Ahmann, Gregory

Abstract Purpose: Naturally occurring tumor-specific T cells derived from peripheral blood (PB) offer a clinically accessible source for adoptive immunotherapy. However, the expansion of these T cells from patients with cancer remains a challenge. We hypothesized that mimicking innate immune activation could optimally stimulate antigen-driven T-cell expansion in vitro, unlocking the therapeutic potential of PB-derived T cells. Patients and Methods: We previously developed an ex vivo culture system that selectively expands tumor antigen–activated T cells from PB mononuclear cells (PBMC), generating multiclonal effector and central memory T cells. In this hypothesis-generating study, we evaluated the therapeutic potential of MUC1-activated T cells in patients with relapsed and/or refractory multiple myeloma (r/rMM) in a phase I clinical trial (NCT05411497). MUC1 is an oncoprotein overexpressed in r/rMM. We translated our small-scale culture into GMP-compliant, large-scale manufacturing, which achieved expansion of T cells from heavily pretreated patients’ PBMCs without exhaustion. Five patients were treated with escalating doses of up to 1 × 1010 T cells. Results: Although the cell infusions were well tolerated, no objective responses occurred. One patient, who received the highest dose, has had stable disease for 2 years after infusion. This patient exhibited transient dermatitis with localized MUC1 and CD3 staining, as potential evidence of on-target, off-tumor T-cell reactivity, perhaps contributing to disease stabilization. T-cell receptor sequencing revealed the T-cell product in four of the patients’ blood, which correlated with the product’s degree of polyfunctionality and MUC1 reactivity. Conclusions: These findings demonstrate the feasibility, safety, and biological activity of PB-derived, MUC1-specific T cells as adoptive immunotherapy. Significance: Expanding tumor antigen–specific T cells for clinical use from the blood of heavily pretreated patients with cancer remains a significant hurdle. We scale up a novel culture method that integrates innate immune signals to stimulate T-cell expansion and demonstrate safety, feasibility, and a case of long-term disease stabilization upon treatment of patients with multiple myeloma in a phase I clinical trial.

Abstract In this study, we investigated tumor intrinsic mechanisms of resistance to cevostamab, a bispecific T-cell engager targeting Fc receptor-like protein 5 (FCRL5) and CD3 that has shown promising clinical activity and durable responses in advanced MM. Longitudinal bone marrow aspirates from nine MM patients treated with cevostamab were analyzed. All patients had previously been treated with both immunomodulatory drugs (IMiDs) and proteasome inhibitors (PI) and eight patients had received daratumumab prior to cevostamab, which was, on average, the 6th line of treatment (mean treatment duration 220±273 days). Five of nine patients responded with ≥PR. Whole genome and transcriptome sequencing was performed on sorted CD138+ MM cells at baseline, and whole exome and transcriptome sequencing was performed on five and three samples, respectively, following cevostamab discontinuation. Baseline FCRL5 expression levels did not correlate with clinical response. However, in one non-responder patient, FCRL5 expression increased four-fold after 28 days of treatment. Given the location of the FCRL5 gene on chromosome 1q—commonly amplified in advanced MM—we evaluated the presence of Gain(1q) in our study cohort. All patients whose best response was ≤PR (n=6) harbored a Gain(1q), whereas it was absent in those who achieved VGPR and sCR (n=3). Crucially, one of the patients that achieved sCR was initially treated with 60 mg of cevostamab for one year and then escalated to a higher dose cohort (90 mg) in the second year. At progression, a clonal FCRL5E115A missense mutation (allelic frequency = 56%) and a subclonal FCRL5G455Efs*19(allelic frequency = 5%) frameshift deletion mutation were detected, both absent at baseline. The missense mutation was located on the extracellular domain of FCRL5 targeted by cevostamab. To validate the functional impact of FCRL5E115Aand FCRL5G455Efs*19 on the sensitivity to cevostamab in vitro, we generated a FCRL5 knockout (KO) model utilizing CRISPR-Cas9 technology in OPM2 MM cells. In vitro cevostamab cytotoxicity assay with PBMCs and pan T cells from healthy donors at various effector to target (E:T) ratios confirmed a complete loss of cevostamab sensitivity in our FCRL5KO models. cDNA encoding patient-derived FCRL5 alterations (FCRL5E115Aand FCRL5G455Efs*19) as well as FCRL5wtwere cloned into mammalian expression vector and reintroduced into FCRL5KO cells. Cevostamab sensitivity was reinstated by FCRL5wt expression in KO cells. In contrast, FCRL5E115A conferred complete resistance to cevostamab (p<0,001) and FCRL5G455Efs*19 depicted a significant decline in cevostamab sensitivity relative to FCRL5wt expressing models. Additional experiment utilizing direct stochastic optical reconstruction microscopy (dSTORM) are ongoing to monitor the membrane localization of FCRL5 mutants. By integrating genomic, transcriptomic and functional data with clinical outcomes, this work is the first to report on acquired FCRL5 mutations causing resistance to cevostamab. We hypothesize that the prolonged treatment duration may have contributed to the acquisition of the target mutations and note a subsequent durable sCR response to BCMA-targeted immunotherapy in the same patient, suggesting immune-related factors such as T cell exhaustion, were not drivers of cevostamab resistance. Due to the limited sample size, these findings require validation in larger cohorts. Nonetheless, our findings begin to elucidate tumor intrinsic mechanisms associated with cevostamab resistance.

