AMX-525 (VIR-5525): A Comprehensive Report on a Novel Masked EGFRxCD3 T-Cell Engager

2. Introduction to AMX-525: A Novel Masked T-Cell Engager

AMX-525, also identified by its developmental code SAR446368 and current clinical identifier VIR-5525, is an investigational biopharmaceutical agent engineered as a next-generation T-cell engager (TCE).[1] It is currently in the early stages of clinical development for the treatment of various solid malignancies. The core therapeutic challenge AMX-525 aims to surmount is the significant dose-limiting systemic toxicities frequently associated with conventional TCE bispecific antibodies. These toxicities, often stemming from on-target, off-tumor engagement of T-cells, can severely curtail therapeutic efficacy and restrict the broader clinical applicability of TCEs.

AMX-525 represents an advancement in this field, employing a sophisticated protease-activated masking technology, known as XPAT™ (XTENylated Protease-Activated bispecific T-Cell Engagers) or Pro-XTEN™. This platform is designed to ensure that the potent T-cell-mediated cytotoxicity is predominantly unleashed within the tumor microenvironment (TME), thereby enhancing tumor-specific killing while minimizing systemic adverse events.[3] The fundamental strategy is to improve the therapeutic index by localizing potent immune activation specifically to the tumor site.

The design philosophy underpinning AMX-525 reflects a significant evolution in the field of cancer immunotherapy. It signifies a progression from generalized immune stimulation or simple targeted T-cell redirection towards a more refined and conditional strategy of localized immune activation. This level of sophistication is a direct response to the clinical limitations observed with earlier generations of TCEs. Such limitations are particularly pronounced for targets like the Epidermal Growth Factor Receptor (EGFR), which, while a valid tumor target, also exhibits widespread, low-level expression on normal tissues. This normal tissue expression makes EGFR a challenging target for unmasked TCEs due to the high risk of on-target, off-tumor toxicity. First-generation immunotherapies, such as high-dose cytokines, often induced broad and severe systemic toxicity. While monoclonal antibodies improved target specificity, their efficacy was not always sufficient. TCEs offered a leap in potency by directly linking T-cells to tumor cells; however, this potency often came at the cost of on-target, off-tumor toxicity or systemic cytokine release if the target antigen was also present on healthy cells. Masking technologies, as utilized in AMX-525, introduce an additional layer of control: conditional activation. This is typically achieved by requiring a second condition beyond target antigen binding, such as the presence of specific enzymatic activity (e.g., proteases) within the TME. This "AND-gate" logic—requiring both TAA presence and TME-specific protease activity—is engineered to dramatically improve the therapeutic window, making potent immunotherapy safer and more effective.

Should the XPAT™/Pro-XTEN™ platform, as exemplified by AMX-525, achieve successful clinical validation, it could pave the way for a new class of safer and more effective TCEs. This could broaden the applicability of TCE-based therapies to a wider range of tumor antigens and patient populations, including those targeting antigens previously considered "undruggable" by TCEs due to unacceptable toxicity profiles. Furthermore, it may enable more effective combination therapies where systemic toxicity might otherwise be an additive concern.

3. Drug Profile and Development Trajectory

AMX-525 is an investigational agent that has progressed through several developmental stages and corporate entities, reflected in its multiple identifiers.

Nomenclature and Aliases

The compound is primarily known as AMX-525, its original designation from Amunix Pharmaceuticals. Following acquisition by Sanofi, it was assigned the developmental code SAR446368. Currently, under development by Vir Biotechnology, Inc., it is identified as VIR-5525 for clinical trial purposes.[2] The consistent use of these identifiers across various sources, including company communications and clinical trial registries, is essential for accurately tracking its development. VIR-5525 is the identifier associated with its ongoing first-in-human clinical trial, NCT06960395.[1]

Pharmacological Class

AMX-525 is classified as a bispecific T-cell engager (BiTE), or more broadly, a T-cell engager (TCE).[2] Specifically, it is a dual-masked TCE engineered to target the Epidermal Growth Factor Receptor (EGFR) on tumor cells and the CD3 complex on T-lymphocytes.[2] TCEs are typically antibody-based constructs designed with two distinct binding specificities. One arm binds to a tumor-associated antigen (TAA)—EGFR in the case of AMX-525—while the other arm engages an activating receptor on T-cells, most commonly the CD3 component of the T-cell receptor complex. This dual binding physically juxtaposes T-cells with tumor cells, triggering T-cell activation and subsequent tumor cell lysis. The "dual-masked" characteristic of AMX-525 refers to the integrated prodrug technology that reversibly silences these binding activities until specific activating conditions are encountered, primarily within the tumor microenvironment.

Developmental Lineage and Licensing Agreements

The core masking technology, known as XPAT™ (XTENylated Protease-Activated bispecific T-Cell Engagers) or Pro-XTEN™, was originally conceived and developed by Amunix Pharmaceuticals.[3] Sanofi subsequently acquired Amunix Pharmaceuticals, thereby gaining control of the XPAT™ platform and its associated pipeline assets, including AMX-525 (then SAR446368).[3]

In a significant strategic move announced in August 2024, Vir Biotechnology, Inc. entered into an exclusive worldwide license agreement with Sanofi. This agreement granted Vir Biotechnology rights to three clinical-stage masked TCEs developed using this platform: SAR446309 (AMX-818, an anti-HER2 TCE), SAR446329 (AMX-500, an anti-PSMA TCE), and SAR446368 (AMX-525, an anti-EGFR TCE, now designated VIR-5525). The agreement also included exclusive use of the protease-cleavable masking platform for applications in oncology and infectious diseases.[3] This licensing pathway underscores the perceived value and potential of the masking technology, with Vir Biotechnology strategically acquiring multiple clinical-stage assets based on this technology to bolster its oncology pipeline.

