TACI Research Articles

Abstract 1889: Dual-specific antibodies that bind to BCMA and TACI (HMBD-009) enable better targeting of multiple myeloma in pre-clinical models

Abstract Existing immunotherapies for multiple myelomas (MM), including bispecific antibodies, antibody-drug conjugates and chimeric antigen receptor-based strategies, have targeted the B-cell maturation antigen (BCMA) that is expressed at the surface of malignant plasma cells (PCs). Although promising, clinical outcomes are hampered by resistance mechanisms that include BCMA extracellular domain shedding and cell-surface downregulation. In addition, BCMA expression is known to be variable among MM patients. Transmembrane activator and CAML interactor (TACI), another member of the tumor necrosis factor receptor (TNFR) super family, represents another viable drug target. TACI expression largely overlaps with BCMA and expression is also increased in PCs of MM patients. Interestingly, TACI may have a role in maintaining an immunosuppressive tumor microenvironment as its expression was observed to be upregulated in Tregs in MM patient's tumor load. Activation of TACI in Tregs triggers their proliferation and survival further supporting a pro-tumoral role for this receptor. Altogether, the dual targeting of both BCMA and TACI should be beneficial to the efficient removal of malignant PCs. Despite low sequence homology, BCMA and TACI share a similar 3D structure, which is likely the reason that APRIL (A proliferation-Inducing ligand) binds to both receptors. We applied our proprietary Rational Antibody Discovery platform to successfully direct the generation of mouse IgG clones able to bind a conserved epitope on BCMA and TACI with sub-nanomolar affinities. In addition, these antibodies were able to inhibit the binding of BCMA to APRIL - an interaction known to favor MM tumor growth and survival. Two IgG clones were successfully humanized and retained their capacity to bind to both BCMA and TACI when heterologously expressed on HEK293, or endogenously expressed in MM cancer cell lines such as H929. No cross-reactivity to other TNFR family members was observed. In antibody-dependent cell-mediated cytotoxicity (ADCC) assays, these clones were able to induce efficient cell killing by binding either BCMA or TACI. Moreover, our lead dual-specific anti-BCMA/anti-TACI antibody (HMBD-009) showed better tumor control compared to other anti-BCMA only antibodies in pre-clinical tumor models of MM. Dual-specific antibodies able to efficiently target both BCMA and TACI represent a promising new avenue to improve the elimination of malignant PCs from MM patients. Citation Format: Fabien M. Decaillot, Dipti Thakkar, Siyu Guan, Konrad Paszkiewicz, Piers J. Ingram, Jerome D. Boyd-Kikurp. Dual-specific antibodies that bind to BCMA and TACI (HMBD-009) enable better targeting of multiple myeloma in pre-clinical models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1889.

TBK1 and TNFRSF13B mutations and an autoinflammatory disease in a child with lethal COVID-19

Among children, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections are typically mild. Here, we describe the case of a 3.5-year-old girl with an unusually severe presentation of coronavirus disease (COVID-19). The child had an autoinflammatory disorder of unknown etiology, which had been treated using prednisolone and methotrexate, and her parents were half cousins of Turkish descent. After 5 days of nonspecific viral infection symptoms, tonic-clonic seizures occurred followed by acute cardiac insufficiency, multi-organ insufficiency, and ultimate death. Trio exome sequencing identified a homozygous splice-variant in the gene TBK1, and a homozygous missense variant in the gene TNFRSF13B. Heterozygous deleterious variants in the TBK1 gene have been associated with severe COVID-19, and the variant in the TNFRSF13B gene has been associated with common variable immunodeficiency (CVID). We suggest that the identified variants, the autoinflammatory disorder and its treatment, or a combination of these factors probably predisposed to lethal COVID-19 in the present case.

Abstract 495: TNFR2 and PD-1/PD-L1/TNFR2 humanized mice aiding exploration of combination strategies to overcome tumor immune escape mechanisms

