UNITED STATES

SECURITIES AND EXCHANGE COMMISSION

Washington, D.C. 20549

 

FORM 8-K

  

CURRENT REPORT

Pursuant to Section 13 or 15(d) of the Securities Exchange Act of 1934

Date of Report (Date of earliest event reported): January 22, 2021

 

ATYR PHARMA, INC.

(Exact name of registrant as specified in its charter)

 

 

 Registrant’s telephone number, including area code: (858) 731-8389

Not Applicable

(Former Name or Former Address, if Changed Since Last Report)

 

Check the appropriate box below if the Form 8-K filing is intended to simultaneously satisfy the filing obligations of the registrant under any of the following provisions:

 

 

 

 

 

Securities registered pursuant to Section 12(b) of the Act:

 

 

Indicate by check mark whether the registrant is an emerging growth company as defined in Rule 405 of the Securities Act of 1933 or Rule 12b-2 of the Securities Exchange Act of 1934.

Emerging growth company     ☒

If an emerging growth company, indicate by check mark if the registrant has elected not to use the extended transition period for complying with any new or revised financial accounting standards provided pursuant to Section 13(a) of the Exchange Act.      ☒

 

 

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Item 7.01 Regulation FD Disclosure.

aTyr Pharma, Inc. (the “Company”) is participating at the Society for Laboratory Automation and Screening Digital International Conference and Exhibition (SLAS) to be held virtually from January 25-27, 2021.  The Company will be presenting a poster presentation entitled, “A Mass Spectrometry Proteomics-Based Approach to Identify Target Receptors for Novel Extracellular tRNA Synthetase Fragments,” via a prerecorded audio recording on the SLAS website.  The press release announcing the poster presentation is attached as Exhibit 99.1.  The poster presentation has been posted on the Company’s website and is attached hereto as Exhibit 99.2.

 

The information under this Item 7.01, including Exhibits 99.1 and 99.2 hereto, is being furnished herewith and shall not be deemed “filed” for the purposes of Section 18 of the Securities Exchange Act of 1934, as amended (the “Exchange Act”), or otherwise subject to the liabilities of that section, nor shall such information be deemed incorporated by reference into any filing under the Securities Act of 1933, as amended, or the Exchange Act, except as expressly set forth by specific reference in such filing.

Item 9.01 Financial Statements and Exhibits.  

(d)  Exhibits

 

 

 

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 SIGNATURE

 

Pursuant to the requirements of the Securities Exchange Act of 1934, the registrant has duly caused this report to be signed on its behalf by the undersigned hereunto duly authorized.

 

 

 

 

 

 

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Exhibit 99.1

 

aTyr Pharma to Present Poster Highlighting Research Approach for Identifying Receptor Targets for Extracellular tRNA Synthetases

Poster to be presented at the 2021 Society for Laboratory Automation and Screening Digital International Conference and Exhibition.

Research approach led to the identification of target receptors for two tRNA synthetases and further validates company’s biology platform.

SAN DIEGO – January 22, 2021 – aTyr Pharma, Inc. (Nasdaq: LIFE), a biotherapeutics company engaged in the discovery and development of innovative medicines based on novel biological pathways, today announced that it will present a poster and audio presentation at the upcoming Society for Laboratory Automation and Screening Digital International Conference and Exhibition (SLAS) to be held virtually January 25-27, 2021. The abstract and poster are available on the SLAS website.

The poster describes a mass spectrometry proteomics-based approach to identify target receptors for two distinct extracellular tRNA synthetase fragments. The utilization of this workflow, which included recombinant protein expression, flow cytometry, and receptor screening in living cells using the ligand-receptor capture TriCEPS technology, resulted in the identification of receptor targets for fragments of the tRNA synthetases AARS and DARS as well as insights into their potential biological activity in immunology, cancer and fibrosis. Furthermore, this approach can be applied more broadly to identify receptor targets of extracellular proteins and other ligands such as peptides, antibodies or viruses.