The progression of the XPAT™ platform and its associated drug candidates, such as AMX-525, from a specialized biotechnology company (Amunix) to a major pharmaceutical corporation (Sanofi), and subsequently to another biotechnology company focused on immune-based therapies (Vir), suggests a multi-stage validation of the technology's potential. Each transaction implies that the acquiring or licensing entity conducted thorough due diligence and recognized substantial scientific and commercial promise. Amunix, as the innovator, likely established strong proof-of-concept with preclinical data. Sanofi's acquisition indicates a perceived broad applicability and strategic value in integrating this platform. Vir Biotechnology's subsequent licensing, particularly with its focus on "powering the immune system," suggests a belief that this platform offers a differentiated approach within the competitive immuno-oncology landscape, especially for overcoming the toxicity hurdles of conventional TCEs.

Furthermore, Vir Biotechnology's acquisition of three distinct TCEs targeting different antigens (EGFR, HER2, PSMA) but all utilizing the same masking technology points to a platform-based strategy.[3] This approach allows for diversified risk and maximizes the potential of the underlying technology. Learnings from the clinical development of one asset, such as the safety profile of the mask or pharmacokinetic and pharmacodynamic characteristics, can directly inform and potentially accelerate the development programs of the other assets sharing the same core technology.

Table 1: AMX-525 (VIR-5525) Key Characteristics

Characteristic	Details
Alternative Names	AMX-525, SAR446368, VIR-5525
Drug Class	Dual-masked Epidermal Growth Factor Receptor (EGFR) x CD3 T-Cell Engager (TCE)
Target Antigens	Epidermal Growth Factor Receptor (EGFR) on tumor cells; CD3 complex on T-lymphocytes
Mechanism of Action (Concise)	A prodrug TCE that, upon protease-mediated unmasking within the tumor microenvironment, redirects T-cell cytotoxicity against EGFR-expressing tumor cells.
Technology Platform	XPAT™ (XTENylated Protease-Activated bispecific T-Cell Engagers) / Pro-XTEN™ protease-cleavable masking technology.
Current Developer	Vir Biotechnology, Inc.
Original Developer/Licensing Path	Amunix Pharmaceuticals → Sanofi → Vir Biotechnology, Inc. (exclusive worldwide license for oncology and infectious diseases for the platform and specified assets).

This table serves as a crucial quick-reference guide, summarizing the fundamental identity, targets, mechanism of action, and developmental context of AMX-525. For an expert audience, this consolidated information facilitates a quicker understanding before delving into the more complex mechanistic and clinical details.

4. Mechanism of Action: The XPAT™ / Pro-XTEN™ Platform

The therapeutic activity of AMX-525 is predicated on its sophisticated molecular design, which combines the potent tumor cell-killing ability of a T-cell engager with a conditional activation mechanism conferred by the XPAT™ / Pro-XTEN™ platform.

Core T-Cell Engager Functionality

At its core, the unmasked AMX-525 molecule functions as a bispecific antibody construct. One binding domain is engineered to recognize and bind to the Epidermal Growth Factor Receptor (EGFR), a transmembrane protein frequently overexpressed or dysregulated on the surface of various solid tumor cells. The second binding domain targets the CD3 epsilon (CD3ε) subunit, a critical component of the T-cell receptor (TCR) complex present on virtually all T-lymphocytes.[2]

The simultaneous engagement of EGFR on a tumor cell and CD3 on a T-cell by AMX-525 creates a physical bridge, effectively forming an artificial immunological synapse. This forced juxtaposition leads to polyclonal T-cell activation, proliferation, and the release of cytotoxic effector molecules (such as perforin and granzymes) and inflammatory cytokines (such as IFN-γ and TNF-α) by the engaged T-cell. This process results in the direct lysis of the EGFR-expressing tumor cell, a mechanism that is independent of the T-cell's native antigen specificity and MHC-peptide recognition pathways.[7]

Protease-Activated Masking Technology (XPAT™ / Pro-XTEN™)

The defining feature of AMX-525 is its XPAT™ / Pro-XTEN™ masking technology, which transforms the inherently potent TCE into a prodrug that is selectively activated within the tumor microenvironment. This platform is designed to address the primary limitation of conventional TCEs: on-target, off-tumor toxicity.

• XTEN™ Polypeptide Masks: These are long, unstructured, hydrophilic, and biologically inert polypeptide sequences. In the AMX-525 construct, XTEN™ masks are genetically fused to both the N- and C-termini of the TCE's antigen-binding domains (typically single-chain variable fragments, scFvs).[6]
• Steric Shielding: The primary role of the XTEN™ masks is to physically occlude the antigen-binding sites of the TCE. This steric hindrance prevents the anti-EGFR and anti-CD3 domains from binding to their respective targets on tumor cells and T-cells when the molecule is in systemic circulation or healthy tissues. This effectively silences the TCE's activity outside the tumor. Preclinical studies with HER2-XPAT™ and EGFR-XPAT™ have demonstrated that this masking can reduce cytotoxic potency by up to 4-log-fold (10,000 to 14,000-fold) in vitro compared to their unmasked counterparts, highlighting the profound dampening effect of the masks.[6]
• Half-Life Extension: A secondary benefit of the XTEN™ polymers is the extension of the molecule's plasma half-life. Their large hydrodynamic volume, similar to the effect of PEGylation, reduces renal clearance. This property can facilitate less frequent dosing regimens and ensure sustained systemic exposure of the masked prodrug, allowing for better tumor penetration. XTEN™ polypeptides are also designed for low immunogenicity.[6]
• Protease-Cleavable Linkers: The XTEN™ masks are tethered to the TCE core via specialized peptide linker sequences. These linkers are meticulously engineered to be susceptible to cleavage by specific classes of proteases that are known to be highly active or overexpressed within the TME. These include matrix metalloproteinases (MMPs), serine proteases (e.g., urokinase-type plasminogen activator (uPA), matriptase), and cysteine proteases (e.g., cathepsins).[3]
• Conditional Activation within the Tumor Microenvironment (TME):
• Systemic Inactivity: In healthy tissues and systemic circulation, the activity of proteases is typically tightly regulated by endogenous inhibitors. Consequently, the XPAT™ protein is expected to remain predominantly in its masked, inactive (prodrug) state, minimizing engagement with EGFR or CD3 on normal cells.[6]
• TME-Specific Unmasking: Many solid tumors exhibit a dysregulated proteolytic environment characterized by elevated levels and activity of various proteases. This aberrant protease activity within the TME facilitates the cleavage of the specialized linkers in AMX-525. This enzymatic cleavage leads to the sequential removal of the XTEN™ masks, thereby exposing the antigen-binding domains of the TCE and restoring its full biological activity specifically at the tumor site.[3] The unmasking process can occur in two steps, potentially yielding partially active intermediates before full activation is achieved.[6]
• Pharmacokinetics of the Activated TCE: A critical aspect of the XPAT™ design is that the fully unmasked, active TCE (uTCE) is engineered to possess a relatively short systemic half-life. This ensures that if any activated TCE diffuses away from the TME and enters systemic circulation, it is rapidly cleared. This rapid clearance further minimizes the potential for systemic on-target, off-tumor toxicities, contributing to the overall safety profile.[6]