Abstract Tumor necrosis factor receptor 2 (TNFR2), also known as tumor necrosis factor receptor superfamily member 1B (TNFRSF1B), is a transmembrane protein, which can mediate both pro- and anti-inflammatory activities of T cells. TNFR2 is mainly expressed on the surface of activated effector T cells and regulatory T cells (Tregs) and promotes proliferation and survival of Tregs through nuclear factor kappa B (NF-κB). Blockade of TNF-TNFR2 interaction with monoclonal antibodies can inhibit the activation of Tregs, possibly resulting in inhibition of their function and/or reduction of their numbers. Yet, until recently the potential of TNFR2 as a therapeutic target for cancer therapy has been underappreciated. Discovery of anti-TNFR2 antibodies, developed to inhibit NF-κB driven growth, has revived excitement for the use of TNFR2as a cancer therapy target and stressed the demand for suitable pre-clinical models to evaluate the safety and efficacy of TNFR2-targeted therapeutics. Biocytogen has generated a humanized TNFR2 mouse model for both in vitro function validation of signaling pathways and in vivo efficacy evaluation of TNFR2 antibodies. In this model, the exons 2~6 of mouse Tnfrsf1b gene which encode the extracellular domain were replaced by human TNFRSF1B counterparts. In the humanized mouse, human TNFR2 was detectable on Tregs in the spleen and the anti-human TNFR2 antibodies associated well with the splenocytes. In addition, anti-human TNFR2 antibodies bound well to CD3+ T cells and inhibited tumor growth. Furthermore, Biocytogen has also generated PD-1/PD-L1/TNFR2 multi-gene humanized mice, in which blood cell composition, morphology and the levels of ALT, AST and other indicators were similar to the wild-type counterpart. Basal leukocyte subpopulations including T/B/NK cells, DC, granulocytes, and monocytes/macrophages, were similar between humanized and wild-type mice. Moreover, using the MC38 tumor model, the combination of anti-TNFR2 antibody and anti-PD-1 (or anti-PD-L1) shows enhanced effect compared to the individual groups, demonstrating that the PD-1/PD-L1/TNFR2 humanized mice have a potential to provide a powerful preclinical model for in vivo evaluation of combination therapies. In conclusion, TNFR2 and PD-1/PD-L1/TNFR2 humanized mice are a useful tool to explore anti-PD-1, anti-PD-L1 and anti-TNFR2 combination therapy strategies with promising activity and low cytotoxicity. Citation Format: Huilin Li, Xiaofei Zhou, Jing Zhang, Veronika Chromikova, Qingcong Lin. TNFR2 and PD-1/PD-L1/TNFR2 humanized mice aiding exploration of combination strategies to overcome tumor immune escape mechanisms [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 495.

Expression of B Cell Maturation Antigen (BCMA) and Transmembrane Activator and Caml Interactor (TACI) Are Correlated and Can be Targeted at Multiple Points during the Course of Multiple Myeloma

Therapies directed to specific antigens expressed on multiple myeloma [MM] cells are desirable given the high likelihood of relapse in patients [pts] and the toxicities associated with current treatment options. BCMA and TACI are surface proteins of interest as they promote the survival and proliferation of mature plasma cells. Given that their gene expression by RT-PCR is higher in MM cells compared to normal plasma cells [PCs], and the association of increasing serum BCMA levels with disease progression, we sought to assess if similar patterns were seen on PCs measured by flow cytometry. We specifically examined changes in BCMA/TACI expression (frequency and intensity) with progression of MM, whether expression of BCMA and TACI are correlated, and if levels of their expression correlates with PC burden as determined by IHC.Flow cytometric data were collected using a validated multi-color panel used for clinical assessment of PC burden, with the addition of antibodies to BCMA and TACI, by the Flow Cytometry Clinical Laboratory at our Center. In total, 29 myeloma pts samples were analyzed, and a retrospective chart review of these individuals was performed under an IRB-approved protocol.Pts were separated into 4 sets of groups based on their number of prior lines of therapy, status of disease, prior autologous stem cell transplant [ASCT], and early relapse post-ASCT. Frequency and intensity of BCMA/ TACI expression were examined across these groups by assessing the following variables within each set of groups: % BCMA+ total PCs, % TACI+ total PCs, as well as BCMA/TACI mean fluorescence intensity (MFI). Correlation analysis was performed to assess whether BCMA expression correlated to TACI expression (n=25). To evaluate the correlation of PC burden in BM aspirate with antigen expression, correlation analysis was performed between PC burden and the BCMA/TACI expression (frequency and intensity). Statistical analysis was performed using GraphPad Prism 7 Software. Statistical tests included 1 way ANOVA (comparison of ≥3 means), student t-test (comparison of 2 means), and Pearson correlation (correlation analysis).The median patient age was 62, with 65.5% male and 34.5% female. IgG kappa (37.9%) was the most common type of MM. 17.2% were newly diagnosed, 27.6% had stable disease, 20.7% were in remission, and 39.5% had relapsed/refractory disease. A majority of pts (41.4%) received 1-3 prior lines of therapy. 51.7% underwent ASCT, 10% of which had relapsed, with 13.8% of relapses occurring ≤ 1 year post-ASCT.Four of 29 pts were excluded from data analysis due to insufficient PCs as detected by flow cytometry. ≥Four lines of therapy (n=9), relapsed/refractory disease (n=8), relapse post-ASCT (n=7), and relapse ≤ 1 year post-ASCT (n=2), were all markers of disease progression. There were no significant differences in BCMA/ TACI expression (frequency and intensity) among these groups and their counterparts, but less variability of expression was observed in newly diagnosed pts.There was a positive correlation between % BCMA+ total PCs with % TACI+ total PCs (r= 0.6051, p= 0.0014, n=25), and BCMA MFI with TACI MFI (r=0.5191, p=0.0078, n=25). A similar relationship was noted where aberrant PCs were identified (r=0.9199, p= 0.0094, n=6).Of the patients without minimal residual disease, there was a negative correlation between PCs in BM aspirate with % TACI+ total PCs(r= -0.5211, p= 0.0320, n=17). There were no significant correlations with PCs in BM aspirate and % of BCMA+ total PCs, BCMA MFI, and TACI MFI. BCMA/TACI expression was spread across a broad range for pts with ≤ 10% clonal PCs in BM aspirate, and a narrower range in pts with > 10% clonal PCs in BM aspirate.BCMA and TACI expression alone are not markers of progression of MM and should be used in conjunction with other antigens associated with progressive disease. However, the persistent and frequent expression of BCMA and TACI identified suggests that BCMA and TACI directed therapies can be used at multiple points during the course of disease. Since expression seems to be more uniform in newly diagnosed patients, patients treated at later time points may benefit from quantitative assessment of BCMA and/or TACI expression prior to receiving therapies targeting these molecules. Finally, expression of BCMA and TACI are correlated, but it remains to be seen if loss of one antigen could precipitate obligate loss of the other. DisclosuresNo relevant conflicts of interest to declare.

