Try a new search

Format these results:

Searched for:

person:hattot01

in-biosketch:yes

Total Results:

43


Binding mode-guided development of high-performance antibodies targeting site-specific posttranslational modifications

Riso, Mariapia; Shah, Rohan N; Koide, Akiko; Ruthenburg, Alexander J; Koide, Shohei; Hattori, Takamitsu
Posttranslational modifications (PTMs) of proteins play critical roles in regulating many cellular events. Antibodies targeting site-specific PTMs are essential tools for detecting and enriching PTMs at sites of interest. However, fundamental difficulties in molecular recognition of both PTM and surrounding peptide sequence have hindered the efficient generation of highly sequence-specific anti-PTM antibodies. Furthermore, the widespread use of potentially inconsistent, nonrenewable, and molecularly undefined antibodies presents experimental challenges thought to contribute to the reproducibility problem in biomedical research. In this study, we describe the binding mode-guided development of a platform that efficiently generates potent and selective recombinant antibodies to PTMs that are molecularly defined and renewable. Our platform is built on our previous discovery of an unconventional binding mode of anti-PTM antibodies, antigen clasping, where two antigen binding sites cooperatively sandwich a single antigen, creating extensive interactions with the antigen and leading to high selectivity and potency. We designed the platform that generates clasping antibodies with two distinct binding units, resulting in efficient generation of antibodies to a set of trimethylated histone H3 with high levels of specificity and affinity. Performance comparison in chromatin immunoprecipitation, a common application in epigenomics, revealed that a clasping antibody to trimethylated histone H3 at lysine 27 exhibited superior specificity to a widely used conventional antibody and captured symmetric and asymmetric nucleosomes in a less biased manner. We further generated clasping antibodies to phosphotyrosine antigens by using the same principle. These results suggest the broad applicability of our platform to generating high-performance clasping antibodies to diverse PTMs.
PMCID:11725865
PMID: 39793060
ISSN: 1091-6490
CID: 5775412

Selective targeting of oncogenic hotspot mutations of the HER2 extracellular domain

Bang, Injin; Hattori, Takamitsu; Leloup, Nadia; Corrado, Alexis; Nyamaa, Atekana; Koide, Akiko; Geles, Ken; Buck, Elizabeth; Koide, Shohei
Oncogenic mutations in the extracellular domain (ECD) of cell-surface receptors could serve as tumor-specific antigens that are accessible to antibody therapeutics. Such mutations have been identified in receptor tyrosine kinases including HER2. However, it is challenging to selectively target a point mutant, while sparing the wild-type protein. Here we developed antibodies selective to HER2 S310F and S310Y, the two most common oncogenic mutations in the HER2 ECD, via combinatorial library screening and structure-guided design. Cryogenic-electron microscopy structures of the HER2 S310F homodimer and an antibody bound to HER2 S310F revealed that these antibodies recognize the mutations in a manner that mimics the dimerization arm of HER2 and thus inhibit HER2 dimerization. These antibodies as T cell engagers selectively killed a HER2 S310F-driven cancer cell line in vitro, and in vivo as a xenograft. These results validate HER2 ECD mutations as actionable therapeutic targets and offer promising candidates toward clinical development.
PMID: 39438724
ISSN: 1552-4469
CID: 5739862

Molecular basis for antibody recognition of multiple drug-peptide/MHC complexes

Maso, Lorenzo; Rajak, Epsa; Bang, Injin; Koide, Akiko; Hattori, Takamitsu; Neel, Benjamin G; Koide, Shohei
The HapImmuneTM platform exploits covalent inhibitors as haptens for creating major histocompatibility complex (MHC)-presented tumor-specific neoantigens by design, combining targeted therapies with immunotherapy for the treatment of drug-resistant cancers. A HapImmune antibody, R023, recognizes multiple sotorasib-conjugated KRAS(G12C) peptides presented by different human leukocyte antigens (HLAs). This high specificity to sotorasib, coupled with broad HLA-binding capability, enables such antibodies, when reformatted as T cell engagers, to potently and selectively kill sotorasib-resistant KRAS(G12C) cancer cells expressing different HLAs upon sotorasib treatment. The loosening of HLA restriction could increase the patient population that can benefit from this therapeutic approach. To understand the molecular basis for its unconventional binding capability, we used single-particle cryogenic electron microscopy to determine the structures of R023 bound to multiple sotorasib-peptide conjugates presented by different HLAs. R023 forms a pocket for sotorasib between the VH and VL domains, binds HLAs in an unconventional, angled way, with VL making most contacts with them, and makes few contacts with the peptide moieties. This binding mode enables the antibody to accommodate different hapten-peptide conjugates and to adjust its conformation to different HLAs presenting hapten-peptides. Deep mutational scanning validated the structures and revealed distinct levels of mutation tolerance by sotorasib- and HLA-binding residues. Together, our structural information and sequence landscape analysis reveal key features for achieving MHC-restricted recognition of multiple hapten-peptide antigens, which will inform the development of next-generation therapeutic antibodies.
PMID: 38781214
ISSN: 1091-6490
CID: 5654922