Details of the abstract and poster presentation are as follows:

Title: A mass spectrometry proteomics-based approach to identify target receptors for novel extracellular tRNA synthetase fragments
Authors: Blythe C. Dillingham, Jennifer Brasseit, Björn Hegemann, Ann L. Menefee, Justin Rahman, Zhiwen Xu, Paul Helbling, Leslie A. Nangle, Ryan A. Adams. aTyr Pharma, San Diego, CA, CSL Behring, Dualsystems Biotech.
Track: Assay Development and Screening
Date: January 25 – 27, 2021

The poster is also available on the aTyr website.

 

 

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“We are pleased to present our findings utilizing a novel approach for identifying receptor targets of extracellular proteins, and for the first time provide insight into the potential biological function of extracellular tRNA synthetases AARS and DARS,” said Sanjay S. Shukla, M.D., M.S., President and Chief Executive Officer of aTyr. “These findings further validate the relevance of our tRNA synthetase biology platform to important disease pathways and demonstrate its ability to generate new drug targets. Identifying receptor targets for these two tRNA synthetases, which are relevant to immunology, cancer and fibrosis, will help guide the development of potential new therapies in these areas of high unmet need.”

About aTyr

aTyr is a biotherapeutics company engaged in the discovery and development of innovative medicines based on novel biological pathways. aTyr’s research and development efforts are concentrated on a newly discovered area of biology, the extracellular functionality and signaling pathways of tRNA synthetases. aTyr has built a global intellectual property estate directed to a potential pipeline of protein compositions derived from 20 tRNA synthetase genes and their extracellular targets. aTyr’s primary focus is ATYR1923, a clinical-stage product candidate which binds to the neuropilin-2 receptor and is designed to down-regulate immune engagement in inflammatory lung diseases. For more information, please visit http://www.atyrpharma.com.

Forward-Looking Statements

This press release contains forward-looking statements within the meaning of the Private Securities Litigation Reform Act of 1995. Forward-looking statements are usually identified by the use of words such as “anticipates,” “believes,” “estimates,” “expects,” “intends,” “may,” “plans,” “projects,” “seeks,” “should,” “will,” and variations of such words or similar expressions. We intend these forward-looking statements to be covered by such safe harbor provisions for forward-looking statements and are making this statement for purposes of complying with those safe harbor provisions. These forward-looking statements include statements regarding potential further research and development activities related to, and potential utility of, the newly identified receptor targets, the potential therapeutic benefits and applications of our current and future product candidates; timelines and plans with respect to certain development activities (such as the timing of data from clinical trials); and certain development goals. These forward-looking statements also reflect our current views about our plans, intentions, expectations, strategies and prospects, which are based on the information currently available to us and on assumptions we have made. Although we believe that our plans, intentions, expectations, strategies and prospects, as reflected in or suggested by these forward-looking statements, are reasonable, we can give no assurance that the plans, intentions, expectations or strategies will be attained or achieved. All forward-looking statements are based on estimates and assumptions by our management that, although we believe to be reasonable, are inherently uncertain. Furthermore, actual results may differ materially from those described in these forward-looking statements and will be affected by a variety of risks and factors that are beyond our control including, without limitation, uncertainty regarding the COVID-19 pandemic, risks associated with the discovery, development and regulation of our product candidates, the risk that we or our partners may cease or delay preclinical or clinical development activities for any of our existing or future product candidates for a variety of reasons (including difficulties or delays in patient enrollment in planned clinical trials), the possibility that existing

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collaborations could be terminated early, and the risk that we may not be able to raise the additional funding required for our business and product development plans, as well as those risks set forth in our most recent Annual Report on Form 10-K, Quarterly Reports on Form 10-Q and in our other SEC filings. Except as required by law, we assume no obligation to update publicly any forward-looking statements, whether as a result of new information, future events or otherwise.