Pivotal factors associated with the immunosuppressive tumor microenvironment and melanoma metastasis.

BackgroundConsidering melanoma is the deadliest malignancy among dermatoma and presently lacks effective therapies, there is an urgent need to investigate the potential mechanisms underlying melanoma metastasis and determine prospective therapeutic targets for precise treatment of melanoma.MethodHub genes in melanoma metastasis were identified by analyzing RNA‐seq data (mRNA, miRNA, and lncRNA) obtained from TCGA database. Then the identified hub genes were validated in human tissues with qRT‐PCR, followed by survival analysis. Competing endogenous RNAs of the hub genes were defined to clarify potential molecular mechanism of melanoma progression. Then central gene‐related signaling pathways were analyzed, followed by immune cell abundance analysis in tumor microenvironment with CYTERSORTx.ResultA tetrad of IL2RA, IL2RG, IFNG, and IL7R genes were determined as hub genes and verified by qRT‐PCR, which were significantly associated with unfavorable prognosis in melanoma. LINC02446, LINC01857, and LINC02384 may act as competing endogenous lncRNAs of IL2RA and IL7R through absorbing their shared miR.891a.5p and miR.203b.3p. JAK—STAT signaling pathway identified as the most relevant pathway in melanoma metastasis, as well as a wealthy of genes including TNFRSF 13B, TNFRSF17, TNFRSF9, TNFRSF8, TNFRSF13C, TNFRSF11B, LAG3, NRP1, ENTPD1, NT5E, CCL21, and CCR7, may induce tumor autoimmune suppression through enhancing regulatory T‐cell abundance and performance in the tumor microenvironment. And regulatory T‐cell proportion was indeed critically elevated in metastatic melanoma relative to primary melanoma, as well as in highly expressed IL2RA, IL2RG, IL7R, and IFNG group than their respective counterparts.ConclusionElevated IL2RA, IL2RG, IL7R, and IFNG expression may play a central role in promoting melanoma metastasis through up regulation of intratumoral regulatory T—cell proportion mainly by activation of JAK—STAT signaling pathway. LINC02446, LINC01857, and LINC02384 may stimulate melanoma progression by reducing tumor‐protecting miR.891a.5p and miR.203b.3p. A number of identified molecules including TNFRSF13B, LAG3, NRP1, ENTPD1, NT5E, CCL21, and CCR7 can serve as future therapeutic targets in melanoma treatment.

Exploring pharmacogenetic variants for predicting response to anti-TNF therapy in autoimmune diseases: a meta-analysis.

Aim: The aim of this study is to explore how SNPs may affect the response to anti-TNF-α therapy in the major autoimmune diseases, such as psoriasis, rheumatoid arthritis, inflammatory bowel diseases and Spondyloarthritis. Methodology: We conducted a systematic overview on the field, by assessing all studies that examined the association between polymorphisms and response to anti-TNF-α therapy in participants of European descent. Results: In total, six independent SNPs located in FCGR2A, FCGR3A, TNF-α and TNFRSF1B genes were significantly associated with response to TNF-α blockers, found mainly in disease-subgroup analyses. Conclusion: No common pharmacogenetic variant was identified for all autoimmune diseases under study, suggesting the requirement of more studies in the field in order to capture such predictive variants that will aid treatment selection.

Effect of Anti-TNF Therapy on Mucosal Apoptosis Genes Expression in Crohn's Disease.