The expression profile and tumorigenic mechanisms of CD97 (ADGRE5) in glioblastoma render it a targetable vulnerability

Ravn-Boess, Niklas; Roy, Nainita; Hattori, Takamitsu; Bready, Devin; Donaldson, Hayley; Lawson, Christopher; Lapierre, Cathryn; Korman, Aryeh; Rodrick, Tori; Liu, Enze; Frenster, Joshua D; Stephan, Gabriele; Wilcox, Jordan; Corrado, Alexis D; Cai, Julia; Ronnen, Rebecca; Wang, Shuai; Haddock, Sara; Sabio Ortiz, Jonathan; Mishkit, Orin; Khodadadi-Jamayran, Alireza; Tsirigos, Aris; Fenyö, David; Zagzag, David; Drube, Julia; Hoffmann, Carsten; Perna, Fabiana; Jones, Drew R; Possemato, Richard; Koide, Akiko; Koide, Shohei; Park, Christopher Y; Placantonakis, Dimitris G
Glioblastoma (GBM) is the most common and aggressive primary brain malignancy. Adhesion G protein-coupled receptors (aGPCRs) have attracted interest for their potential as treatment targets. Here, we show that CD97 (ADGRE5) is the most promising aGPCR target in GBM, by virtue of its de novo expression compared to healthy brain tissue. CD97 knockdown or knockout significantly reduces the tumor initiation capacity of patient-derived GBM cultures (PDGCs) in vitro and in vivo. We find that CD97 promotes glycolytic metabolism via the mitogen-activated protein kinase (MAPK) pathway, which depends on phosphorylation of its C terminus and recruitment of β-arrestin. We also demonstrate that THY1/CD90 is a likely CD97 ligand in GBM. Lastly, we show that an anti-CD97 antibody-drug conjugate selectively kills tumor cells in vitro. Our studies identify CD97 as a regulator of tumor metabolism, elucidate mechanisms of receptor activation and signaling, and provide strong scientific rationale for developing biologics to target it therapeutically in GBM.
PMID: 37938973
ISSN: 2211-1247
CID: 5590372

Exploring switch II pocket conformation of KRAS(G12D) with mutant-selective monobody inhibitors

Akkapeddi, Padma; Hattori, Takamitsu; Khan, Imran; Glasser, Eliezra; Koide, Akiko; Ketavarapu, Gayatri; Whaby, Michael; Zuberi, Mariyam; Teng, Kai Wen; Lefler, Julia; Maso, Lorenzo; Bang, Injin; Ostrowski, Michael C; O'Bryan, John P; Koide, Shohei
The G12D mutation is among the most common KRAS mutations associated with cancer, in particular, pancreatic cancer. Here, we have developed monobodies, small synthetic binding proteins, that are selective to KRAS(G12D) over KRAS(wild type) and other oncogenic KRAS mutations, as well as over the G12D mutation in HRAS and NRAS. Crystallographic studies revealed that, similar to other KRAS mutant-selective inhibitors, the initial monobody bound to the S-II pocket, the groove between switch II and α3 helix, and captured this pocket in the most widely open form reported to date. Unlike other G12D-selective polypeptides reported to date, the monobody used its backbone NH group to directly recognize the side chain of KRAS Asp12, a feature that closely resembles that of a small-molecule inhibitor, MTRX1133. The monobody also directly interacted with H95, a residue not conserved in RAS isoforms. These features rationalize the high selectivity toward the G12D mutant and the KRAS isoform. Structure-guided affinity maturation resulted in monobodies with low nM K
PMCID:10334749
PMID: 37399416
ISSN: 1091-6490
CID: 5536822

In vivo metabolomics identifies CD38 as an emergent vulnerability in LKB1 -mutant lung cancer