Overall Rationale for Enhanced Therapeutic Index

The primary objective of this intricate molecular engineering is to mitigate the severe dose-limiting toxicities frequently observed with conventional, unmasked TCEs. These toxicities include systemic cytokine release syndrome (CRS) and on-target damage to normal tissues that express the target antigen (EGFR, in the case of AMX-525).[3] By confining potent T-cell activation to the tumor site, the XPAT™ platform aims to enable the administration of higher, more therapeutically effective doses of the TCE, thereby significantly widening the therapeutic window and improving the benefit-risk profile of the therapy.

The XPAT™ platform operates on a sophisticated "AND-gate" principle for TCE activation. Full therapeutic potency is achieved only when two conditions are met: (1) the presence of the target antigen (EGFR) on cells, allowing for initial, albeit weak, localization of the masked TCE, AND (2) the presence of sufficiently high protease activity in the local microenvironment to efficiently cleave the masks and liberate the active TCE. This dual-requirement system is inherently more selective than targeting strategies based on single-antigen recognition alone. Traditional TCEs essentially rely on a single "key"—the presence of the TAA. If normal cells also possess this TAA, they can be inadvertently targeted, leading to toxicity. The XPAT™ platform introduces a second "lock"—the protease-cleavable mask. The TME, with its characteristic high protease activity, possesses the "key" to this second lock. Therefore, only within the TME, where both the TAA is present and proteases are sufficiently active, can the TCE be fully "unlocked" to exert its potent cytotoxic effect. This design dramatically enhances specificity compared to single-lock systems.

Furthermore, the platform leverages a dichotomous pharmacokinetic strategy. A long half-life is engineered for the masked prodrug, facilitated by the XTEN™ polymers, to ensure adequate tumor delivery and potentially more convenient dosing schedules. Conversely, a short half-life is designed for the unmasked, active TCE. This ensures that if the active drug escapes the TME, its systemic exposure is limited, thereby reducing the risk of systemic toxicities. This careful pharmacokinetic engineering is a key differentiator of the XPAT™ platform.

The success of this platform in a clinical setting will depend, in part, on the consistency and level of differential protease activity between tumor tissues and healthy tissues. Variability in TME protease profiles across different tumor types, or even inter-patient and intra-tumoral heterogeneity, could influence the efficiency of AMX-525 activation and, consequently, its therapeutic efficacy. This highlights a potential avenue for future biomarker development, such as assessing TME protease signatures to identify patient populations most likely to benefit from this therapeutic approach.

5. Preclinical Evidence for AMX-525 (EGFR-XPAT™) and its Underlying Platform

The preclinical data supporting the clinical development of AMX-525 (VIR-5525) are derived from studies on the EGFR-XPAT™ protein itself and from analogous studies on the HER2-XPAT™ protein, which serve to validate the broader XPAT™/Pro-XTEN™ platform technology. These studies have provided critical insights into the molecule's activity, selectivity, and safety profile.

In Vitro Characterization

• Binding Affinity and Masking Efficacy: The unmasked TCE core of EGFR-XPAT™ (and its HER2-XPAT™ counterpart) demonstrates high-affinity binding to its respective tumor-associated antigen (EGFR or HER2) and to the CD3 complex on T-cells.[6] The XTEN™ masking was shown to be highly effective in attenuating this binding and subsequent activity. In vitro, EGFR-XPAT™ exhibited a greater than 4-log-fold (i.e., >10,000-fold) reduction in T-cell mediated cytotoxic activity compared to its unmasked form.[6] Similarly, the HER2-XPAT™ showed up to a 14,000-fold protection from cytotoxicity.[7] This substantial reduction in potency of the masked form is crucial for preventing premature T-cell activation in systemic circulation. Studies with HER2-XPAT™ also showed minimal cytotoxicity against normal cells, such as cardiomyocytes, even at high concentrations (1µM), underscoring the safety aspect of the masking towards non-tumor cells.[7]
• Protease-Dependent Activation: The cytotoxic potential of the masked TCEs was effectively restored upon exposure to TME-relevant proteases. This confirmed the designed cleavability of the linkers and validated the conditional activation mechanism, which relies on the enzymatic activity prevalent in the tumor environment.[6]

In Vivo Antitumor Activity (Xenograft Models)