Introduction The major primary genetic events in multiple myeloma (MM) include translocations of immunoglobulin heavy chain (IgH) and a hyperdiploidy (HRD) karyotype (Kuehl, 2002). The main drivers of pathogenesis in HRD MM are unknown (Fonseca, 2003), therefore further research is needed in this subgroup. Translational research in HRD MM has relied on analysis of genomic and transcriptomic alterations, however changes at the genome and transcriptome level do not always correlate directionally with protein abundance. Because proteins are considered the main and final effectors of most cellular processes, proteogenomics can improve our understanding of the drivers of HRD MM. We employed it here to identify potential biomarkers in HRD MM and determine whether any given trisomy may lead to a selective advantage. Methods: Protein and mRNA were extracted from CD138+ cells from 47 HRD MM and 14 t(11;14) MM samples. RNA-seq data was processed using MAPR-Seq pipeline using default parameters and gene-wise raw counts were extracted for each sample. Proteomics data was processed using Specranaut and protein-wise intensities were extracted for each sample. A generalized linear model configured with negative binomial distribution was used to analyze gene-wise count data in edgeR software. Genes with an adjusted p-value of <=0.05 were considered significantly differentially expressed. Protein intensity data were normalized using trimmed mean of M-values and log2 transformed. A generalized linear model configured with Gaussian distribution was used to compare protein intensities between the groups. Proteins with an adjusted p-value <= 0.05 were considered significantly differentially expressed. Two separate rank lists for protein and mRNA data were generated for pathway analysis. Results When comparing our small cohort of HRD MM patient samples to those with t(11;14) MM, out of approximately 16,600 total transcripts and 9,700 proteins evaluated we identified approximately 2,100 significantly variant transcripts and 500 significantly variant proteins in HRD MM. GSEA pathway analysis of the transcriptome and proteome revealed significantly upregulated translational and ribosomal processing pathways in HRD MM. Transcriptional MYC target pathways were also significantly upregulated. RNAseq revealed significantly higher expression of MYC (Rank 24.5, p=5.3E-6) in HRD MM. It is known that patients with t(11;14) MM have increased expression of CCND1 (Kuehl and Bergsagel, 2012). As expected, we found that the HRD MM subgroup had significantly lower expression of CCND1 compared to the t(11;14) MM subgroup (Rank -79.9, p=2.8E-21) and this was also true at the protein level (Rank -21.7, p=2.2E-5). The most significantly abundant protein in the HRD MM subgroup compared to t(11;14) MM was BAHD1, which is a protein that targets H3K27me3 and recruits transcriptional co-repressors (Fan, 2020). BAHD1 has been shown to repress erythroid genes and may recruit HDAC5, which plays a role in osteoblast differentiation (Rank 72.3, p=2.9E-20; Xu, 2021). The second most abundant protein in HRD MM was CKS2, which binds to the catalytic subunit of cyclin-dependent kinases (CDKs) and therefore positively regulates cell proliferation, invasion, and migration and has been shown to be implicated in tumor progression in several solid tumors (Rank 72.3, p=2.9E-20; You, 2015). TRIM proteins promote tumorigenesis and cancer development. TRIM14 mediates cancer development through mediating the JAK/STAT, PI3K/AKT, NFkB, and p53 pathways and was found to be significantly higher in HRD MM (Rank 70.6, p=9.0E-20; Huang, 2022). We also found increased protein abundance of LAMP5 in HRD MM (Rank 20.1, p=6.0E-5). LAMP5 may play an important role in MM progression and has been shown to be highly expressed in patients with mixed-lineage leukemia and is associated with a poor prognosis (Wang, 2023; Gracia-Maldonado, 2022). We identified many proteins (not all discussed here) that have not been well described in HRD MM and thus may be biomarkers/targets of disease pathogenesis of this subgroup. Conclusion Proteins are the final effectors of most cellular processes. Proteogenomics can improve our understanding of the drivers of HRD MM. Using a proteogenomic approach we identified proteins that may lead to selective advantage and/or pathogenesis of trisomies in HRD MM.

