TNG-348: A Comprehensive Review of a USP1 Inhibitor for HRD+ Cancers

1. Introduction to TNG-348

TNG-348 emerged as an orally bioavailable, small molecule inhibitor designed to target the ubiquitin-specific protease 1 (USP1).[1] Its development was rooted in the principle of synthetic lethality, aiming to exploit specific vulnerabilities in cancer cells with deficient DNA repair mechanisms.[2]

1.1. Chemical Identity and Basic Properties

TNG-348 is chemically identified by the CAS Number 2839740-79-1.[2] Its molecular formula is C27​H23​F6​N9​O, corresponding to a molecular weight of 603.5 g/mol.[2] For computational drug-like property assessment, its CLogP (a measure of lipophilicity) is 3, and its Topological Polar Surface Area (TPSA) is 104 Ų.[2] These parameters are crucial in early drug development for predicting pharmacokinetic properties such as absorption and membrane permeability. The SMILES (Simplified Molecular Input Line Entry System) string for TNG-348 is CN1C=C(N=C1C2=CC=C(C=C2)CN3C4=NC(=NC=C4N(C3=N)CC(F)(F)F)C5=C(N=CN=C5OC)C6CC6)C(F)(F)F.[2]

1.2. Developer and Origin

TNG-348 was developed by Tango Therapeutics, a clinical-stage biotechnology company specializing in precision oncology and synthetic lethality.[2] The foundational preclinical USP1 inhibitor program that led to the discovery and development of TNG-348 was in-licensed by Tango Therapeutics from Medivir AB in 2020.[4] This collaboration highlights a common strategy in the pharmaceutical industry where specialized biotechnology companies advance promising preclinical assets originating from other research entities. Tango Therapeutics' focus on synthetic lethality suggests that the precursor program from Medivir demonstrated significant potential in selectively targeting cancer cells with specific DNA repair deficiencies, aligning with Tango's core research and development strategy.

1.3. Therapeutic Class

TNG-348 is classified as a Ubiquitin-Specific Protease 1 (USP1) inhibitor.[1] More specifically, it functions as an allosteric, selective, and reversible inhibitor of the USP1 enzyme.[12] USP1 is a deubiquitinating enzyme (DUB) that plays a critical role in DNA damage repair processes. The development of an allosteric, selective, and reversible inhibitor like TNG-348 likely represented a deliberate drug design approach. This strategy may have been aimed at achieving greater target specificity and potentially a more favorable safety profile compared to earlier or non-allosteric USP1 inhibitors, although, as later discussed, safety objectives were not ultimately met.

Table 1: TNG-348 Key Characteristics

Characteristic	Details
Name	TNG-348
Alternative Designations	Company Code: TNG348
Developer	Tango Therapeutics
Origin	Preclinical program in-licensed from Medivir
CAS Number	2839740-79-1
Molecular Formula	C27​H23​F6​N9​O
Molecular Weight	603.5 g/mol
Therapeutic Class	USP1 inhibitor (allosteric, selective, reversible)
SMILES String	CN1C=C(N=C1C2=CC=C(C=C2)CN3C4=NC(=NC=C4N(C3=N)CC(F)(F)F)C5=C(N=CN=C5OC)C6CC6)C(F)(F)F
CLogP	3
TPSA	104 Ų

Data compiled from [1]

2. Mechanism of Action

The therapeutic rationale for TNG-348 revolves around the inhibition of USP1, a critical enzyme in the DNA damage response (DDR) network. Understanding USP1's role provides context for its selection as an anti-cancer target.

2.1. The Role of USP1 in DNA Damage Repair and Cancer

Ubiquitin-Specific Protease 1 (USP1) is a deubiquitinating enzyme (DUB) that is found to be overexpressed in a variety of tumor cell types.[1] Its primary function involves the regulation of key DNA repair pathways, notably the Fanconi Anemia (FA) pathway and the translesion synthesis (TLS) pathway.[1] USP1 achieves this by deubiquitinating critical protein substrates, including Proliferating Cell Nuclear Antigen (PCNA) and the FANCD2-FANCI complex.[3] Deubiquitination of these proteins is essential for the proper progression of DNA replication and repair, particularly in response to DNA damage induced by genotoxic agents like interstrand crosslinking (ICL) agents.[18] Beyond its direct role in DNA repair, USP1 also contributes to cancer cell survival and aggressiveness by stabilizing proteins such as ID1 (Inhibitor of DNA binding 1) and CHEK1 (Checkpoint Kinase 1), thereby promoting cancer stem cell maintenance and radioresistance, particularly observed in glioblastoma.[18]

2.2. TNG-348 as a Selective, Allosteric, and Reversible USP1 Inhibitor

TNG-348 was engineered as a small molecule that specifically targets and inhibits USP1 through an allosteric mechanism.[1] This means TNG-348 binds to a site on the USP1 enzyme distinct from its catalytic active site, inducing a conformational change that inhibits its deubiquitinating activity. The interaction is characterized as selective for USP1 and reversible.[12] The precise nature of this allosteric binding was further elucidated by cryo-electron microscopy (cryo-EM) studies, which provided structural insights into the USP1-TNG348 complex.[13] Such allosteric inhibitors can offer advantages in terms of selectivity over active-site inhibitors, which often face challenges with off-target effects due to conserved active site motifs across enzyme families.

2.3. Molecular Interactions and Impact on Cellular Pathways

The inhibition of USP1's deubiquitinating function by TNG-348 has direct consequences on downstream cellular processes. A key outcome is the dose-dependent accumulation of ubiquitinated forms of USP1 substrates, particularly mono- and poly-ubiquitinated PCNA (ub-PCNA) and ubiquitinated FANCD2.[3] This accumulation, observable both in vitro and in vivo, serves as a pharmacodynamic biomarker indicating target engagement by TNG-348.