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A Mass Spectrometry Proteomics-Based Approach to Identify Target Receptors for Novel Extracellular tRNA Synthetase Fragments Purpose: To identify target receptors and determine the biological functions of novel extracellular tRNA synthetase fragments with links to immune modulation Methods: tRNA synthetase fragments were expressed recombinantly, and their binding to various human cell lines was assessed by flow cytometry. Receptor screening was completed in three cell-lines using the ligand-receptor capture technology LRC-TriCEPS followed by mass spectrometry proteomics analysis1,2. Target validation was completed by siRNA knock-down and flow cytometry, and biological function was determined using a FRET-based enzyme-inhibitor assay. Results: Utilizing this workflow, we successfully identified target cell-surface receptors for tRNA synthetase fragments and have gained insight into their previously unknown biological functions. In doing so, we have also created a novel approach which can be applied more broadly to identify receptor targets of extracellular proteins in an endogenous system. Introduction Materials and Methods Overview Acknowledgements AARS-1 and DARS-1 Binding to Human Cells ⍺-V5 AF647 Antibody V5-tagged Synthetase Fragment Unknown Receptor While canonically known for their intracellular role in protein synthesis, full-length and splice or proteolytic variants of tRNA synthetases have been found to exist in the extracellular space where they may play an immunomodulatory role. Full-length Histidyl-tRNA synthetase (HARS) has been established as a molecule present in circulation that modulates T-cell activity, and a HARS variant has been shown to bind to Neuropilin-2 and to inhibit proinflammatory chemokines and cytokines. Alanyl-tRNA Synthetase (AARS) and Aspartyl-tRNA Synthetase (DARS) are also present extracellularly and have links to immune modulation; however, their receptor targets and downstream biological function remain unknown: Auto-antibodies targeting AARS and other synthetases are present in rare anti-synthetase syndromes associated with inflammatory phenotypes such as myositis and interstitial lung disease3. Full-length DARS protein and a DARS fragment are secreted from THP-1 Macrophages when stimulated with LPS (shown to the right). Results Receptor Candidates Identified from 3 Receptor Screens Conclusions and Workflow FACS Cell Binding Assay LRC-TriCEPS Mass Spectrometry Proteomics Receptor Screen Recombinant Expression of AARS and DARS Fragments Plasma Membrane U251 cells Glioblastoma THP-1 cells M0 Macrophage Blythe C. Dillingham1, Jennifer Brasseit2, Björn Hegemann2, Ann L. Menefee1, Justin Rahman1, Zhiwen Xu1, Paul Helbling3, Leslie A. Nangle1, Ryan A. Adams1* 1. aTyr Pharma, 2. CSL Behring, 3. Dualsystems Biotech *Contact: radams@atyrpharma.com A549 cells Lung Adenocarcinoma siRNA knockdown of DARS-1 candidate TIMP-1 validated by flow cytometry: Multiple undisclosed candidate receptors were identified for both AARS-1 and DARS-1 (as indicated by numbers above). Some candidates appear to be cell-type specific, while others were identified across multiple cell-types. The candidates identified indicate potential involvement in multiple biological pathways including TGF-β and IGF signaling, oncology and fibrosis. TIMP-1 protein, a regulator of extracellular matrix (ECM) degradation, was identified as a candidate for DARS-1 and selected for follow-up experiments. DARS-1 candidate TIMP-1: Binding validation and Functional Activity AARS-1 and DARS-1 binding to unknown cell surface proteins on various cell types was tested by flow cytometry. Cells were incubated with 300 nM