Elena Sotillo

Stanford University | SU · Stanford Cancer Institute

Lysosome-targeting chimeras (LYTACs) are a promising therapeutic modality to drive the degradation of extracellular proteins. However, early versions of LYTAC contain synthetic glycopeptides that cannot be genetically encoded. Here we present our designs for a fully genetically encodable LYTAC (GELYTAC), making our tool compatible with integration...

Background CAR T cell therapy is a promising therapeutic modality for cancer treatment, but poor CAR T cell persistence limits efficacy in patients.¹ CAR T cells with a memory-like phenotype are associated with durable persistence in patients and response to therapy,² thereby implicating memory as an important therapeutic axis. Thus, strategies to...

Background In acute myelogenous leukemia (AML), CD93 is an appealing target for chimeric antigen receptor (CAR) T-cell therapy, offering leukemic killing without myelosuppression in preclinical studies.¹ However, CD93 CAR T-cell development is hampered by CD93 expression on healthy endothelium. Our group previously demonstrated the efficacy of an i...

Background CAR T cells have been highly effective against refractory B cell malignancies but have not demonstrated sustained antitumor effects against solid tumors. Intense effort is underway to augment the potency of CAR T cells in order to overcome the suppressive tumor microenvironment, which is associated with T cell exhaustion. Adenosine is a...

Background Chimeric Antigen Receptor (CAR) T cells are now standard of care (SOC) for some patients with B cell and plasma cell malignancies and could disrupt the therapeutic landscape of solid tumors. However, access to CAR-T cells is not adequate to meet clinical needs, in part due to high cost and long lead times for manufacturing clinical grade...

Adoptively transferred T cells and agents designed to block the CD47/SIRPα axis are promising antitumor therapeutics, which activate distinct arms of the immune system. We administered anti-CD47 (αCD47) with adoptively transferred T cells with the goal of enhancing antitumor efficacy but observed rapid macrophage-mediated clearance of T cells expre...

Adenosine (Ado) mediates immune suppression in the tumor microenvironment and exhausted CD8 ⁺ CAR T cells mediate Ado-induced immunosuppression through CD39/73-dependent Ado production. Knockout of CD39, CD73 or A2aR had modest effects on exhausted CAR T cells, whereas overexpression of Ado deaminase (ADA), which metabolizes Ado to inosine (INO), i...

Poor CAR T persistence limits CAR T cell therapies for B cell malignancies and solid tumors 1,2 . The expression of memory-associated genes such as TCF7 (protein name TCF1) is linked to response and long-term persistence in patients ³⁻⁷ , thereby implicating memory programs in therapeutic efficacy. Here, we demonstrate that the pioneer transcriptio...

Chimeric antigen receptor T cells (CAR-Ts) have demonstrated remarkable efficacy in leukemia and lymphomas but limited responses in solid tumors. We conducted a phase I trial (NCT02107963) of GD2 CAR-Ts (GD2-CAR.OX40.28.z.ICD9), demonstrating feasibility and safety of administering GD2 CAR-Ts in children and young adults with neuroblastoma and, for...

Chimeric antigen receptors (CARs) are synthetic receptors that link an extracellular tumor specific domain to intracellular signaling domains. Despite of remarkable results against refractory B cell malignancies, intense effort is underway to augment the potency of CAR T cells in order to overcome the suppressive tumor microenvironment, which is as...

Chimeric antigen receptor T (CAR T) cells are synthetically engineered to target specific tumor antigens. CD47 is a ubiquitous receptor that serves as a “don’t eat me” signal by binding to SIRPɑ on macrophages and is often over-expressed by cancer cells. Despite individual promise of CAR T and anti-CD47 therapies, neither has demonstrated clear eff...

Although chimeric antigen receptor (CAR) T cells have altered the treatment landscape for B cell malignancies, the risk of on-target, off-tumour toxicity has hampered their development for solid tumours because most target antigens are shared with normal cells1,2. Researchers have attempted to apply Boolean-logic gating to CAR T cells to prevent to...

T cells are the major arm of the immune system responsible for controlling and regressing cancers. To identify genes limiting T cell function, we conducted genome-wide CRISPR knockout screens in human chimeric antigen receptor (CAR) T cells. Top hits were MED12 and CCNC , components of the Mediator kinase module. Targeted MED12 deletion enhanced an...

Background CAR-T cell therapies have demonstrated remarkable clinical benefit in liquid tumors, but significant obstacles such as antigen loss, tumor heterogeneity, T cell exhaustion, and poor persistence limit the frequency and durability of complete responses. To overcome these challenges and expand this class of therapeutics to solid tumors, T c...

