Common sharesPre-funded warrants to purchase Common shares Definium Therapeutics, Inc. We are offering $500,000,000 of our common shares, without par value, at a public offering priceof $per share and, to certain investors, pre-funded warrants to purchase common shares.The purchase price of each pre-funded warrant will equal the price per share at which thecommon shares are being sold to the public in this offering, minus $, which is the exerciseprice of each pre-funded warrant. This prospectus supplement also relates to the offering of thecommon shares issuable upon exercise of the pre-funded warrants. Our common shares are listed on the Nasdaq Global Select Market (“Nasdaq”) under the symbol“DFTX”. The last reported sale price of our common shares on Nasdaq on June18, 2026 was$24.48 per share. We do not intend to list the pre-funded warrants on Nasdaq, any other nationalsecurities exchange or any other nationally recognized trading system. Investing in our securities involves a high degree of risk. See “Risk Factors” beginning onpageS-10of this prospectus supplement, page 8 of the accompanying prospectus andunder similar headings in the documents incorporated by reference into this prospectussupplement and the accompanying prospectus. We are an “emerging growth company” under applicable Securities and Exchange Commissionrules and are subject to reduced public company reporting requirements. See “ProspectusSupplement Summary—Implications of Being an Emerging Growth Company.” Neither the Securities and Exchange Commission nor any state or other securitiescommission has approved or disapproved of these securities or determined if thisprospectus supplement or the accompanying prospectus is truthful or complete. Anyrepresentation to the contrary is a criminal offense. We have granted the underwriters an option to purchase up to an additional $75,000,000 ofcommon shares from us at the public offering price, less the underwriting discounts andcommissions payable by us. This option is exercisable, in whole or in part, for a period of 30daysfollowing the date of this prospectus supplement. If the underwriters exercise the option in full,the total underwriting discounts and commissions payable by us will be approximately $,and the total proceeds to us, before expenses, will be approximately $.The information in this preliminary prospectus supplement is not complete and may be changed. A registration statement relating to thesesecurities has been filed with the Securities and Exchange Commission and is effective. This preliminary prospectus supplement and theaccompanying prospectus are not an offer to sell these securities and they are not soliciting an offer to buy these securities in anyjurisdiction where the offer or sale is not permitted. Delivery of the common shares and the pre-funded warrants is expected to be made on or aboutJune, 2026. Prospectus Supplement dated June, 2026 Table of contents About this prospectus supplementS-iiProspectus supplement summaryS-1The offeringS-7Risk factorsS-10Special note regarding forward-looking statementsS-13Use of proceedsS-16Dividend policyS-17DilutionS-18Description of the securities we are offeringS-20Certain Canadian federal income tax considerationsS-23Certain U.S. federal income tax considerationsS-25UnderwritingS-33Legal mattersS-39ExpertsS-39Where you can find more informationS-39Incorporation of certain information by referenceS-40Enforceability of civil liabilitiesS-41 About this prospectusiiSpecial note regarding forward-looking statements1Market, industry and other data3The company4Risk factors8Use of proceeds9Selling securityholders10Plan of distribution11General description of our securities14Description of our common shares15Description of our warrants16Description of our debt securities18Description of our units23Where you can find more information24Incorporation by reference24Legal matters24Experts25 About this prospectus supplement This prospectus supplement and the accompanying prospectus are part of a “shelf” registrationstatement on Form S-3 that went effective upon filing with the Securities and ExchangeCommission (the “SEC”) on June28, 2024. Under this shelf registration process, we may sell anycombination of the securities described in our base prospectus included in the shelf registrationstatement in one or more offerings. This prospectus supplement relates to the offering of our common shares, pre-funded warrantsand common shares issuable upon exercise of the pre-funded warrants. Before buying any ofsuch common shares, pre-funded warrants or common shares issuable upon exercise of the pre-funded warrants, we urge you to carefully read this prospectus supplement, together with theaccompanying prospectus and the information incorporated by reference as described under theheadings “Where You Can Find More Information” and “Incorporation of Certain Information byReference” in this prospectus supplement. These documents contain important