Deng, Jiehui; Peng, David H; Fenyo, David; Yuan, Hao; Lopez, Alfonso; Levin, Daniel S; Meynardie, Mary; Quinteros, Mari; Ranieri, Michela; Sahu, Soumyadip; Lau, Sally C M; Shum, Elaine; Velcheti, Vamsidhar; Punekar, Salman R; Rekhtman, Natasha; Dowling, Catríona M; Weerasekara, Vajira; Xue, Yun; Ji, Hongbin; Siu, Yik; Jones, Drew; Hata, Aaron N; Shimamura, Takeshi; Poirier, John T; Rudin, Charles M; Hattori, Takamitsu; Koide, Shohei; Papagiannakopoulos, Thales; Neel, Benjamin G; Bardeesy, Nabeel; Wong, Kwok-Kin
UNLABELLED:. Surprisingly, compared with other genetic subsets, murine and human LKB1-mutant NSCLC show marked overexpression of the NAD+-catabolizing ectoenzyme, CD38 on the surface of tumor cells. Loss of LKB1 or inactivation of Salt-Inducible Kinases (SIKs)-key downstream effectors of LKB1- induces CD38 transcription induction via a CREB binding site in the CD38 promoter. Treatment with the FDA-approved anti-CD38 antibody, daratumumab, inhibited growth of LKB1-mutant NSCLC xenografts. Together, these results reveal CD38 as a promising therapeutic target in patients with LKB1 mutant lung cancer. SIGNIFICANCE/CONCLUSIONS:tumor suppressor of lung adenocarcinoma patients and are associated with resistance to current treatments. Our study identified CD38 as a potential therapeutic target that is highly overexpressed in this specific subtype of cancer, associated with a shift in NAD homeostasis.
PMCID:10153147
PMID: 37131623
ISSN: 2692-8205
CID: 5507602

Creating MHC-restricted neoantigens with covalent inhibitors that can be targeted by immune therapy

Hattori, Takamitsu; Maso, Lorenzo; Araki, Kiyomi Y; Koide, Akiko; Hayman, James; Akkapeddi, Padma; Bang, Injin; Neel, Benjamin G; Koide, Shohei
Intracellular oncoproteins can be inhibited with targeted therapy, but responses are not durable. Immune therapies can be curative, but most oncogene-driven tumors are unresponsive to these agents. Fragments of intracellular oncoproteins can act as neoantigens presented by the major histocompatibility complex (MHC) but recognizing minimal differences between oncoproteins and their normal counterparts is challenging. We have established a platform technology that exploits hapten-peptide conjugates generated by covalent inhibitors to create distinct neoantigens that selectively mark cancer cells. Using the FDA-approved covalent inhibitors sotorasib and osimertinib, we developed "HapImmuneTM" antibodies that bind to drug-peptide conjugate/MHC complexes but not to the free drugs. A HapImmuneTM-based bispecific T cell engager selectively and potently kills sotorasib-resistant lung cancer cells upon sotorasib treatment. Notably, it is effective against KRASG12C mutant cells with different HLA supertypes, HLA-A*02 and A*03/11, suggesting loosening of MHC restriction. Our strategy creates targetable neoantigens by design, unifying targeted and immune therapies.
PMID: 36250888
ISSN: 2159-8290
CID: 5360222

DEFINING A NOVEL ROLE FOR CD97 IN REGULATING GBM GLYCOLYTIC METABOLISM [Meeting Abstract]