• EGFR-XPAT™: In immunodeficient mouse models engrafted with human peripheral blood mononuclear cells (huPBMCs) and bearing HT-29 human colorectal tumor xenografts (which express EGFR), EGFR-XPAT™ demonstrated potent, dose-dependent antitumor activity.[6]
• HER2-XPAT™ (Platform Validation):
• In HER2-high expressing BT-474 breast cancer xenograft models, HER2-XPAT™ induced complete tumor regressions, with efficacy comparable to equimolar doses of the unmasked HER2-TCE.[7]
• A critical validation of the mechanism came from studies using a non-cleavable version of HER2-XPAT™, where the protease-sensitive linkers were altered. This non-cleavable construct lacked in vivo efficacy, directly demonstrating that protease cleavage within the TME is essential for the therapeutic activity of XPAT™ proteins.[6]
• Importantly, efficacy was also observed in HER2-low expressing HT-55 colorectal cancer models, suggesting that the platform may have utility beyond tumors with high antigen expression levels.[7] This is a significant finding, as it implies that the enhanced safety margin allowing for higher dosing might compensate for lower target density.
• Tumor-Specific Cleavage: In vivo studies using fluorescently-labeled HER2-XPAT™ in tumor-bearing mice provided direct evidence of TME-selective unmasking. These experiments showed preferential proteolytic cleavage of the XPAT™ protein and accumulation of the active (unmasked) TCE form within the tumor, with significantly less cleavage observed in healthy organs.[6]

Pharmacokinetics and Safety in Non-Human Primates (NHPs)

NHPs serve as a crucial translational model for assessing potential human toxicities of immunotherapies, particularly concerning cytokine release syndrome and on-target, off-tumor effects.

• EGFR-XPAT™:
• In NHP studies, a single intravenous (IV) dose of EGFR-XPAT™ at 0.46 mg/kg resulted in only minor toxicity and transient cytokine release. The Maximum Tolerated Dose (MTD) for a single IV administration was established at 1 mg/kg.[6]
• A key finding highlighting the improved therapeutic window was that the peak plasma concentration (Cmax) achieved at the EGFR-XPAT™ MTD (1 mg/kg) was at least 200-fold higher than the Cmax observed at the MTD of its corresponding unmasked EGFR-TCE (which was 0.033 mg/kg/day administered via a 48-hour continuous IV infusion).[6] This substantial difference underscores the significant safety benefit conferred by the masking technology.
• HER2-XPAT™ (Platform Validation):
• HER2-XPAT™ demonstrated a robust safety profile in NHPs, with an MTD of up to 42 mg/kg.[7]
• This translated to a greater than 400-fold increase in the tolerated Cmax in NHPs compared to its unmasked HER2-TCE counterpart.[6]
• Significantly, minimal Cytokine Release Syndrome (CRS) or systemic T-cell activation was observed even at very high doses (e.g., up to 50 mg/kg for AMX-818/HER2-XPAT™). In contrast, the unmasked HER2-TCE induced lethal CRS at much lower doses (e.g., >0.3 mg/kg/day).[7] Early T-cell margination observed with HER2-XPAT™ at 2mg/kg was transient, further supporting a mitigated systemic immune response with the masked form.
• Pharmacokinetic analysis in NHPs revealed high plasma stability of HER2-XPAT™, with only 1-3% of singly-cleaved metabolites detected in plasma and no detectable fully unmasked TCE, even at high administered doses.[7]
• Translational Relevance of Stability: In vitro studies demonstrated that HER2-XPAT™ exhibited low and comparable levels of cleavage in plasma samples from healthy humans, diseased humans (cancer patients), and NHPs. This finding supports the hypothesis that the stability and masking characteristics observed in preclinical NHP studies are likely to be translatable to the human clinical setting, providing confidence in the platform's behavior in patients.[6]

The striking quantitative improvements in tolerated Cmax in NHPs for both EGFR-XPAT™ (>200-fold) and HER2-XPAT™ (>400-fold) compared to their unmasked counterparts provide compelling evidence that the XPAT™ masking technology effectively mitigates systemic toxicity. This is not a marginal improvement but rather a potentially transformative shift in the safety profile of TCEs. The observation that these masked TCEs are tolerated at systemic exposures orders of magnitude higher than unmasked versions implies that the masking is effectively preventing widespread, dose-limiting immune activation in healthy tissues. Such a substantial increase in the tolerated dose is a strong indicator of an expanded therapeutic window, a primary objective of prodrug strategies in oncology.

Furthermore, the in vivo efficacy of HER2-XPAT™ being critically dependent on protease cleavability (as the non-cleavable version was inactive) is a crucial piece of mechanistic validation.[6] This directly supports the proposed mechanism of TME-specific activation and largely rules out significant therapeutic activity from the masked form itself or through non-specific unmasking pathways. This confirmation of the "conditional activation" design reduces the risk that the drug might operate through an alternative, unintended mechanism.

The efficacy of HER2-XPAT™ in HER2-low expressing tumor models is also highly significant.[7] It suggests that the improved safety profile, which allows for higher drug exposure, might enable effective targeting of tumors with lower antigen densities. Many solid tumors express TAAs at low or heterogeneous levels, often rendering them poor candidates for conventional targeted therapies that require high antigen density for efficacy. The XPAT™ platform could potentially make such targets accessible to TCE therapy by allowing for higher drug concentrations within the TME, thereby increasing the probability of productive T-cell engagement even with fewer TAAs per tumor cell.

Table 2: Summary of Key Preclinical Findings for EGFR-XPAT™ Platform (Relevant to AMX-525/VIR-5525)

Parameter	Model System	Key Finding for EGFR-XPAT™ (AMX-525/VIR-5525)	Key Finding for Platform (e.g., HER2-XPAT™)	Source Snippet(s)
In Vitro Masking Efficacy (vs. unmasked TCE)*	Tumor cell lines + T-cells	>4-log-fold protection from cytotoxicity	Up to 14,000-fold protection	6
In Vivo Antitumor Efficacy*	HT-29 (colorectal) xenograft + huPBMCs	Potent antitumor activity	Protease-dependent complete regressions; efficacy in HER2-low models (BT-474, HT-55 xenografts)	6
Tumor-Specific Activation	Murine Xenograft (fluorescent label)	Data not specified for EGFR-XPAT	Preferential cleavage in tumor vs. healthy tissue	6
NHP MTD (Single IV Dose)	Non-Human Primates	1 mg/kg	Up to 42 mg/kg	6
NHP Safety Margin (Tolerated Cmax vs. unmasked TCE)	Non-Human Primates	>200-fold increase	>400-fold increase	6
NHP Systemic Effects (e.g., CRS, T-cell activation)	Non-Human Primates	Minor toxicity/cytokine release at 0.46 mg/kg	Minimal CRS/systemic T-cell activation even at high doses	6
Plasma Stability (Cleavage)	NHP plasma; Human plasma (healthy & diseased)	Data not specified for EGFR-XPAT	High stability; minimal cleavage (1-3% singly-cleaved metabolites in NHP plasma); low & similar cleavage in human & NHP plasma in vitro	6