Introduction Multiple myeloma (MM) patients often become unresponsive to immunomodulatory drugs (IMiDs) over time. Research has indicated that the cellular ability to counteract oxidative stress plays a crucial role in determining the sensitivity to IMiDs in MM. This study aims to comprehend the metabolic pathways responsible for MM's antioxidative capability and develop new treatments to augment the effectiveness of IMiDs. Methods We experimented using MM cells to investigate how different nutrient conditions affect cell proliferation and drug response. We utilized stable isotope-labeled amino acids and mass spectrometry to analyze the metabolites in these cells. We evaluated cell viability through the MTT assay, the CellTiter-Glo® Luminescent Assay, and direct cell counting. Furthermore, we conducted Western blots to measure the levels of various proteins expressed in MM cells grown under different nutrient conditions. To create CRBN knockout cells, we employed the CRBN CRISPR/Cas9 KO plasmid, and for the overexpression of CRBN in MM cells, we used a lentivirus system. Results Our research findings reveal that the production of cellular cysteine enhances the cell's ability to counteract oxidative damage, leading to resistance to IMiDs even with the presence of the wild-type CRBN protein. However, MM cells not producing enough cysteine rely on consuming extracellular cystine, which increases sensitivity to lenalidomide and reduces antioxidative capacity. Lenalidomide-sensitive cells acquire more cystine from the extracellular environment than resistant cells. The analysis of cysteine uptake in various MM cell lines using C13-labeled cysteine revealed that the lenalidomide-sensitive cell line, OCIMY5-CRBN, consumed more cysteine from the media, and the presence of L-glutamine further increased this uptake. Lenalidomide treatment increased the catabolism of cysteine into taurine in sensitive cell lines, confirming the link between lenalidomide's molecular action and cysteine catabolism. Our study also identified that lenalidomide treatment led to an accumulation of CDO protein, which adds molecular oxygen to cysteine, transforming it into cysteine sulfinic acid. Our study suggests that CRBN may play a role in maintaining intracellular CDO protein levels, and the accumulation of CDO protein induced by lenalidomide is the molecular factor responsible for cystine catabolism and sensitivity to IMiDs in multiple myeloma. We found that by inhibiting intracellular cysteine biosynthesis, we could improve the effectiveness of IMiD against MM. One drug that can achieve this is 5-aza-2-deoxycytidine (DAC), which acts as a DNA methylation inhibitor. Additionally, combining DAC with IMiDs improves their effectiveness, even in cell lines that are resistant to treatment. Furthermore, we attempted to inhibit cysteine biosynthesis by promoting the synthesis of methionine from homocysteine with the supplementation of 5-methyltetrahydrofolate. We discovered that treatment with 5-methyltetrahydrofolate further enhanced the sensitivity of IMiDs in resistant MM cell lines. Our results confirm that interfering with intracellular cysteine biosynthesis is a promising therapeutic approach for MM and can improve the effectiveness of IMiDs. Conclusions The biosynthesis of L-Cysteine is vital for the cellular ability to resist oxidation. Cancer cells that create high levels of cysteine within the cell from methionine generate a more significant amount of reduced cysteine, which can act as antioxidants and make MM cells resistant to IMiDs. On the other hand, cells that cannot produce enough cysteine increase their consumption of extracellular cystine and are more susceptible to IMiDs. Cells that consume an oxidized form of cystine from outside the cell require a reduction process inside the cell, which further reduces their ability to resist oxidation. Additionally, the accumulation of CDO protein induced by lenalidomide can enhance the breakdown of intracellular cysteine and cystine uptake in vulnerable cells. Overall, MM cells that rely on extracellular cysteine are sensitive to IMiDs. Therefore, interfering with intracellular cysteine biosynthesis is a better option for myeloma treatment and enhancing the sensitivity of IMiDs.