Crohn's disease (CD) is a chronic immune-mediated disorder for which there is not a fully effective treatment. Moreover, biological therapy with anti-tumor necrosis factor-α (anti-TNF-α) monoclonal antibodies leads to an effective response in only 60–70% of patients. Our previous data suggested that specific loci polymorphism of the TNFRSF1B, FCGR3A, IL1R, IL1B, and FAS genes could be a predictor of the primary non-response to anti-TNF therapy in CD patients. In this work, we propose to explain this hypothesis by functional analysis in colon biopsies and in a cell culture model. Using the RT-qPCR analysis, we estimated the FCGR3A, IL1R, TNFRSF1B, IL1B, FAS, and ADAM17 genes mRNA level in colon biopsies material from inflamed and non-inflamed tissue from 21 CD patients (14 responders and 7 non-responders to anti-TNF therapy) and 6 controls, as well as in vitro in a peripheral blood mononuclear cells (PBMCs) from 14 CD patients (seven responders and seven non-responders to anti-TNF therapy) and eight controls cultured for 72 h with 10 μg/ml of anti-TNF antibody. Our findings demonstrated a significant down-regulation of TNFRSF1B gene expression in non-responders both in inflamed and in non-inflamed colon tissue, while the expression of the FCGR3A and IL1B genes was significantly up-regulated in non-responders in the inflamed colon region. In vitro research results indicate that the anti-TNF drug induced a significant decrease in TNFRSF1B, FCGR3A, and FAS gene expression in non-responders. These results show that altered TNFRSF1B, FCGR3A, and IL1B genes expression can be a predictor of the primary non-response to anti-TNF therapy in CD patients.

Further analyses of APRIL/APRIL-receptor/glycosaminoglycan interactions by biochemical assays linked to computational studies.

A proliferation-inducing ligand (APRIL) is a member of the tumor necrosis factor superfamily. APRIL is quite unique in this superfamily for at least for two reasons: (i) it binds to glycosaminoglycans (GAGs) via its positively charged N-terminus; (ii) one of its signaling receptor, the transmembrane activator and CAML interactor (TACI), was also reported to bind GAGs. Here, as provided by biochemical evidences with the use of an APRIL deletion mutant linked to computational studies, APRIL-GAG interaction involved other regions than the APRIL N-terminus. Preferential interaction of APRIL with heparin followed by chondroitin sulfate E was confirmed by in silico analysis. Both computational and experimental approaches did not reveal the heparan sulfate binding to TACI. Together, computational results corroborated experiments contributing with atomistic details to the knowledge on this biologically relevant trimolecular system. Additionally, a high-throughput rigorous analysis of the free energy calculations data was performed to critically evaluate the applied computational methodologies.

TNFRSF1B Gene Variants and Related Soluble TNFR2 Levels Impact Resilience in Alzheimer's Disease.

Tumor necrosis factor receptor 2 (TNFR2) promotes neuronal survival downstream. This longitudinal study evaluated whether the TNFRSF1B gene encoding TNFR2 and levels of its soluble form (sTNFR2) affect Alzheimer disease (AD) biomarkers and clinical outcomes. Data analyzed included 188 patients in the Alzheimer's Disease Neuroimaging Initiative (ADNI) who had mild cognitive impairment (MCI) and AD dementia. Further, a replication study was performed in 48 patients with MCI with positive AD biomarkers who were treated at a memory clinic. Cerebrospinal fluid (CSF) sTNFR2 levels along with two related TNFRSF1B gene single nucleotide polymorphisms (SNPs) rs976881 and rs1061622 were assessed. General linear models were used to evaluate the effect of CSF sTNFR2 levels and each SNP in relationship to CSF t-tau and p-tau, cognitive domains, MRI brain measures, and longitudinal cognitive changes after adjustments were made for covariates such as APOE ε4 status. In the ADNI cohort, a significant interaction between rs976881 and CSF sTNFR2 modulates CSF t-tau and p-tau levels; hippocampal and whole brain volumes; and Digit Span Forwards subtest scores. In the replication cohort, a significant interaction between rs976881 and CSF sTNFR2 modulates CSF p-tau. A significant interaction between rs976881 and CSF sTNFR2 also impacts Clinical Dementia Rating Sum of Boxes scores over 12 months in the ADNI cohort. The interaction between TNFRSF1B variant rs976881 and CSF sTNFR2 levels was noted to modulate multiple AD-associated severity markers and cognitive domains. This interaction impacts resilience-related clinical outcomes in AD and lends support to sTNFR2 as a promising candidate for therapeutic targeting to improve clinical outcomes of interest.

TNFRSF13B Diversification Fueled by B Cell Responses to Environmental Challenges-A Hypothesis.

B cell differentiation and memory are controlled by the transmembrane activator and CAML interactor (TACI), a receptor encoded by TNFRSF13B. TNFRSF13B mutations are frequently found in common variable immunodeficiency (CVID) and in IgA -deficiency; yet, ~98% of those with mutant TNFRSF13B are healthy. Indeed, TNFRSF13B is among the 5% most polymorphic genes in man. Other mammals evidence polymorphism at comparable loci. We hypothesize that TNFRSF13B diversity might promote rather than detract from well-being by controlling key elements of innate immunity. We shall discuss how extraordinary diversity of TNFRSF13B could have evolved and persisted across diverse species of mammals by controlling innate and adaptive B cell responses in apparently paradoxical ways.