Adoptive T cell immune therapies mediate impressive clinical benefit in a fraction of patients, but anti-tumor effects are often limited by inadequate T cell potency. To identify genes limiting T cell effector function, we conducted genome-wide CRISPR knock-out screens in human primary CAR-T cells. The top hits were MED12 and CCNC , components of t...

Introductory paragraph While CAR T cells have altered the treatment landscape for B cell malignancies, the risk of on-target, off-tumor toxicity has hampered their development for solid tumors because most target antigens are shared with normal cells 1,2 . Researchers have attempted to apply Boolean logic gating to CAR T cells to prevent on-target,...

Chimeric Antigen Receptor (CAR) T cell therapy has resulted in remarkable clinical outcomes in the context of acute and chronic lymphoblastic leukemia, but remains unsuccessful in the treatment of solid tumors. One reason for this failure is thought to be T cell dysfunction or exhaustion promoted by suppressive soluble factors within the tumor micr...

Chimeric antigen receptor (CAR) T cell therapy has revolutionized oncology through engineered targeting of antigens on previously untreatable cancers. However, less than half of patients on CAR T cell therapy experience long-term disease control, with better outcomes observed in pediatric compared to adult populations. Senescent T cells have been s...

Adoptive T cell immune therapies mediate impressive clinical benefit in a fraction of patients, but anti-tumor effects are often limited by inadequate T cell potency. To identify genes that limit T cell effector function, we conducted genome-wide CRISPR knock-out screens in human primary CAR-T cells. The top hits were components of the CDK8 kinase...

Regulatable CAR platforms could circumvent toxicities associated with CAR-T therapy, but existing systems have shortcomings including leakiness and attenuated activity. Here, we present SNIP CARs, a protease-based platform for regulating CAR activity using an FDA-approved small molecule. Design iterations yielded CAR-T cells that manifest full func...

The disialoganglioside GD2 is overexpressed on several solid tumors, and monoclonal antibodies targeting GD2 have substantially improved outcomes for children with high-risk neuroblastoma. However, approximately 40% of patients with neuroblastoma still relapse, and anti-GD2 has not mediated significant clinical activity in any other GD2⁺ malignancy...

Pediatric cancers often mimic fetal tissues and express proteins normally silenced postnatally that could serve as immune targets. We developed T cells expressing chimeric antigen receptors (CARs) targeting glypican-2 (GPC2), a fetal antigen expressed on neuroblastoma (NB) and several other solid tumors. CARs engineered using standard designs contr...

Despite impressive response to front-line treatment, relapsed or refractory B-cell acute lymphoblastic leukemia (r/r B-ALL) remains one of the most common causes of death in children with cancer. In pediatric and adult r/r B-ALL, CD19 CAR T cell (CART19) therapy has shown impressive results with over 80% of patients achieving complete remission. Ho...

Chimeric antigen receptor (CAR) T cells hold promise for the treatment of acute myeloid leukemia (AML), but optimal targets remain to be defined. We demonstrate that CD93 CAR T cells engineered from a novel humanized CD93-specific binder potently kill AML in vitro and in vivo but spare hematopoietic stem and progenitor cells (HSPC). No toxicity is...

MYC oncoprotein promotes cell proliferation and serves as the key driver in many human cancers; therefore, considerable effort has been expended to develop reliable pharmacological methods to suppress its expression or function. Despite impressive progress, MYC-targeting drugs have not reached the clinic. Recent advances suggest that within a limit...

Significance T cell exhaustion is a major barrier to cancer immunotherapy. T cell exhaustion is the state of T cell dysfunction after chronic stimulation, and recent studies indicate that exhaustion is epigenetically controlled and associated with unique chromatin profiles. This work reports the genome-wide map of active DNA regulatory elements and...

Background: CAR T-cells targeting solid tumor antigens have not yet demonstrated similar success as seen for CD19, BCMA and CD22 in B-cell malignancies. Truly tumor-specific antigens are rare, yet pediatric solid tumors manifest stalled fetal developmental programs and often overexpress oncofetal antigens, normally restricted to prenatal tissues. A...

Resistance to CD19-directed immunotherapies in lymphoblastic leukemia has been attributed, among other factors, to several aberrant CD19 pre-mRNA splicing events, including recently reported excision of a cryptic intron embedded within CD19 exon 2. While “exitrons” are known to exist in hundreds of human transcripts, we discovered, using reporter a...

CAR-T cells rest to get back in the race Chimeric antigen receptor (CAR)–T cells, which are engineered to target specific tumor antigens, are increasingly used as an immunotherapy. CAR-T cells have shown promising results in patients, particularly in hematologic cancers, but their anticancer activity can be limited by the onset of exhaustion and th...