Introduction Fluorescent In-Situ Hybridization (FISH) has been an indispensable tool to characterize the cytogenetic landscape of multiple myeloma (MM), mainly for identifying primary abnormalities such as heavy chain gene (IgH) translocations and odd-number chromosome trisomies. These aberrations play a crucial role in risk stratifying the disease given their correlation with disease aggressiveness, treatment response and overall prognosis. High-risk (HR) cytogenetics are identified through the presence of t(14;16), t(14;20) or loss of the p53 gene locus. Less is known about the prognostic implication of secondary abnormalities, in particular gain of 1q21. We aimed to evaluate the association between primary and secondary cytogenetic abnormalities and their impact on overall survival (OS) in MM patients. Methods We performed a retrospective analysis on a FISH records database from three Mayo Clinic campuses. Cohort included patients >18 years of age, diagnosed with multiple myeloma between 2005 and 2022, with FISH performed within two years of diagnosis. Demographics including age, date of diagnoses and OS data were retrieved from electronic medical records. Statistical analyses were done using BlueSky Statistics software. Study was conducted in accordance with the principles of the Declaration of Helsinki; informed consent was obtained from all participants. Results We analyzed a total of 2221 patients, among which 59.8% were male, with a median age of diagnosis 65 years. Hyperdiploidy was the most prevalent primary cytogenetic abnormality observed in 44.3% of patients, followed by t(11;14) as the most common translocation (23%), and t(4;14) (8.2%). Gain(1q21) was assessed in 925 patients, among which 64%, 30.3% and 5.7% had 2, 3 and ≥4 copies respectively. Furthermore, 11.4% (n=253) exhibited either mutated TP53 or del17p, and 37.8% (n=839) presented with 13q loss or monosomy 13. The median follow-up for the overall population was 102 months (8.5 years), with a median OS of 75.5 months. Patients with high-risk translocations had significantly shorter median OS of 40.01 months compared to 80.65 months for those with standard risk translocations (p <0.001). The presence of ≥4 1q copies was associated with significantly reduced overall survival (2 copies: not reached, 3 copies: 69.4 months, ≥4 copies: 40.1774; p <0.001). This was consistent even when considering the presence of high-risk translocations (Figure 1). To confirm the independent significance of these findings, we performed multivariate Cox regression models, incorporating age at diagnosis, year of diagnosis, gender, presence of hyperdiploid disease, and presence of high-risk translocations. Given that overall survival outcomes were analyzed irrespective of treatment received, we incorporated year of diagnosis as a variable to control for potential impact of evolving treatment regimens over the years. The presence of ≥4 1q copies was associated with reduced OS, with a hazard ratio (HR) of 2.12 (CI: 1.38-3.26; p <0.001). The presence of 3 copies also showed prognostic significance (HR: 1.43; p: 0.012). Similarly, both the presence of TP53mut/mon17 (HR: 1.96, CI: 1.66-2.31; p <0.001) and 13qdel/mon13 (HR: 1.21, CI: 1.08-1.38) were associated with decreased OS. On the other hand, 1p deletion did not have any significant prognostic implications (HR: 1.84, CI: 0.38-1.59; p 0.113). When incorporating all cytogenetic events into a multivariate model, variables associated with reduced OS included age at diagnosis, the presence of both of 3 and ≥4 1q copies, TP53mut/mon17, t(4;14), t(14;16) and t(14;20). However, 13qDel/mon13 lost significance when incorporated with other abnormalities (Table 1). Conclusion Our investigation sheds light on the less explored realm of secondary cytogenetic abnormalities and confirm a strong association between increasing number of 1q copies and reduced overall survival, independent of other cytogenetic abnormalities and treatment regimens. These results underscore the utility of FISH-based comprehensive cytogenetic profiling at baseline, which enables reliable identification of high-risk patients. Future prospective trials should further elucidate the pathogenic mechanism for these findings. Our analysis used OS as primary endpoint and included over 2000 MM patients, reinforcing the clinical relevance of our findings.