Ravn-Boess, N; Bhowmick, N; Hattori, T; Prakash, V; Stephan, G; Frenster, J; Bready, D; Lawson, C; Wilcox, J; Placantonakis, D
Glioblastoma (GBM) is the most common and aggressive primary brain malignancy. Despite multimodal therapy, disease recurrence is inevitable. To identify novel vulnerabilities of GBM, we performed an arrayed CRISPR/ Cas9 screen against select adhesion G protein-coupled receptors (aGPCRs), many of which we found to be de novo expressed in GBM. Knockout of CD97, previously implicated in GBM cell migration, produced the most striking proliferative disadvantage in patient-derived GBM cultures (PDGC) among aGPCRs tested. We found high CD97 surface expression in all our PDGCs, while levels remained nearly undetectable in non-neoplastic brain cells, confirming that CD97 is de novo expressed in GBM. Upon shRNAmediated knockdown of CD97 in PDGCs from all three TCGA transcriptional subtypes, we observed reduced proliferation, as measured by cell cycle analysis. Notably, CD97 knockdown also significantly reduced tumorsphere formation capacity as measured by limiting dilution assays; an effect that was partially rescued upon CD97 overexpression. To elucidate mechanisms of action of CD97, we performed RNA-sequencing and GO pathway enrichment analysis from PDGCs following CD97 knockdown. The top downregulated pathways involved glycolytic metabolism, specifically involving many genes relevant for glucose-6-phosphate (G6P) and fructose- 6-phosphate (F6P) processing. Indeed, when we measured metabolite levels under both steady-state and flux conditions using mass spectrometry, we observed an accumulation of G6P and a depletion of most downstream glycolytic and Krebs cycle metabolites upon CD97 knockdown. Furthermore, Seahorse metabolic assays revealed deficits in both glycolytic metabolism and oxygen consumption. We aim to interrogate the activity of specific glycolytic enzymes involved in processing G6P and F6P, pinpointing how these are influenced by CD97 signaling pathways (MAPK or Akt). Overall; our studies suggest a novel role of CD97 in regulating GBM metabolism (Warburg effect), and provide a strong scientific rationale for developing biologics to target CD97 which appears to be universally and de novo expressed in GBM
EMBASE:639940023
ISSN: 1523-5866
CID: 5513282

Monobody adapter for functional antibody display on nanoparticles for adaptable targeted delivery applications

Albert, C; Bracaglia, L; Koide, A; DiRito, J; Lysyy, T; Harkins, L; Edwards, C; Richfield, O; Grundler, J; Zhou, K; Denbaum, E; Ketavarapu, G; Hattori, T; Perincheri, S; Langford, J; Feizi, A; Haakinson, D; Hosgood, S A; Nicholson, M L; Pober, J S; Saltzman, W M; Koide, S; Tietjen, G T
Vascular endothelial cells (ECs) play a central role in the pathophysiology of many diseases. The use of targeted nanoparticles (NPs) to deliver therapeutics to ECs could dramatically improve efficacy by providing elevated and sustained intracellular drug levels. However, achieving sufficient levels of NP targeting in human settings remains elusive. Here, we overcome this barrier by engineering a monobody adapter that presents antibodies on the NP surface in a manner that fully preserves their antigen-binding function. This system improves targeting efficacy in cultured ECs under flow by >1000-fold over conventional antibody immobilization using amine coupling and enables robust delivery of NPs to the ECs of human kidneys undergoing ex vivo perfusion, a clinical setting used for organ transplant. Our monobody adapter also enables a simple plug-and-play capacity that facilitates the evaluation of a diverse array of targeted NPs. This technology has the potential to simplify and possibly accelerate both the development and clinical translation of EC-targeted nanomedicines.
PMCID:9553936
PMID: 36220817
ISSN: 2041-1723
CID: 5352032

High-valency anti-CD99 antibodies toward the treatment of T cell acute lymphoblastic leukemia

Romero, Larizbeth A; Hattori, Takamitsu; A E Ali, Mohamed; Ketavarapu, Gayatri; Koide, Akiko; Park, Christopher Y; Koide, Shohei
T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive form of leukemia that currently requires intensive chemotherapy. While childhood T-ALL is associated with high cure rates, adult T-ALL is not, and both are associated with significant short- and long-term morbidities. Thus, less toxic and effective strategies to treat T-ALL are needed. CD99 is overexpressed on T-ALL blasts at diagnosis and at relapse. Although targeting CD99 with cytotoxic antibodies has been proposed, the molecular features required for their activity are undefined. We identified human antibodies that selectively bound to the extracellular domain human CD99 and the most potent clone, 10A1, shared an epitope with a previously described cytotoxic IgM antibody. We engineered clone 10A1 in bivalent, trivalent, tetravalent, and dodecavalent formats. Increasing the antibody valency beyond two had no effects on binding to T-ALL cells. In contrast, a valency of ≥3 was required for cytotoxicity, suggesting a mechanism of action in which an antibody clusters ≥3 CD99 molecules to induce cytotoxicity. We developed a human IgG-based tetravalent version of 10A1 that exhibited cytotoxic activity to T-ALL cells but not to healthy peripheral blood cells. The crystal structure of the 10A1 Fab in complex with a CD99 fragment revealed that the antibody primarily recognizes a proline-rich motif (PRM) of CD99 in a manner reminiscent of SH3-PRM interactions. This work further validates CD99 as a promising therapeutic target in T-ALL and defines a pathway toward the development of a selective therapy against T-ALL.
PMID: 34958778
ISSN: 1089-8638
CID: 5108052