Dysfunction in T cells limits the efficacy of cancer immunotherapy. We profiled the epigenome, transcriptome, and enhancer connectome of exhaustion-prone GD2-targeting HA-28z chimeric antigen receptor (CAR) T cells and control CD19-targeting CAR T cells, which present less exhaustion-inducing tonic signaling, at multiple points during their ex vivo...

The disialoganglioside GD2 is consistently overexpressed in neuroblastoma and osteosarcoma, and is variably expressed in other sarcomas, gliomas, neuroendocrine tumors, and epithelial cancers. Anti-GD2 antibodies have improved the survival rates of patients with neuroblastoma only when administered as part of intense chemotherapy-based cytotoxic re...

To identify disease-relevant T cell receptors (TCRs) with shared antigen specificity, we analyzed 778,938 TCRβ chain sequences from 178 non-small cell lung cancer patients using the GLIPH2 (grouping of lymphocyte interactions with paratope hotspots 2) algorithm. We identified over 66,000 shared specificity groups, of which 435 were clonally expande...

Introduction: Acute myeloid leukemia (AML) is the most prevalent acute leukemia in the United States, accounting for more than 11,000 deaths each year and with a 5-year overall survival rate of less than 30%. With the exception of Gemtuzumab ozogamycin, an anti-CD33 antibody drug conjugate, the landmark success of immunotherapy in other hematologic...

CD19 CAR T cells have revolutionized the treatment of relapsed and refractory (R/R) large B cell lymphomas (LBCL), mediating durable complete responses in approximately 40-50% of patients. Besides a loss or decrease in CD19 expression, no studies have identified tumor specific factors driving inherent or acquired resistance to CAR T cells in LBCL....

Atypical teratoid/rhabdoid tumors (ATRT) are highly malignant embryonal brain tumors, arising in very young children, and are associated with inactivation of SMARCB1. The prognosis is extremely poor. Hence new treatments are urgently needed. We show that 100% of ATRT express B7-H3 and residual SMARCA4 SWI/SNF activity drives B7-H3 expression. Furth...

The use of anti-GD2 antibodies for neuroblastoma (NBL) has resulted in enhanced survival, but many patients still relapse and ultimately die of their disease. Additionally, despite expression of GD2 on osteosarcoma (OS), anti-GD2 antibodies have not proven widely effective in that disease. Enhancing the efficacy of anti-GD2 antibodies could result...

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Atypical teratoid/rhabdoid tumors (ATRTs) typically arise in the central nervous system (CNS) of children under 3 years of age. Despite intensive multimodal therapy (surgery, chemotherapy and, if age permits, radiotherapy), median survival is 17 months1,2. We show that ATRTs robustly express B7-H3/CD276 that does not result from the inactivating mu...

Insufficient reactivity against cells with low antigen density has emerged as an important cause of chimeric antigen receptor (CAR) T-cell resistance. Little is known about factors that modulate the threshold for antigen recognition. We demonstrate that CD19 CAR activity is dependent upon antigen density and that the CAR construct in axicabtagene c...

T cell exhaustion limits immune responses against cancer and is a major cause of resistance to CAR- T cell therapeutics. Using a model wherein tonic CAR signaling induces hallmark features of exhaustion, we employed a drug-regulatable CAR to test the impact of transient cessation of receptor signaling (i.e. "rest") on the development and maintenanc...

Chimeric antigen receptor (CAR) T cells mediate anti-tumour effects in a small subset of patients with cancer1,2,3, but dysfunction due to T cell exhaustion is an important barrier to progress4,5,6. To investigate the biology of exhaustion in human T cells expressing CAR receptors, we used a model system with a tonically signaling CAR, which induce...

CD19-directed chimeric antigen receptor T cell (CAR-T) therapy has shown impressive results in children and adults with relapsed or refractory B-ALL or diffuse large B-cell lymphoma. However, 30 - 70% of initial responders will eventually relapse with CD19 antigen loss (CD19Neg) (Maude SL, et al. N Engl J Med. 2018). To avoid CD19Neg relapse, patie...

Therapeutic targeting of initiating oncogenes is the mainstay of precision medicine. Considerable efforts have been expended toward silencing MYC, which drives many human cancers including Burkitt lymphomas (BL). Yet, the effects of MYC silencing on standard-of-care therapies are poorly understood. Here we found that inhibition of MYC transcription...