This comparative table effectively highlights the consistency of the XPAT™ platform's performance across different targets (EGFR and HER2). It underscores the platform's core benefits—substantial improvement in safety margins and tumor-selective activation—which are the primary justifications for advancing AMX-525/VIR-5525 into clinical trials. The direct comparison of MTD and tolerated Cmax improvements is particularly impactful for an expert audience.

6. Clinical Development Program for AMX-525 (VIR-5525)

The transition of AMX-525 (now VIR-5525) into clinical development marks a critical phase in evaluating its therapeutic potential in humans.

Regulatory Status

An Investigational New Drug (IND) application for AMX-525 (SAR446368) has been cleared by the U.S. Food and Drug Administration (FDA). This regulatory clearance is a prerequisite for initiating clinical trials in the United States and signifies that the FDA has reviewed preclinical data and found it adequate to support testing in human subjects.[4]

Phase 1 Clinical Trial (NCT06960395)

This is the first-in-human (FIH) study designed to assess VIR-5525.

• Official Title: The study is registered under the title: "A Phase 1, First-in-Human Study of the Safety, Pharmacokinetics, and Preliminary Efficacy of VIR-5525 Alone and in Combination With Pembrolizumab in Participants With Locally Advanced or Metastatic Solid Tumors".[8]
• Study Design: NCT06960395 is an open-label, multicenter, Phase 1 study. It follows a standard design for early-phase oncology trials, incorporating a dose-escalation phase to determine safety and optimal dosing, followed by a dose-expansion phase to further evaluate safety and preliminary efficacy in specific patient cohorts.[8] The study includes two main arms:
• Arm 1 (Monotherapy): VIR-5525 will be administered as a single agent.
• Arm 2 (Combination Therapy): VIR-5525 will be administered in combination with pembrolizumab, an established anti-PD-1 monoclonal antibody. The inclusion of a combination arm with a checkpoint inhibitor from the outset is a notable feature of the trial design, aiming to explore potential synergistic anti-tumor effects early in development.[8]
• Key Objectives and Endpoints: Based on the trial registration details [8]:
• Primary Objectives:
• To evaluate the safety and tolerability profile of VIR-5525 when administered as monotherapy and when combined with pembrolizumab.
• To identify any dose-limiting toxicities (DLTs).
• To determine the Maximum Tolerated Dose (MTD) and/or the Recommended Phase 2 Dose (RP2D) for VIR-5525, both as a single agent and in the combination regimen.
• Secondary Objectives:
• To characterize the pharmacokinetic (PK) profile of VIR-5525.
• To obtain preliminary assessments of anti-tumor activity for VIR-5525 monotherapy and combination therapy. Efficacy measures will likely include Objective Response Rate (ORR), Duration of Response (DOR), and Disease Control Rate (DCR), assessed according to Response Evaluation Criteria in Solid Tumors (RECIST) v1.1.
• Target Patient Population: The trial is designed for adult participants with locally advanced or metastatic solid tumors that are known to express EGFR.[8]
• Specific Tumor Types for Enrollment: Initial enrollment will focus on patients with various EGFR-expressing malignancies, including but not limited to colorectal cancer (CRC), squamous cell carcinoma of the head and neck (SCCHN), non-small cell lung cancer (NSCLC), and renal cell carcinoma (RCC).[4]
• Key Eligibility Criteria (Illustrative): While full protocol details are not provided in the available information, general eligibility would include:
• Histologically or cytologically confirmed diagnosis of an EGFR-expressing solid tumor.
• Locally advanced or metastatic disease.
• Documented disease progression on, or intolerance to, prior standard therapies, or patients for whom no standard treatment options are available.
• (Specific prior therapy requirements, Eastern Cooperative Oncology Group (ECOG) performance status, adequate organ function, and other standard oncology trial criteria would be detailed in the full protocol.)
• Current Status and Projected Timelines:
• The trial status has been listed as "Recruiting" in some databases [1], while a clinical trial listing updated May 7, 2025, indicated "Not Yet Enrolling".[8]
• Initial enrollment was anticipated by Vir Biotechnology to commence in the first quarter of 2025 or sooner.[4] Corporate presentations from Vir Biotechnology in May 2025 projected the Phase 1 study start and first-in-human (FIH) dosing in the second quarter of 2025.[11] The report will reflect the latest available information, noting that slight variations in projected start dates are common during the initiation phase of clinical trials.
• Sponsor: The clinical trial is sponsored by Vir Biotechnology, Inc..[1]

The decision to evaluate VIR-5525 in combination with pembrolizumab from the outset in a Phase 1 setting, rather than pursuing monotherapy development to a more advanced stage first, suggests an aggressive and scientifically driven development strategy. This approach is likely based on a strong preclinical rationale for synergy between TCE-mediated T-cell activation and PD-1 blockade. TCEs, like unmasked VIR-5525, activate T-cells to target and kill tumor cells. However, activated T-cells can upregulate inhibitory receptors such as PD-1, and tumor cells or other cells in the TME can express PD-L1, leading to T-cell exhaustion and a dampening of the anti-tumor immune response. Pembrolizumab, an anti-PD-1 antibody, blocks this critical inhibitory checkpoint, potentially sustaining or enhancing the T-cell response initiated by the TCE. Testing this combination early allows for a quicker assessment of this synergistic potential, which could accelerate the path to a more effective therapeutic regimen if the hypothesis holds true.