Abstract PS7-03: Changes in body mass index (BMI) over the life course are associated with gene expression in postmenopausal women

Abstract Introduction: Changes in body mass index (BMI) over the life course are associated with both mammographic breast density and breast cancer risk in postmenopausal women. The underlying biological mechanisms driving these associations are, however, yet to be elucidated. Understanding these mechanisms could provide important insights into breast cancer prevention early in life. To provide insight into the biological mechanisms underlying the associations of BMI change over the life course and breast cancer risk in postmenopausal women, we comprehensively investigated the associations of BMI change from ages 10 and 18 to current age with gene expression of biomarkers that have previously been associated with breast cancer risk: growth factors (IGF1, IGFBP3, FGF1, FGF12, TGFB1), sex hormones (PRL, PGR, ESR1, STAT1, STAT5), and receptor activator of nuclear factor-κB (RANK) pathway (RANK, RANKL, OPG, BMP2, TNFRSF13B, TNFRSF18) gene expression in postmenopausal women. Methods: We investigated these in 372 postmenopausal women free from breast cancer recruited during annual screening mammogram. We estimated BMI at age 10 using a validated 9-level pictogram. Gene expression levels were measured using NanoString nCounter system. We investigated the associations of BMI change with gene expression in multivariable linear regression models, adjusted for confounders. Results: The mean age of study participants was 58 years. Increase in BMI over the life course was associated with an increase in BMP2 gene expression but a decrease in RANK, RANKL, and TNFRSF13B gene expression. Compared to women who had a BMI gain of 0.1-5 kg/m2 from age 10, BMP2 gene expression increased in women who had a BMI gain of 5.1-10 kg/m2 (beta coefficient [β] = 0.48, 95% confidence interval [95% CI] = 0.04 to 0.92); BMI gain of 10.1-15 kg/m2 (β = 0.47, 95% CI = 0.03 to 0.91); and BMI gain of > 15 kg/m2 (β = 0.58, 95% CI = 0.15 to 1.02) (p-trend = 0.04). Similar results were observed for BMI gains from age 18 (p-trend < 0.01). Compared to women who had a BMI gain of 0.1-5 kg/m2 from age 10, RANK gene expression decreased in women who had a BMI gain of 5.1-10 kg/m2 (β = -0.21, 95% CI= -0.51 to 0.09); BMI gain of 10.1-15 kg/m2 (β = -0.29, 95% CI= -0.59 to 0.00); and BMI gain of >15 kg/m2 (β = -0.37, 95% CI= -0.66 to -0.07) (p-trend < 0.01). Similar results were observed for weight gains from age 10 and RANKL and TNFRSF13B gene expression, although the associations were weaker for RANKL gene expression. Changes in BMI over the life course were not associated with OPG, growth factors or sex hormone gene expression. Conclusions: Changes in BMI from childhood, late adolescence, and early adulthood were associated with RANK pathway gene expression in postmenopausal women. Our findings offer important and innovative new insights into how childhood adiposity may confer long-term protection against breast cancer risk in postmenopausal women. Citation Format: Yunan Han, Graham A Colditz, Adetunji T Toriola. Changes in body mass index (BMI) over the life course are associated with gene expression in postmenopausal women [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS7-03.

B Cell Dysregulation in Common Variable Immunodeficiency Interstitial Lung Disease.

Common variable immunodeficiency (CVID) is the most frequently diagnosed primary antibody deficiency. About half of CVID patients develop chronic non-infectious complications thought to be due to intrinsic immune dysregulation, including autoimmunity, gastrointestinal disease, and interstitial lung disease (ILD). Multiple studies have found ILD to be a significant cause of morbidity and mortality in CVID. Yet, the precise mechanisms underlying this complication in CVID are poorly understood. CVID ILD is marked by profound pulmonary infiltration of both T and B cells as well as granulomatous inflammation in many cases. B cell depletive therapy, whether done as a monotherapy or in combination with another immunosuppressive agent, has become a standard of therapy for CVID ILD. However, CVID is a heterogeneous disorder, as is its lung pathology, and the precise patients that would benefit from B cell depletive therapy, when it should administered, and how long it should be repeated all remain gaps in our knowledge. Moreover, some have ILD recurrence after B cell depletive therapy and the relative importance of B cell biology remains incompletely defined. Developmental and functional abnormalities of B cell compartments observed in CVID ILD and related conditions suggest that imbalance of B cell signaling networks may promote lung disease. Included within these potential mechanisms of disease is B cell activating factor (BAFF), a cytokine that is upregulated by the interferon gamma (IFN-γ):STAT1 signaling axis to potently influence B cell activation and survival. B cell responses to BAFF are shaped by the divergent effects and expression patterns of its three receptors: BAFF receptor (BAFF-R), transmembrane activator and CAML interactor (TACI), and B cell maturation antigen (BCMA). Moreover, soluble forms of BAFF-R, TACI, and BCMA exist and may further influence the pathogenesis of ILD. Continued efforts to understand how dysregulated B cell biology promotes ILD development and progression will help close the gap in our understanding of how to best diagnose, define, and manage ILD in CVID.

Cerebrospinal Fluid Biomarkers in Relation to MRZ Reaction Status in Primary Progressive Multiple Sclerosis.