Background: Ewing sarcoma (EWS) manifests one of the lowest somatic mutation rates of any cancer, leading to a scarcity of druggable mutations and neoantigens. Immunotherapeutics targeting differentially expressed cell surface antigens could provide therapeutic benefit for such tumors. Pregnancy-associated plasma protein A (PAPP-A) is a cell membr...

CAR T cells mediate antitumor effects in a small subset of cancer patients, but dysfunction due to T cell exhaustion is an important barrier to progress. To investigate the biology of exhaustion in human T cells expressing CAR receptors, we used a model system employing a tonically signaling CAR, which induces hallmarks of exhaustion described in o...

Key Points Dasatinib potently and reversibly suppresses CAR-T cell cytotoxicity, cytokine secretion, and proliferation. Dasatinib could be repurposed as a safety switch to mitigate CAR-mediated toxicity in patients.

Purpose: Patients with relapsed pediatric solid tumors and CNS malignancies have few therapeutic options and frequently die of their disease. Chimeric antigen receptor (CAR) T cells have shown tremendous success in treating relapsed pediatric acute lymphoblastic leukemia, but this has not yet translated to treating solid tumors. This is partially...

Target antigen density has emerged as a major factor influencing the potency of CAR T cells. Our laboratory has demonstrated that the activity of numerous CARs is highly dependent on target antigen density (Walker et al., Mol Ther, 2017), and high complete response rates in a recent trial of CD22 CAR T cells for B-ALL were tempered by frequent rela...

Patients with high risk neuroblastoma have a poor prognosis and survivors are often left with debilitating long term sequelae from treatment. Even after integration of anti-GD2 monoclonal antibody therapy into standard, upftont protocols, 5-year overall survival rates are only about 50%. The success of anti-GD2 therapy has proven that immunotherapy...

We have previously described a mechanism of acquired resistance of B-cell acute lymphoblastic leukemia to CD19-directed chimeric antigen receptor T-cell (CART) immunotherapy. It was based on in-frame insertions in or skipping of CD19 exon 2. To distinguish between epitope loss and defects in surface localization, we used retroviral transduction and...

In patients with chronic viral infection or cancer, continuous antigen exposure results in T cell exhaustion, which is characterized by sustained co-expression of multiple inhibitory receptors (ex. PD-1, TIM-3, LAG-3) a hierarchical loss of effector function. Human T cells expressing a high-affinity anti-GD2 chimeric antigen receptor (CAR, HA.28z)...

Chimeric antigen receptor (CAR) T cell therapy has revolutionized the treatment of B cell leukemia. However, application of CAR T cell therapy in solid tumors has been disappointing. We believe the development of T cell exhaustion, a phenomenon well described in chronic viral infection, is a major barrier to widespread success of CAR T cell therapy...

T cell receptor therapy; primary mediastinal large B cell lymphoma; rituximab; brentuximab

Background: CD19 is a near-universal surface marker of B-cell malignancies. Therefore, it is regarded as a target of choice for various immunotherapies, such as bi-specific T-cell engagers and chimeric antigen receptor-armed T-cells. Although the latter have proven remarkably effective in treating B-cell acute lymphoblastic leukemia (B-ALL), relaps...

CD19 is expressed broadly on the surface of B-cells during normal development and malignant growth, making it a good target for immunotherapy. While immunotherapies targeting CD19 have had great success against pediatric B-cell acute lymphoblastic leukemia (B-ALL), relapses lacking the CD19 epitope still occur (Maude et al., 2014). We have discover...

Unlabelled: The CD19 antigen, expressed on most B-cell acute lymphoblastic leukemias (B-ALL), can be targeted with chimeric antigen receptor-armed T cells (CART-19), but relapses with epitope loss occur in 10% to 20% of pediatric responders. We detected hemizygous deletions spanning the CD19 locus and de novo frameshift and missense mutations in e...

The Myc proto-oncogene is known to regulate p53-dependent apoptosis through the Myc -> ARF -| MDM2 -| p53 pathway. We had previously shown that this pathway could be exploited to increase sensitivity to anti-cancer drugs. Specifically, small increases in Myc levels obtained by inhibiting the Myc-targeting microRNA-34a resulted in enhanced apoptotic...

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA CD19 is expressed on most B-cell neoplasms, including acute lymphoblastic leukemias (B-ALL). Chimeric antigen receptor (CAR)-armed T cells targeting CD19 (CART-19) yield complete remission rate of 70-90% in B-ALL patients; the bispecific anti-CD19/CD3 antibody blinatu...