Furthermore, VIR-5525 (targeting EGFR) is one of three clinical-stage masked TCEs that Vir Biotechnology is concurrently advancing, the others being VIR-5818 (targeting HER2) and VIR-5500 (targeting PSMA).[9] These assets all share the core XPAT™/Pro-XTEN™ masking technology. Consequently, early clinical data emerging from any of these trials—particularly concerning the safety profile of the platform, its pharmacokinetic behavior (e.g., half-life of the masked drug), evidence of TME-specific unmasking versus systemic cleavage, and any immunogenicity related to the XTEN™ polypeptides—will be highly informative for the entire portfolio. Positive read-across from one program could significantly de-risk and build confidence in the others that utilize the same foundational technology.

While not explicitly detailed for NCT06960395 in the provided information, a critical component in the development of TME-activated prodrugs like VIR-5525 will be the comprehensive exploration of predictive biomarkers. This could involve assessing EGFR expression levels and heterogeneity, characterizing TME protease activity profiles, and evaluating the immune cell infiltrate in patient tumors. Such translational research will be essential to identify patient populations most likely to derive benefit from VIR-5525 and to understand the determinants of response and resistance. The inclusion of diverse tumor types (CRC, SCCHN, NSCLC, RCC) in the initial cohorts of the Phase 1 trial may facilitate the identification of differential responses based on these factors, guiding future development strategies.

Table 3: Overview of Clinical Trial NCT06960395 for VIR-5525 (AMX-525)

Category	Details
Official Title	A Phase 1, First-in-Human Study of the Safety, Pharmacokinetics, and Preliminary Efficacy of VIR-5525 Alone and in Combination With Pembrolizumab in Participants With Locally Advanced or Metastatic Solid Tumors
Phase	Phase 1
Study Design	Open-label, multicenter, first-in-human, dose-escalation and dose-expansion study. Includes monotherapy (VIR-5525) and combination therapy (VIR-5525 + pembrolizumab) arms.
Primary Objectives	Evaluate safety and tolerability of VIR-5525 (monotherapy and combination); determine Maximum Tolerated Dose (MTD) and/or Recommended Phase 2 Dose (RP2D).
Key Secondary Endpoints	Pharmacokinetics of VIR-5525; preliminary anti-tumor activity (e.g., Objective Response Rate (ORR), Duration of Response (DOR), Disease Control Rate (DCR) per RECIST v1.1).
Target Population	Adult participants with locally advanced or metastatic EGFR-expressing solid tumors who have progressed on or are intolerant to standard therapies.
Planned Tumor Cohorts (Initial)	Colorectal cancer, Squamous Cell Carcinoma of the Head and Neck (SCCHN), Non-Small Cell Lung Cancer (NSCLC), Renal Cell Carcinoma (RCC).
Key Interventions	VIR-5525 (AMX-525); Pembrolizumab.
Sponsor	Vir Biotechnology, Inc.
Current Status (as of May 2025)	Recruiting / Not Yet Enrolling; First-in-human dosing anticipated Q2 2025.

This table is fundamental to the report as it details the human testing phase of AMX-525. It outlines how the drug's safety and initial efficacy are being assessed, the patient populations being studied, and the overall structure of this pivotal first-in-human trial. This information is critical for understanding the drug's path to potential approval.

7. Therapeutic Potential and Target Indications

The therapeutic strategy for AMX-525 (VIR-5525) is centered on targeting the Epidermal Growth Factor Receptor (EGFR) in various solid tumors, leveraging the novel XPAT™/Pro-XTEN™ masking technology to potentially overcome limitations of existing EGFR-targeted therapies and conventional T-cell engagers.

Rationale for EGFR-Targeted Therapy in Solid Tumors

EGFR is a well-established oncogenic driver and a validated therapeutic target in oncology. It is a transmembrane tyrosine kinase receptor that, upon activation by ligands such as EGF and TGF-α, triggers intracellular signaling pathways (e.g., RAS/MAPK, PI3K/AKT) crucial for cell proliferation, survival, differentiation, invasion, and metastasis. Overexpression, amplification, or activating mutations of EGFR are frequently observed in a variety of epithelial cancers, contributing to malignant transformation and progression.[4]

Existing therapeutic strategies targeting EGFR include monoclonal antibodies (mAbs) that block ligand binding or receptor dimerization (e.g., cetuximab, panitumumab) and small molecule tyrosine kinase inhibitors (TKIs) that inhibit EGFR's intracellular kinase activity (e.g., gefitinib, erlotinib, osimertinib). While these agents have shown clinical benefit in certain patient populations, their efficacy can be limited by primary or acquired resistance mechanisms and significant on-target toxicities due to EGFR expression in normal tissues (e.g., skin, gastrointestinal tract). TCEs like AMX-525 offer a distinct mechanism of action by directly harnessing the cytotoxic potential of T-cells against EGFR-expressing tumor cells, which could be effective even in tumors resistant to mAbs or TKIs.

Identified Tumor Types for Initial Investigation

The Phase 1 clinical trial (NCT06960395) for VIR-5525 plans to enroll patients with a range of EGFR-expressing solid tumors. The initially specified tumor types for investigation include:

• Colorectal Cancer (CRC) [4]
• Squamous Cell Carcinoma of the Head and Neck (SCCHN) [4]
• Non-Small Cell Lung Cancer (NSCLC) [4]
• Renal Cell Carcinoma (RCC) [4]

These indications represent areas of significant unmet medical need where EGFR is frequently expressed and novel therapeutic approaches are actively being sought. The selection of these diverse tumor types will allow for an initial assessment of VIR-5525's activity across different histological contexts and varying tumor microenvironments, which may differ in their protease profiles and immune landscapes.