The MRZ reaction (MRZR) comprises the three antibody indices (AIs) against measles, rubella, and varicella zoster virus, reflecting an intrathecal polyspecific B cell response highly specific for multiple sclerosis (MS). Thus, MRZR can be used to confirm a diagnosis of primary progressive MS (PPMS) but its pathophysiological and wider clinical relevance is unclear. This study aimed to investigate whether PPMS patients with a positive MRZR (MRZR+) differ from those with a negative MRZR (MRZR-) according to cerebrospinal fluid (CSF) biomarkers of B cell activity, neuroaxonal damage or glial activity, and clinical features. (1) Methods: In a multicenter PPMS cohort (n = 81) with known MRZR status, we measured B cell-activating factor (BAFF), chemokine CXC ligand 13 (CXCL-13), soluble B cell maturation antigen (sBCMA), soluble transmembrane activator and CAML interactor (sTACI), and chitinase-3-like protein 1 (CHI3L1) in the CSF with enzyme-linked immunosorbent assays (ELISAs). Glial fibrillary acidic protein (GFAP) and neurofilament light chain (NfL) were detected in serum and CSF using single molecule array (SIMOA) technology. (2) Results: MRZR+ patients (45.7% of all PPMS patients) revealed higher levels of NfL in CSF compared to MRZR- patients (54.3%). There were positive correlations between each of sBCMA, sTACI, and intrathecal immunoglobin G (IgG) synthesis. Additionally, NfL concentrations in serum positively correlated with those in CSF and those of GFAP in serum. However, MRZR+ and MRZR- patients did not differ concerning clinical features (e.g., age, disease duration, Expanded Disability Status Scale (EDSS) at diagnosis and follow-up); CSF routine parameters; CSF concentrations of BAFF, CXCL-13, sBCMA, sTACI, CHI3L1, and GFAP; or serum concentrations of GFAP and NfL. (3) Conclusions: In PPMS patients, MRZR positivity might indicate a more pronounced axonal damage. Higher levels of the soluble B cell receptors BCMA and transmembrane activator and CAML interactor (TACI) in CSF are associated with a stronger intrathecal IgG synthesis in PPMS.

Is Polymorphism in the Apoptosis and Inflammatory Pathway Genes Associated With a Primary Response to Anti-TNF Therapy in Crohn's Disease Patients?

Anti-tumor necrosis factor (TNF) therapy is used for the induction and maintenance of remission in Crohn’s disease (CD) patients. However, primary nonresponders to initial treatment constitute 20%–40% of cases. The causes of this phenomenon are still unknown. In this study, we aimed to determine the genetic predictors of the variable reactions of CD patients to anti-TNF therapy. Using long-range PCR libraries and the next-generation sequencing (NGS) method, we performed broad pharmacogenetic studies including a panel of 23 genes (TNFRSF1A, TNFRSF1B, CASP9, FCGR3A, LTA, TNF, FAS, ADAM17, IL17A, IL6, MMP1, MMP3, S100A8, S100A9, S100A12, TLR2, TLR4, TLR9, CD14, IL23R, IL23, IL1R, and IL1B) in a group of 107 diagnosed and clinically characterized CD patients following anti-TNF therapy. In the studied group, we indicated, in total, 598 single nucleotide variants for all analyzed genomic targets. Twelve patients (11.2%) did not respond to the induction therapy, which was associated with alleles in 11 loci located in FCGR3A (rs7539036, rs6672453, rs373184583, and rs12128686), IL1R (rs2041747), TNFRSF1B (rs5746053), IL1B (rs1071676, rs1143639, rs1143637, and rs1143634), and FAS (rs7896789) genes. After multiple comparison corrections, the results were not statistically significant, however for nonresponders the alleles distribution for those loci presented large differences and specified scheme compared to responders and populations. These findings require further investigation in an independent larger cohort before introducing them for a clinical setting, however, we identified an interesting direction. Polymorphism of the FCGR3A, IL1R, TNFRSF1B, IL1B, and FAS genes could be a predictor of the primary response to anti-TNF therapy in CD patients.

Association of CRP, CD14, Pro-Inflammatory Cytokines and Their Receptors (TNFA, LTA, TNFRSF1A, TNFRSF1B, IL1B, and IL6) Genes with Chronic Obstructive Pulmonary Disease Development

The aim of the present study was to investigate the association of COPD and frequent exacerbator COPD phenotype with CRP, CD14, and pro-inflammatory cytokines and their receptors (TNFA, LTA, TNFRSF1A, TNFRSF1B, IL1B, and IL6) genes. It was found that COPD was associated with allele A of the TNFA gene (rs1800629G>A) (P = 0.002, OR = 1.45); the association was established in the log-additive model (P = 0.0022, Pcor-FDR = 0.01705, OR = 1.47); this association was confirmed in the frequent exacerbator COPD phenotype group (P = 0.001, Pcor-FDR = 0.007, OR = 1.59). Allele G of the LTA gene (rs909253A>G) (P = 0.002, OR = 1.33) was also shown to be a marker for COPD risk; the association was established in the log-additive model (P = 0.0021, Pcor-FDR = 0.01705, OR = 1.31) and it was confirmed in patients with rare exacerbations (P = 0.003, Pcor-FDR = 0.0084, OR = 1.39). The genotype GG of the TNFRSF1B gene (rs1061622T>G) was a marker of resistance to the development of the frequent exacerbator COPD phenotype (P = 0.003, Pcor-FDR = 0.0084, OR = 0.46). The genotype CC of the CD14 gene (rs2569190T>C) was associated with higher forced expiratory volume in 1 s (P = 0.006); subjects with genotype AA of the TNFRSF1B gene (rs1061624A >G) and genotype GG of the LTA gene (rs909253A>G) exhibited lower forced expiratory volume in 1 s (P = 0.04 and P = 0.01, respectively). Genotype AA of the TNFRSF1A gene (rs767455A>G) was associated with higher smoking pack-years (P = 0.0036).

