucagon-like peptide 1 hormone receptor (GLP-1R) that is widely used in clinics and has shown a neuroprotective effect. Our previous studies demonstrated the protective effect of E4 in early experimental DR; however, the molecular and cellular mechanisms that mediate this protective effect are not fully known. The BRB plays a key role in DR. We speculated that E4 may exert its protective effects on the BRB. To test this hypothesis, E4 (0.1 µg/2 µL/eye) or vehicle were intravitreally injected into diabetic Goto-Kakizaki(GK) rats and control animals. The results revealed that E4 significantly inhibited the reductions in electroretinogram (ERG) amplitudes in the GK rats, particularly in the b-wave and oscillatory potentials (OPs). E4 upregulated retinal GLP-1R expression and downregulated the expressions of placental growth factor (PLGF) and vascular endothelial growth factor (VEGF) via the ERK and AKT/PKB pathways. Decreases in tight junction protein (i.e., claudin-5 and occludin) expression and increases in Evans blue permeation (EBP) were inhibited by E4. Similar results were also found in primary rat Müller cells in high glucose concentration cultures in vitro. We conclude that E4 may protect the BRB from diabetic insults by decreasing PLGF and ICAM-1 expression and maintaining the integrity of the BRB. Thus, E4 treatment may be an effective therapeutic approach for DR.

Taurine: the comeback of a neutraceutical in the prevention of retinal degenerations.

Taurine is the most abundant amino acid in the retina. In the 1970s, it was thought to be involved in retinal diseases with photoreceptor degeneration, because cats on a taurine-free diet presented photoreceptor loss. However, with the exception of its introduction into baby milk and parenteral nutrition, taurine has not yet been incorporated into any commercial treatment with the aim of slowing photoreceptor degeneration. Our recent discovery that taurine depletion is involved in the retinal toxicity of the antiepileptic drug vigabatrin has returned taurine to the limelight in the field of neuroprotection. However, although the retinal toxicity of vigabatrin principally involves a deleterious effect on photoreceptors, retinal ganglion cells (RGCs) are also affected. These findings led us to investigate the possible role of taurine depletion in retinal diseases with RGC degeneration, such as glaucoma and diabetic retinopathy. The major antioxidant properties of taurine may influence disease processes. In addition, the efficacy of taurine is dependent on its uptake into retinal cells, microvascular endothelial cells and the retinal pigment epithelium. Disturbances of retinal vascular perfusion in these retinal diseases may therefore affect the retinal uptake of taurine, resulting in local depletion. The low plasma taurine concentrations observed in diabetic patients may further enhance such local decreases in taurine concentration. We here review the evidence for a role of taurine in retinal ganglion cell survival and studies suggesting that this compound may be involved in the pathophysiology of glaucoma or diabetic retinopathy. Along with other antioxidant molecules, taurine should therefore be seriously reconsidered as a potential treatment for such retinal diseases.

The wave-field from an array of periodic emitters driven simultaneously by a broadband pulse.

The use of phased array methods are commonplace in ultrasonic applications, where controlling the variation of the phase between the narrowband emitters in an array facilitates beam steering and focusing of ultrasonic waves. An approach is presented here whereby emitters of alternating polarity arranged in a one-dimensional array are pulsed simultaneously, and have sufficiently wide, controlled bandwidth to emit a two-dimensional wave. This pulsed approach provides a rapid means of simultaneously covering a region of space with a wave-front, whereby any wave that scatters or reflects off a body to a detector will have a distinct arrival time and frequency. This is a general wave phenomenon with a potential application in radar, sonar, and ultrasound. The key result is that one can obtain a smooth, continuous wave-front emitted from the array, over a large solid angle, whose frequency varies as a function of angle to the array. Analytic and finite element models created to describe this phenomenon have been validated with experimental results using ultrasonic waves in metal samples.

FEZF2, a novel 3p14 tumor suppressor gene, represses oncogene EZH2 and MDM2 expression and is frequently methylated in nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is an Epstein-Barr virus-associated tumor prevalent in southern China and southeast Asia, with the 3p14-p12 locus reported as a critical tumor suppressor gene (TSG) region during its pathogenesis. We identified a novel 3p14.2 TSG, FEZF2 (FEZ family zinc finger 2), for NPC. FEZF2 is readily expressed in normal tissues including upper respiratory epithelium, testis, brain and ovary tissues, as well as in immortalized nasopharyngeal epithelial cell line NP69, but it is completely silenced in NPC cell lines due to CpG methylation of its promoter, although no homozygous deletion of FEZF2 was detected. 5-Aza-2'-deoxycytidine treatment restored FEZF2 expression in NPC cell lines along with its promoter demethylation. FEZF2 was frequently downregulated in NPC tumors, with promoter methylation detected in 75.5% of tumors, but only in 7.1% of normal nasopharyngeal tissues. Restored FEZF2 expression suppressed NPC cell clonogenicity through inducing G2/M cell cycle arrest and apoptosis and also inhibited NPC cell migration and stemness. FEZF2 acted as a histone deacetylase-associated repressor downregulating multiple oncogenes including EZH2 and MDM2, through direct binding to their promoters. Concomitantly, overexpression of EZH2 was frequently detected in NPC tumors. Thus, we have identified FEZF2 as a novel 3p14.2 TSG frequently inactivated by promoter methylation in NPC, which functions as a repressor downregulating multiple oncogene expression.