Potential Advantages of a Masked EGFR-Targeted TCE

The primary anticipated advantage of AMX-525, owing to its XPAT™/Pro-XTEN™ masking technology, is the potential to overcome the severe dose-limiting toxicities that would likely plague an unmasked EGFR-TCE. EGFR is expressed on various normal epithelial tissues, including skin keratinocytes and cells of the gastrointestinal mucosa. Direct and systemic engagement of T-cells with these normal EGFR-expressing cells by an unmasked TCE could lead to severe dermatologic toxicities (e.g., rash, desquamation) and gastrointestinal toxicities (e.g., diarrhea, mucositis), similar to or potentially more severe than those seen with anti-EGFR mAbs. Such toxicities would likely limit the achievable therapeutic dose of an unmasked EGFR-TCE to sub-optimal levels.

By confining potent T-cell activation to the TME through protease-dependent unmasking, AMX-525 aims to achieve a significantly improved therapeutic index.[3] This could allow for:

• Higher Effective Dosing: Administration of AMX-525 at doses that result in greater tumor exposure and more effective T-cell engagement within the tumor, without inducing prohibitive systemic toxicities.
• Treatment of Tumors with Broader EGFR Expression: Potential utility in tumors where EGFR is a viable target but where conventional TCEs would be too toxic due to normal tissue expression.
• Overcoming Resistance: Offering a therapeutic option for patients whose tumors have developed resistance to other EGFR-targeted therapies (mAbs or TKIs) due to its distinct immune-mediated mechanism of action.

The success of AMX-525 could significantly expand the landscape of "druggable" targets for TCEs. Many compelling tumor antigens, like EGFR, are also expressed on vital normal tissues, making them challenging for highly potent immunotherapies. If the XPAT™ platform effectively mitigates on-target, off-tumor toxicity for EGFR, it would provide strong validation for applying this masking strategy to other such antigens. This could unlock a new class of TCEs with improved safety and efficacy, potentially transforming the treatment paradigms for numerous solid tumors. The choice of initial tumor types for the Phase 1 trial—CRC, SCCHN, NSCLC, and RCC—likely reflects a strategic balance between the prevalence of EGFR expression in these cancers, the existing unmet medical need, and potentially, an assessment of TME characteristics (e.g., protease profiles) that might favor drug activation. Early clinical signals from these diverse cohorts will be crucial in guiding the future development path of AMX-525 and identifying the patient populations most likely to benefit.

8. Strategic Implications and Future Perspectives

The development of AMX-525 (VIR-5525) and the broader licensing of the XPAT™/Pro-XTEN™ platform carry significant strategic implications for Vir Biotechnology and the field of immuno-oncology.

Positioning within Vir Biotechnology's Oncology Portfolio

AMX-525 (VIR-5525) is a cornerstone of Vir Biotechnology's strategic expansion into the oncology therapeutic area. Known primarily for its work in infectious diseases (e.g., treatments for COVID-19, Hepatitis B and D), Vir is leveraging licensed, clinically advanced masked TCE technology to build a competitive oncology pipeline.[3] This move represents a significant diversification for the company, capitalizing on its core expertise in immunology and applying it to a new, high-value therapeutic domain.

AMX-525 is one of three masked TCEs licensed from Sanofi that Vir is advancing into or through early clinical development. The others are VIR-5818 (AMX-818), targeting HER2, and VIR-5500 (AMX-500), targeting PSMA.[3] This portfolio indicates a platform-based approach, where the underlying XPAT™/Pro-XTEN™ technology is the common denominator. The company's stated strategy involves "powering the immune system" against both cancer and infectious diseases, and these masked TCEs align directly with this mission in the oncology space.[3]

Potential Advantages Over Conventional (Unmasked) T-Cell Engagers

The primary differentiating factor and anticipated advantage of AMX-525 and other XPAT™-based TCEs over conventional, unmasked TCEs lie in the potential for a significantly improved therapeutic index. This is expected due to:

• TME-Selective Activation: The protease-dependent unmasking mechanism is designed to restrict potent T-cell engagement primarily to the tumor site, sparing healthy tissues.[3]
• Reduced Systemic Toxicity: This localized activation is hypothesized to lead to a better safety and tolerability profile, with a reduced risk of systemic cytokine release syndrome (CRS) and other on-target, off-tumor toxicities that often limit the utility of unmasked TCEs, especially those targeting antigens like EGFR with notable normal tissue expression.[6]
• Broader Applicability: The improved safety profile might allow for effective treatment of tumors with lower or more heterogeneous antigen expression levels, or for targeting antigens with more widespread normal tissue expression, which are often intractable with conventional TCEs.

Future Research Directions

The clinical development of AMX-525 will likely inform several future research avenues:

• Indication Expansion: Based on signals of activity observed in the Phase 1 trial, VIR-5525 could be expanded into additional EGFR-expressing solid tumor indications.
• Biomarker Development: A critical area will be the identification and validation of predictive biomarkers. These could include quantification of EGFR expression levels, assessment of TME protease signatures (to predict efficient drug unmasking), and characterization of the tumor immune infiltrate. Such biomarkers could help in patient selection and optimizing therapeutic outcomes.
• Combination Strategies: Beyond the initial combination with pembrolizumab, further exploration of synergistic combinations with other immunotherapies, targeted agents, or chemotherapy could be pursued to enhance efficacy or overcome resistance.
• Platform Application: Vir Biotechnology may leverage its exclusive rights to the Pro-XTEN™ masking platform to develop novel masked therapeutics based on its internally discovered antibodies, potentially utilizing its proprietary dAIsY™ (data AI structure and antibody) AI engine for antibody discovery and engineering.[9]

Potential Challenges

Despite the promising preclinical data and innovative design, the development of AMX-525 is not without potential challenges:

• TME Heterogeneity: The efficacy of AMX-525 will depend on the consistent presence and sufficient activity of the relevant proteases within diverse human tumor microenvironments. TME protease profiles can vary significantly between tumor types, between patients, and even within different regions of the same tumor. Insufficient or heterogeneous protease activity could lead to suboptimal drug activation and reduced efficacy.
• Immunogenicity: While the XTEN™ polypeptide masks are designed for low immunogenicity [7], the potential for immune responses against any component of the complex biologic (XTEN™ masks, linker remnants, or the TCE itself) in humans remains a consideration that will be monitored in clinical trials.
• Manufacturing Complexity: The production of dual-masked, multi-component TCEs like AMX-525 can be complex and costly, requiring sophisticated biomanufacturing processes to ensure consistency and quality at scale.