Abstract Myeloma is a neoplasia characterized by the accumulation of malignant plasma cells in the bone marrow. In these studies, we have demonstrated that CD40 is expressed in human myeloma cells and have used a recently established IL-6-dependent myeloma cell line, ANBL-6, to examine the potential function of CD40 expression in myeloma cells. In addition to its expression on the ANBL-6 cells, we show that CD40 is expressed on freshly isolated myeloma cells from seven of seven patients tested. To address the role of CD40 expression in myeloma cells, we have examined the responsiveness of the ANBL-6 cell line to a CD40-specific mAb, G28-5. This cell line has previously been shown to proliferate only in response to IL-6. Of interest in this study, G28-5 also induced proliferation of the ANBL-6 cells. This proliferation was substantially inhibited by an IL-6-neutralizing mAb. Analysis of ANBL-6 cell culture supernatants by ELISA demonstrated that G28-5-stimulated cells secreted significant levels of IL-6, whereas unstimulated cell culture supernatants contained undetectable levels of IL-6. Furthermore, CV-1/EBNA cells expressing the human CD40 ligand also induced the proliferation of the ANBL-6 cell line, an effect that was inhibited by the anti-IL-6 mAb. Lastly, RNA blot analysis demonstrated an increase in IL-6 message in G28-5-stimulated ANBL-6 cells over unstimulated cells. These results indicate that the primary mechanism of anti-CD40-stimulated proliferation of the ANBL-6 cells is the induction of autocrine IL-6 production. Moreover, these data suggest that the expression of CD40 in malignant plasma cells may play a role in tumor cell expansion, possibly by stimulating the induction of autocrine IL-6 secretion.

Abstract Venetoclax (VTX) is a selective small-molecule inhibitor of BCL-2 that exhibits antitumoral activity against MM cells presenting lymphoid features and those with translocation t(11;14). Despite its impressive clinical activity, VTX therapy for a prolonged duration can lead to drug resistance. Therefore, it is important to understand the underlying mechanisms of resistance in order to develop strategies to prevent or overcome resistance. In the present study, we established four VTX resistant human myeloma cell lines (HMCLs) from four sensitive HMCLs, including three with t(11;14), in culture with a stepwise increase in treatment dose with VTX. To identify the molecular basis of acquired VTX resistance, whole exon sequencing (WES), mRNA-sequencing (mRNAseq), and protein expression assays were performed in the four isogenic VTX-sensitive/resistant HMCLs and three MM patients with samples collected before VTX administration and after clinical resistance to the drug. Compared with sensitive cell lines and patient samples collected before VTX administration, mRNAseq analysis identified downregulation of BIM and upregulation of BCLXL in both resistant cell lines and MM cells from relapse patients. Other transcriptional changes detected included upregulation of AURKA, BIRC3, BIRC5, and IL32. Enrichment analysis of differentially expressed genes suggested involvement of PI3K and MAPK signaling, likely associated with cytokines, growth factors (EGF, FGF and IGF family members), and receptor tyrosine kinase (EGF and FGF). Western blot analysis was performed to compare BCL2 family expression in resistant cell lines versus sensitive cell lines and it showed upregulation of BCL2 survival members (such as MCL-1 and BCLXL), and downregulation of pro-apoptotic BH3 members (such as BIM and PUMA). BIM expression was completely lost in one resistant cell line, and introduction of exogenous BIM into this cell line enhanced VTX sensitivity. Interestingly, BCL2 was upregulated in some resistant cell lines generated after a long-term treatment with VTX, suggesting BCL2 expression level may not be suitable as a marker of VTX sensitivity for acquired resistance. Unlike in CLL, BCL2 mutations were not identified through WES in any resistant cell lines or primary patient sample harvested after relapse. While 8 genes were mutated in two resistant samples , no clear mutational pattern emerged . Based on the above, we further tested some specific inhibitors in in vitro or ex vivo cell models to help understanding resistant mechanism and identify strategies to overcome VTX resistance. We found that inhibition of MCL-1, with the compound S68345, substantially enhanced VTX sensitivity in three resistant HMCLs and in primary cells from one relapsed MM patient. A BCLXL inhibitor (A155463) only significantly enhanced VTX sensitivity in one resistant cell line after co-treatment with VTX. Co-treatment of the other three resistant cell lines with VTX, S68345 and A155463 resulted in the most synergistic anti-myeloma activity, suggesting those cell lines are co-dependent on MCL-1, BCLXL, and BCL2 for survival, although they are more dependent on MCL-1. We also found