Osteopontin increases the expression of ß1, 4-galactosyltransferase-I and promotes adhesion in human RL95-2 cells.

Beta1, 4-Galactosyltransferase-I (ß1, 4-GalT-I), which transfers galactose from UDP-Gal to N-acetylglucosamine and N-acetylglucosamine-terminated oligosaccharides of N- and O-linked glycans in a ß(1-4) linkage, plays a critical role in cell adhesion, sperm-egg recognition, neurite growth, and tumor cell migration and invasion. Our previously experiments also show that ß1, 4-GalT-I was up-regulated by estrogens and some important cytokines of embryo implantation especially Interleukin-1 (IL-1), TGF-α and Leukemia Inhibitory Factor (LIF) in endometrial cells. In the receptive phase human uterus, osteopontin (OPN) is the most highly up-regulated extracellular matrix/adhesion molecule/cytokine. In this study, we demonstrated the correlated expression of OPN and ß1, 4-GalT-I in endometrium during early pregnancy, and recombinant human OPN (rhOPN) protein induced the ß1, 4-GalT-I up-regulation in RL95-2 cells. Inhibition of MEK/ERK, PI3K/AKT and NF-κB suppressed rhOPN-induced ß1, 4-GalT-I expression. In addition, rhOPN promoted the adhesion of blastocysts cells in vitro in ß1, 4-GalT-I-dependent manner. Moreover, the adhesion is greatly inhibited when ß1, 4-GalT-I was blocked with the specific antibody. Taken together, our data suggest that ß1, 4-GalT-I provides a mechanism to bridge embryo to endometrium during implantation.

Hypomethylation and Over-Expression of the Beta Isoform of BLIMP1 is Induced by Epstein-Barr Virus Infection of B Cells; Potential Implications for the Pathogenesis of EBV-Associated Lymphomas.

B-lymphocyte-induced maturation protein 1 (BLIMP1) exists as two major isoforms, α and ß, which arise from alternate promoters. Inactivation of the full length BLIMP1α isoform is thought to contribute to B cell lymphomagenesis by blocking post-germinal centre (GC) B cell differentiation. In contrast, the shorter ß isoform is functionally impaired and over-expressed in several haematological malignancies, including diffuse large B cell lymphomas (DLBCL). We have studied the influence on BLIMP1ß expression of the Epstein-Barr virus (EBV), a human herpesvirus that is implicated in the pathogenesis of several GC-derived lymphomas, including a subset of DLBCL and Hodgkin's lymphoma (HL). We show that BLIMP1ß expression is increased following the EBV infection of normal human tonsillar GC B cells. We also show that this change in expression is accompanied by hypomethylation of the BLIMP1ß-specific promoter. Furthermore, we confirmed previous reports that the BLIMP1ß promoter is hypomethylated in DLBCL cell lines and show for the first time that BLIMP1ß is hypomethylated in the Hodgkin/Reed-Sternberg (HRS) cells of HL. Our results provide evidence in support of a role for BLIMP1ß in the pathogenesis of EBV-associated B cell lymphomas.

Epigenetic silencing of a proapoptotic cell adhesion molecule, the immunoglobulin superfamily member IGSF4, by promoter CpG methylation protects Hodgkin lymphoma cells from apoptosis.

The malignant Hodgkin/Reed-Sternberg (HRS) cells of Hodgkin lymphoma (HL) are believed to derive from germinal center (GC) B cells, but lack expression of a functional B cell receptor. As apoptosis is the normal fate of B-cell receptor-negative GC B cells, mechanisms that abrogate apoptosis are thus critical in HL development, such as epigenetic disruption of certain pro-apoptotic cancer genes including tumor suppressor genes. Identifying methylated genes elucidates oncogenic mechanisms and provides valuable biomarkers; therefore, we performed a chemical epigenetic screening for methylated genes in HL through pharmacological demethylation and expression profiling. IGSF4/CADM1/TSLC1, a pro-apoptotic cell adhesion molecule of the immunoglobulin superfamily, was identified together with other methylated targets. In contrast to its expression in normal GC B cells, IGSF4 was down-regulated and methylated in HL cell lines, most primary HL, and microdissected HRS cells of 3/5 cases, but not in normal peripheral blood mononuclear cells and seldom in normal lymph nodes. We also detected IGSF4 methylation in sera of 14/18 (78%) HL patients but seldom in normal sera. Ectopic IGSF4 expression decreased HL cells survival and increased their sensitivity to apoptosis. IGSF4 induction that normally follows heat shock stress treatment was also abrogated in methylated lymphoma cells. Thus, our data demonstrate that IGSF4 silencing by CpG methylation provides an anti-apoptotic signal to HRS cells important in HL pathogenesis.