protein for one hour, followed by a 30-minute incubation with a fluorescently-conjugated ⍺-V5 detection antibody. Binding was quantified as median fluorescent intensity (MFI), and fold changes were calculated relative to ⍺-V5 detection antibody alone. AARS-1 and DARS-1 binding, as well as a V5-tag control, to a panel of primary and immortalized cell-lines was assessed by flow cytometry – data overview shown in table to left. Three cell-lines were selected based on cell binding data for downstream receptor screening experiments – shown in graphs above (mean ± SEM; n=2-3). Frei, Andreas P et al. “Direct identification of ligand-receptor interactions on living cells and tissues.” Nature Biotechnology vol. 30,10 (2012): 997-1001. doi:10.1038/nbt.2354 Sobotzki, Nadine et al. “HATRIC-based identification of receptors for orphan ligands” Nat Commun 9, 1519 (2018). https://doi.org/10.1038/s41467-018-03936-z Witt, Leah J et al. “The Diagnosis and Treatment of Antisynthetase Syndrome.” Clinical Pulmonary Medicine vol. 23,5 (2016): 218-226. doi:10.1097/CPM.0000000000000171 References Extracellular AARS and DARS fragments, AARS-1 and DARS-1, were successfully expressed recombinantly, and their target receptors were identified in endogenous systems using the ligand-receptor capture technology LRC-TriCEPS and mass spectrometry proteomics. The receptors identified provide new insight into the biological activity of extracellular tRNA synthetases. DARS-1 was shown to enhance TIMP-1-mediated inhibition of MMP-1 activity, indicating it may have a regulatory role in extracellular matrix remodeling. AARS-1 17 14 28 38 49 98 198 6 62 52.6 kDa DARS-1 198 17 14 28 38 49 98 6 62 19.4 kDa DARS Full-length DARS fragment V5-tagged extracellular tRNA synthetase fragments AARS-1 and DARS-1 were expressed recombinantly in Expi293 or ExpiCHO cells and purified for downstream in vitro assays. Shown to the left are SDS PAGE (blue) and anti-V5 Western blots (grey) detecting purified AARS-1 and DARS-1 under non-reduced conditions. TriCEPS 3.0 Reagent LRC-TriCEPS Receptor Screen Ligand of interest is coupled to TriCEPS reagent. Upon ligand binding, TriCEPS cross-links to the receptor targets via the hydrazide functional group. Ligand-Receptor complexes are isolated using the azide group for purification. Experiments performed in triplicate; Thresholds for significance: Log10 p-value > 2; Log2 fold change > 2 AARS-1 DARS-1 This project was supported by aTyr Pharma and CSL Behring. In addition to the listed authors, we would like to acknowledge Esther Chong and Lauren Guy from aTyr Pharma for assistance with cloning and receptor screening experiments, respectively. Cytosol Media - - + + LPS kDa 60 50 40 30 20 DARS-1 binding to THP-1 M2 Macrophages is TIMP-1 dependent Determine Biological Function Recombinant expression Test Binding to Human Cells (Flow Cytometry) Receptor Identification in Living Cells (LRC-TriCEPS/Mass Spec Proteomics) Western Blot DARS fragment secreted upon stimulation of THP-1 macrophages with LPS: DARS-1 binding to THP-1 M2 macrophages is dependent on TIMP-1 expression: (mean ± SEM; n=2) TIMP-1 regulates ECM degradation by inhibiting matrix metalloproteinases (MMPs). FRET-based MMP-1 enzyme inhibitor assay: TIMP-1 inhibits MMP-1 (IC50 = 10.2 µM). DARS-1 enhances TIMP-1-mediated inhibition of MMP-1 activity in a dose-dependent manner (shown to the right). DARS-1 enhances TIMP-1-mediated inhibition of MMP-1 Matrix Metalloproteinase-1 Activity (mean ± SEM; n=2) Workflow: 1. Validate binding (e.g. siRNA) 2. Functional assay 1 1 3 2 4 4 15 A549 U251 THP-1 9 2 2 1 A549 U251 THP-1 Fold change: ≥ 20, ≥ 3, < 3 Oncology Innate Immune cells Adaptive Immune cells Exhibit 99.2