Human Cumulus Cells in Long-Term In Vitro Culture Reflect Differential Expression Profile of Genes Responsible for Planned Cell Death and Aging-A Study of New Molecular Markers.

In the ovarian follicle, maturation of the oocyte increases in the presence of somatic cells called cumulus cells (CCs). These cells form a direct barrier between the oocyte and external environment. Thanks to bidirectional communication, they have a direct impact on the oocyte, its quality and development potential. Understanding the genetic profile of CCs appears to be important in elucidating the physiology of oocytes. Long-term in vitro culture of CCs collected from patients undergoing controlled ovarian stimulation during in vitro fertilization procedure was conducted. Using microarray expression analysis, transcript levels were assessed on day 1, 7, 15, and 30 of culture. Apoptosis and aging of CCs strictly influence oocyte quality and subsequently the outcome of assisted reproductive technologies (ART). Thus, particular attention was paid to the analysis of genes involved in programmed cell death, aging, and apoptosis. Due to the detailed level of expression analysis of each of the 133 analyzed genes, three groups were selected: first with significantly decreased expression during the culture; second with the statistically lowest increase in expression; and third with the highest significant increase in expression. COL3A1, SFRP4, CTGF, HTR2B, VCAM1, TNFRSF11B genes, belonging to the third group, were identified as potential carriers of information on oocyte quality.

Bispecific CAR T cells for multiple myeloma: natural ligand compared to tandem scFv design

Abstract One of the most promising candidates for Chimeric Antigen Receptor (CAR) T cell therapy beyond CAR19 is treatment of multiple myeloma with CAR T cells targeting B cell maturation antigen (BCMA). However, current reports of BCMA CAR in the clinic have a median progression free survival of 11.8 months, suggesting that targeting BCMA alone may not be sufficient. Evidence suggests patients treated with BCMA-targeted therapies may be thwarted by BCMA negative relapse. We have developed a novel second generation CAR T cell against another multiple myeloma target: transmembrane activator and CAML interactor (TACI). Mice were immunized against TACI and we designed a new scFv targeting TACI based on the resulting antibodies. Anti-TACI CAR T cells are cytotoxic in vitro and in vivo against multiple myeloma. To overcome single antigen loss, we designed bispecific tandem scFv CAR T cells targeting BCMA and TACI. These two antigens share a natural ligand, A Proliferation-Inducing Ligand (APRIL), which has also been used to design a dual-targeting CAR called TriPRIL. We show that these dual targeting CARs, based on tandem scFv or natural ligand design, have similar efficacy against wild type multiple myeloma models. However, this changes in the context of single antigen loss. We have characterized the proliferative (population doublings) and activation capability (CD69) of these CARs, as well as their memory (CCR7, CD45RA) and exhaustion phenotype (PD-1, TIM-3, LAG-3), with long term exposure to single antigen. Our studies show that sensitivity to antigen density differs between the tandem bispecifics and the natural ligand CAR. These data provide insight into how structural differences between dual-targeting CAR T cells affects their function.

Transmembrane Activator and CAML Interactor (TACI): Another Potential Target for Immunotherapy of Multiple Myeloma?

Multiple myeloma (MM) has emerged as the next most likely oncological or hematological disease indication amenable for cellular immunotherapy. Much of the attention has been focused on B cell maturation antigen (BCMA) as a unique cell surface protein on myeloma cells that is available for monoclonal antibodies, antibody drug conjugates (ADCs), T-cell redirecting bispecific molecules, and chimeric antigen receptor (CAR) T cell targeting. BCMA is a member of the tumor necrosis factor receptor (TNFR) superfamily that binds two ligands B-cell activating factor (BAFF) and a proliferation-inducing ligand (APRIL) and mediates the growth and survival of plasma and MM cells. Interestingly, transmembrane activator and CAML interactor (TACI), another TNFR superfamily member, also binds the same ligands and plays largely overlapping roles as BCMA in normal plasma and malignant MM cells. In this article, we review the biology of TACI, focusing on its role in normal B and plasma cells and malignant MM cells, and also discuss various ways to incorporate TACI as a potential target for immunotherapies against MM.