that inhibition of PI3K signaling, IGF1, RTK (EGF and FGF) and AURKA significantly increased VTX sensitivity, partially through downregulation of MCL-1, and BCLXL, and upregulation of BIM. Conventional anti-MM drugs such as dexamethasone, bortezomib and lenalidomide, were shown to have little activity on augmenting VTX sensitivity in most resistant cell lines. In summary, we find that acquired resistance to VTX in MM is largely associated with BCL2 family regulation, including upregulation of survival members such as MCL-1, BCLXL, BCL2, and downregulation of pro-apoptotic members, especially BIM. Our study also indicates that upstream signaling involved in BCL2 family regulation during acquired resistance is likely related to cytokine, growth factor, and/or RTK-induced cell signaling such as PI3K. Co-inhibition of MCL-1, or BCLXL, as well as the upstream PI3K, RTK (FGF and EGF), IGF-1 mediated signaling were effective in overcoming VTX resistance. Disclosures Fonseca: Mayo Clinic in Arizona: Current Employment; Amgen: Consultancy; BMS: Consultancy; Celgene: Consultancy; Takeda: Consultancy; Bayer: Consultancy; Janssen: Consultancy; Novartis: Consultancy; Pharmacyclics: Consultancy; Sanofi: Consultancy; Merck: Consultancy; Juno: Consultancy; Kite: Consultancy; Aduro: Consultancy; OncoTracker: Consultancy, Membership on an entity's Board of Directors or advisory committees; GSK: Consultancy; AbbVie: Consultancy; Patent: Prognosticaton of myeloma via FISH: Patents & Royalties; Scientific Advisory Board: Adaptive Biotechnologies: Membership on an entity's Board of Directors or advisory committees; Caris Life Sciences: Membership on an entity's Board of Directors or advisory committees.

Multiple Myeloma (MM) is a hematological malignancy resulting in abnormal plasma cells proliferating in the bone marrow. One of the most common treatment strategies for newly diagnosed MM is the combination of induction therapy followed by autologous stem cell transplant (ASCT). Due to the underlying disease and/or the effect of the treatment regimens administered, immunosuppression is a common clinical consequence. It has been shown that ASCT recipients, treated with traditional compounds, have a reduction in the levels of antibody titers to vaccine-preventable diseases such as measles, mumps and rubella (MMR), between one and four years post-transplant. Therefore, re-vaccination is recommended at least two years after ASCT. Therapeutic options have expanded in the past decade, with the introduction of novel agents that have significantly improved MM patient outcomes; however, this may also arguably have new and different implications on a patient's immune system. In this study we sought to analyze the presence of IgG antibodies against MMR in 110 patients with MM post-ASCT. All patients received ASCT between 2014 and 2019 at Mayo Clinic Arizona. Plasma samples were collected approximately 100 days after ASCT (median 92 days) and the antibody titers were tested using the Bio-Rad MMR IgG multiplex flow immunoassay. Sample antibody index values were compared with the assay-specific calibration to determine positivity. For a control population, we utilized the results of fully vaccinated and presumptively immune healthcare workers (HCWs) that were evaluated for the presence of antibodies to measles (n=199), mumps (n=197) and rubella (n=209), using the same method, instrument and laboratory. Overall, 70% of the MM patients were positive for antibodies against measles, compared with 77.4% of HCWs. For mumps, the MM cohort had a positivity of 49.1% versus 84.8% of HCWs. Finally, rubella antibodies were found in 64.5% and 83.7% of MM and HWCs, respectively. Next, we performed testing on serial samples collected from 45 MM patients, comparing the presence of MMR antibodies pre-ASCT (median 130 days) and post-ASCT. The number of patients with positive titers detected pre- and post-transplant was unchanged for each of the three viruses. In summary, our findings do not indicate a significant reduction in relative MMR antibody levels in ASCT recipients. This suggests that earlier re-vaccination is not required post-ASCT in the era of novel MM compounds. However, further validation studies in larger cohorts are necessary prior to considering a change in current vaccination guidelines. Disclosures Braggio: DASA: Consultancy; Bayer: Other: Stock Owner; Acerta Pharma: Research Funding. Fonseca:Janssen: Consultancy; Takeda: Consultancy; Novartis: Consultancy; Bayer: Consultancy; Sanofi: Consultancy; Merck: Consultancy; Pharmacyclics: Consultancy; Amgen: Consultancy; BMS: Consultancy; Celgene: Consultancy; Juno: Consultancy; Kite: Consultancy; Aduro: Consultancy; OncoTracker: Consultancy, Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Consultancy; GSK: Consultancy; AbbVie: Consultancy; Adaptive Biotechnologies: Membership on an entity's Board of Directors or advisory committees.