The strategic reliance on a single core technology (XPAT™/Pro-XTEN™) for multiple lead oncology assets (EGFR, HER2, and PSMA-targeted TCEs) creates a concentrated risk-reward profile for Vir Biotechnology. Early clinical data, particularly regarding safety, pharmacokinetics, and evidence of TME-specific activation from any of these three programs, will be highly impactful for the entire platform's valuation and future development trajectory. Positive safety data or clear evidence of tumor-localized unmasking from one trial would significantly de-risk the platform and bolster confidence in the other assets. Conversely, unexpected platform-related toxicities (e.g., immunogenicity of XTEN™ components, off-target cleavage by systemic proteases) in one program could cast doubt on the viability of the others.

The long-term success of AMX-525 will depend not only on its individual clinical performance but also on the broader validation of the protease-activated masking concept in diverse human tumor microenvironments. If successful, it could represent a significant advancement in making highly potent T-cell redirecting therapies safer and more broadly applicable in the fight against cancer.

9. Conclusion

AMX-525 (VIR-5525) emerges from preclinical evaluation as a highly promising, next-generation investigational therapeutic for EGFR-expressing solid tumors. Its identity as a dual-masked EGFRxCD3 T-cell engager, built upon the innovative XPAT™/Pro-XTEN™ protease-activated masking technology, positions it to address a critical unmet need in cancer immunotherapy: the mitigation of severe systemic toxicities associated with conventional T-cell engagers.

The preclinical data, particularly from non-human primate studies, provide a strong rationale for its clinical development. These studies have demonstrated a significantly improved therapeutic index for XPAT™-based TCEs compared to their unmasked counterparts, primarily attributed to the TME-selective cleavage of the masking XTEN™ polypeptides by tumor-associated proteases. This conditional activation mechanism aims to concentrate potent T-cell-mediated cytotoxicity at the tumor site, thereby sparing healthy tissues and reducing the likelihood of dose-limiting systemic adverse events. For EGFR-XPAT™, this translated to a greater than 200-fold increase in the tolerated maximum concentration in NHPs, a key indicator of an enhanced safety margin.

The progression of AMX-525 (VIR-5525) into a Phase 1 first-in-human clinical trial (NCT06960395) is a critical step. This study will evaluate its safety, pharmacokinetics, and preliminary efficacy as both a monotherapy and in combination with the PD-1 inhibitor pembrolizumab in patients with various advanced EGFR-expressing solid tumors, including colorectal cancer, SCCHN, NSCLC, and renal cell carcinoma. The insights gained from this trial will be pivotal in determining the future trajectory of AMX-525 and will also provide valuable clinical validation for the broader XPAT™/Pro-XTEN™ platform technology.

If the promising preclinical profile translates into a favorable safety and efficacy profile in humans, AMX-525 (VIR-5525) has the potential to offer a new, more tolerable, and effective treatment option for patients with challenging EGFR-expressing cancers. More broadly, the success of such advanced masking and conditional activation strategies could significantly expand the applicability of T-cell engaging therapies in oncology, allowing for the targeting of a wider array of tumor antigens while improving patient outcomes.

1. Executive Summary

AMX-525, also known as SAR446368 and currently under the clinical development identifier VIR-5525, is an investigational biopharmaceutical agent designed as a dual-masked, protease-activated T-cell engager (TCE) targeting the Epidermal Growth Factor Receptor (EGFR) on tumor cells and the CD3 complex on T-lymphocytes.[2] Developed using the innovative XPAT™/Pro-XTEN™ platform technology, AMX-525 aims to address the significant challenge of systemic toxicity associated with conventional TCEs by confining potent T-cell activation to the tumor microenvironment (TME).[3]

The core mechanism involves XTEN™ polypeptide masks that sterically shield the TCE's binding domains, rendering it largely inactive in systemic circulation. These masks are linked via protease-cleavable sequences, designed to be selectively cleaved by proteases overexpressed in the TME, thereby unmasking the TCE and enabling localized T-cell-mediated tumor cell lysis.[6] Preclinical studies, including those with an EGFR-XPAT™ prototype and a HER2-XPAT™ platform analogue, have demonstrated a significantly improved therapeutic index. Notably, EGFR-XPAT™ showed a >200-fold higher tolerated maximum concentration in non-human primates compared to its unmasked counterpart, alongside potent, protease-dependent anti-tumor activity in vivo.[6]

AMX-525 (VIR-5525) has received IND clearance from the FDA and is advancing into a Phase 1 clinical trial (NCT06960395). This first-in-human study will evaluate the safety, pharmacokinetics, and preliminary efficacy of VIR-5525 as a monotherapy and in combination with the PD-1 inhibitor pembrolizumab in adult participants with locally advanced or metastatic EGFR-expressing solid tumors, including colorectal cancer, squamous cell carcinoma of the head and neck, non-small cell lung cancer, and renal cell carcinoma.[4]

The development of AMX-525 by Vir Biotechnology, following its acquisition from Sanofi (originating from Amunix Pharmaceuticals), represents a strategic entry into oncology leveraging a platform with the potential to overcome key limitations of current immunotherapies.[3] If clinically validated, AMX-525 could offer a safer and more effective treatment modality for a range of challenging solid tumors, and the underlying XPAT™/Pro-XTEN™ technology could unlock a new class of conditionally activated biologics.

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