Juvenile Paget’s disease: Report of a family with a novel missense mutation and unique radiological manifestations in a heterozygote

Abstract Juvenile Paget’s disease (JPD) is a rare, generalized skeletal disorder characterized by markedly increased bone turnover secondary to enhanced osteoclastic activity. The patients with JPD develop progressive, widespread skeletal involvement and non-skeletal manifestations. Objectives To identify the characteristic molecular, biochemical, radiological, and audiological findings in three siblings with JPD associated with intrafamilial phenotypic variability. Patients and Methods A Saudi family with 3 affected siblings was recruited. Biochemical measurement of serum alkaline phosphatase and markers of bone turnover were performed. Genetic testing and sequencing of a panel of 22 known genes for skeletal dysplasia with increase bone density were performed by next generation sequencing (NGS). Radiological and audiological assessment were also performed. Results Patients showed marked elevation of serum alkaline phosphatase and markers of bone turnover. A novel homozygous mutation c.433T>G (p.Cys145Gly) in exon 3 of TNFRSF11B gene was identified in the 3 affected siblings. Both parents were heterozygous for the mutation. The heterozygote father had thickening of calvarium, a radiological manifestation of JPD. The eldest child (index case) had a unique striking distortion of the skull bones as evidenced by 3-D computed tomography of skull. He had also bilateral inner ear structural deformity and severe sensorineural hearing loss. Conclusions: JPD is a rare disorder that can be highly overlooked due to phenotypic overlap with other disorders of skeletal dysplasia. Mutations affecting cysteine residues result in the most severe JPD phenotypes. NGS is useful for early diagnosis of JPD, a prerequisite for successful treatment strategy.

Development TACI and Bcma Dual-Antigen Targeted Immunotherapy for Multiple Myeloma

Introduction: Despite recent advances in treatment for multiple myeloma(MM), there remains a need for novel therapeutic approaches. Recently, we have reported that antigen-specific CD8+ cytotoxic T lymphocytes (CTL) with anti-MM activity can be induce by immunogenic peptides to XBP1 (X-box binding protein 1), CD138 (Syndecan-1) and CS1 (SLAMF7). Based on these results, multicenter Phase 1/2a trials are completed in patients with smoldering multiple myeloma (SMM) (JAMA Oncol.2018) and on-going in patients with SMM or triple negative breast cancer, alone and in combination with checkpoint inhibitors, lenalidomide, and HDAC6 inhibitor. We here expand the breadth and extent of MM-specific immunotherapies by targeting additional tumor-associated antigens, including B-cell maturation antigen (BCMA) and transmembrane activator and CAML interactor (TACI), TNF receptor family proteins which are involved in maturation of B-cells and highly expressed in MM. Purpose: We aim to develop a dual antigen targeted immunotherapeutic approach to induce central memory-specific anti-MM immunity and improve patient outcome. Results: We demonstrated that engineered BCMA72-80 (YLMFLLRKI) and BCMA54-62 (YILWTCLGL) peptides can evoke BCMA-specific CTL and their specific activities against MM (Bae et al. 2019). Here, we report the identification of a novel heteroclitic TACI peptide with improved HLA-A2 binding/stability compared to the native TACI peptides. The heteroclitic TACI peptide induces antigen-specific memory CD8+ CTL with robust anti-tumor activities (CD107a degranulation, Granzyme B upregulation, Th1 cytokine production) against HLA-A2+ MM (U266, McCAR) cells, but not against HLA-A2- MM (OPM2, RPMI) nor HLA-A2+ breast cancer (MDA-MB231) cells. In response to HLA-A2+ MM cells, the heteroclitic TACI peptide-specific CTL showed the antigen-specific proliferation of CD8+ CTL expressing costimulatory molecules (CD28 > 41BB > CD40L), which was directly associated with functional anti-tumor activity. These results suggest that the heteroclitic TACI peptide identified is a potential novel therapeutic option to effectively generate TACI-specific CD8+ memory CTL targeting MM. In on-going studies, we are evaluating a combination of immunogenic heteroclitic peptides specific to BCMA and TACI to induce highly functional antigen-specific immune responses by CD8+ Tc and CD4+ Th cells to efficiently target tumor cells. Conclusions: A novel heteroclitic TACI peptide can induce MM-specific central memory CD8+ CTL with robust poly-functional anti-tumor activities. These results provide the framework for therapeutic application targeting combination TACI and BCMA antigens in MM patients, alone and in combination with checkpoint inhibitors, epigenetic regulators and immune modulators, to both enhance anti-MM immunity and improve patient outcome. Disclosures Richardson: Janssen: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol-Myers Squibb: Research Funding. Munshi:Oncopep: Consultancy; Janssen: Consultancy; Takeda: Consultancy; Amgen: Consultancy; Celgene: Consultancy; Adaptive: Consultancy; Abbvie: Consultancy. Anderson:Oncopep: Other: Scientific Founder; Amgen: Consultancy, Speakers Bureau; Sanofi-Aventis: Other: Advisory Board; Bristol-Myers Squibb: Other: Scientific Founder; Janssen: Consultancy, Speakers Bureau; Takeda: Consultancy, Speakers Bureau; Celgene: Consultancy, Speakers Bureau.