The c-MYC oncogene deregulation is a common trait in human cancers, with programmed cell death (or apoptosis) among Myc-regulated events. Myc is well-known to induce apoptosis in a p53-dependent manner, via the Myc→ARF ┤MDM2 ┤p53 pathway. We had previously shown that this pathway couldbe exploited to increase sensitivity to anti-cancer drugs. Speci...

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL For patients with high-risk neuroblastoma (NB) survival remains below 40%, and survivors suffer from life-long treatment-related disabilities. This necessitates the development of oncogene-specific targeted therapies. We and others observed that small-molecule inhibitors...

It has been a decade since scientists realized that microRNAs (miRNAs) are not an oddity invented by worms to regulate gene expression at post-transcriptional levels. Rather, many of these 21-22-nucleotide-short RNAs exist in invertebrates and vertebrates alike and some of them are in fact highly conserved. miRNAs are now recognized as an important...

Transcription of genomic loci containing protein-coding genes often yields not only cognate mRNAs but also assorted noncoding RNAs (ncRNAs), which typically map in the vicinity of transcription start sites. A new study shows that far from being random byproducts of gene expression, many long ncRNAs (lncRNAs) are synthesized in a coordinate fashion...

Downregulation of microRNA-34a by Myc is known to be essential for tumorigenesis and improve tumor-cell survival. Conversely, upregulation of miR-34a by p53 is thought to enhance its acetylation and activity and contribute to the pro-apoptotic effects of this tumor suppressor. We sought to determine whether restoration of miR-34a levels in B-lympho...

Pocket proteins negatively regulate transcription of E2F-dependent genes and progression through the G0/G1 transition and the cell cycle restriction point in G1. Pocket protein repressor activities are inactivated via phosphorylation at multiple Pro-directed Ser/Thr sites by the coordinated action of G1 and G1/S cyclin-dependent kinases. These phos...

Hepatocellular carcinoma is the third leading cause of cancer mortality worldwide; current chemotherapeutic interventions for this disease are largely ineffective. The retinoblastoma tumor suppressor (RB) is functionally inactivated at relatively high frequency in hepatocellular carcinoma and hepatoma cell lines. Here, we analyzed the ability of CD...

Quiescent: From Latin quies, referring to a state of being at rest, dormant, inactive, quiet, still (Merriam-Webster, 2009, Online Dictionary: http://www.merriam-webster.com/dictionary/quiescent). This term refers to a state of dormancy as opposed to a proliferative state. However, quiescent cells are in any other regard metabolically active. In ma...

We have previously shown that SV40 small t antigen (st) cooperates with deregulated cyclin E to activate CDK2 and bypass quiescence in normal human fibroblasts (NHF). Here we show that st expression in serum-starved and density-arrested NHF specifically induces up-regulation and loading of CDC6 onto chromatin. Coexpression of cyclin E results in fu...

Cyclin E overexpression is observed in multiple human tumors and linked to poor prognosis. We have previously shown that ectopic expression of cyclin E is sufficient to induce mitogen-independent cell cycle entry in a variety of tumor/immortal cell lines. Here we have investigated the rate-limiting step leading to cell cycle entry in quiescent norm...

To explore the possible association/s of the first reported tumour necrosis factor (TNF-alphaTNF-) alpha promoter gene polymorphisms -308, -238, -376 and -163 (G-->A) with systemic (SoJIA) and oligoarticular subtypes of juvenile idiopathic arthritis (JIA); and to test the association between these polymorphisms and the class I/class II HLA alleles...

It is thought that G(1) cyclin/CDK mediated phosphorylation of pocket proteins from mid G(1) to mitosis is reversed via dephosphorylation in mitosis. We examined the mechanisms involved in the unexpectedly rapid dephosphorylation of the pocket proteins induced via inhibition of cellular protein synthesis by cycloheximide (CHX) as well as direct inh...

The presence of telomerase activity has been analyzed in almost all tumor types and tumor-derived cell lines. However, there are very few studies that focus on the presence of telomerase activity in bone tumors, and most of them report analysis on very few samples or bone-derived cell lines. The objective of this study was to analyze the telomere l...

The authors' objectives were to compare height at diagnosis of children with bone tumors with that of Spanish reference children; to analyze the frequency of the genotypes for the polymorphisms of the vitamin D receptor (VDR), estrogen receptor (ER), and collagen Ialpha1 (COLIalpha1) genes in patients and in healthy controls; and to test the relati...

To analyze the genetic and epigenetic alterations affecting the RB1, TP53, p16INK4, and p21WAF1 tumor suppressor genes, loss of heterozygosity (LOH) at 3q and 18q, and the clinical variables of a series of Spanish children with osteosarcoma. These genetic changes were tested for an association with prognosis. Peripheral blood samples and clinical d...