The persistence of ubiquitinated PCNA, due to USP1 inhibition, disrupts the normal progression of the TLS pathway. TLS is a DNA damage tolerance mechanism that allows DNA replication to proceed past DNA lesions that would otherwise stall the replication fork. TNG-348's interference with TLS occurs via the RAD18-dependent induction of ub-PCNA.[12] The dysregulation of these DNA repair and tolerance mechanisms leads to increased DNA damage, replication fork degradation, inhibition of DNA replication, and ultimately, apoptosis and decreased survival of tumor cells.[1] This cytotoxic effect is particularly pronounced in cancer cells with pre-existing deficiencies in homologous recombination (HRD), such as those harboring BRCA1 or BRCA2 mutations.

2.4. Rationale for Synthetic Lethality in Homologous Recombination Deficient (HRD+) Cancers

The primary therapeutic strategy for TNG-348 was based on the concept of synthetic lethality. Cancer cells with HRD, commonly found in a significant percentage of ovarian (up to 50%), breast (up to 25%), prostate (up to 10%), and pancreatic cancers (up to 5%), are unable to efficiently repair DNA double-strand breaks (DSBs) via the high-fidelity HR pathway.[4] Consequently, these cells become heavily reliant on alternative DNA repair and damage tolerance pathways, such as TLS, in which USP1 plays a critical role.[13] By inhibiting USP1, TNG-348 aims to cripple this compensatory pathway, leading to an overwhelming accumulation of DNA damage and selective cell death in HRD+ cancer cells, while largely sparing normal cells with intact HR function.[12] The targeting of DUBs like USP1 represents an emerging field in oncology. The development of TNG-348, despite its eventual discontinuation, has contributed to the validation of USP1 as a druggable target within this complex DDR network. The specific reliance of HRD+ cells on USP1-mediated TLS provided a strong rationale for the clinical investigation of TNG-348 in this patient population.

2.5. Comparison and Synergistic Potential with PARP Inhibitors (PARPi)

Poly(ADP-ribose) polymerase inhibitors (PARPi) are an established class of drugs that also exploit synthetic lethality in HRD+ cancers. While both TNG-348 and PARPi selectively target HRD tumor cells, their mechanisms of action are distinct.[4] PARPi primarily function by inhibiting PARP enzymes involved in single-strand break repair and by trapping PARP on DNA, leading to the formation of DSBs during replication, which are lethal in HR-deficient cells.

A crucial finding from preclinical studies was that the inhibition of USP1 by TNG348 acts synergistically with PARPi, such as olaparib.[4] This synergy results in enhanced anti-tumor effects in HRD tumor models. Furthermore, and of significant clinical relevance, TNG348 demonstrated the ability to overcome acquired resistance to PARPi in vivo.[12] CRISPR screen data indicated that knockout of PARP1, a mechanism of PARPi resistance, actually sensitizes cells to TNG348 treatment.[12] This suggests non-overlapping resistance mechanisms and a potential sequential vulnerability, where cells that become resistant to PARPi might become even more susceptible to USP1 inhibition. This mechanistic distinction and synergistic interaction formed a strong basis for evaluating TNG348 both as a monotherapy in PARPi-resistant settings and in combination with PARPi.

3. Preclinical Evaluation

The preclinical development of TNG-348 provided the foundational evidence for its advancement into human clinical trials. These studies focused on demonstrating its efficacy, target engagement, and synergistic potential, particularly in the context of HRD+ cancers.

3.1. Summary of in vitro and in vivo Efficacy Findings

In vitro, TNG-348 demonstrated selective cytotoxicity against cancer cell lines harboring BRCA1/2 mutations, which are a hallmark of HRD.[3] This selective killing of HRD+ cells aligns with the intended synthetic lethal mechanism of action. A key pharmacodynamic effect observed both in vitro and in vivo was the dose-dependent accumulation of ubiquitinated USP1 substrates, such as ub-PCNA and ub-FANCD2.[12] This served as a critical biomarker confirming that TNG-348 was engaging its target and modulating the USP1 pathway as intended.

In vivo, TNG-348 showed activity in xenograft models derived from human cancers. Notably, it was effective in models that exhibited primary resistance to PARP inhibitors and also in models that had acquired resistance to PARP inhibitors after prior treatment.[4] This activity in PARPi-resistant settings was a particularly significant finding, as PARPi resistance is a major clinical challenge. The ability of Tango Therapeutics to overcome high microsomal clearance and hERG (human Ether-à-go-go-Related Gene) liabilities, which often plague early drug candidates, during the discovery of TNG348 suggests significant medicinal chemistry efforts were undertaken to optimize its drug-like properties before these efficacy studies.[2] This makes the eventual clinical failure due to a different toxicity (liver toxicity) more notable, emphasizing the unpredictable nature of drug development.

3.2. Demonstration of Synergy with PARP Inhibitors

A cornerstone of the preclinical rationale for TNG-348 was its synergistic interaction with PARP inhibitors. Multiple studies demonstrated that combining TNG-348 with PARPi, such as olaparib, resulted in significantly enhanced anti-tumor effects compared to either agent alone in HRD tumor models.[4] This synergy was observed as increased tumor growth inhibition and, in some cases, tumor regression in various mouse xenograft models.[12] The strong preclinical synergy provided a compelling argument for clinical investigation of TNG-348 in combination with PARPi, either to enhance first-line efficacy or to overcome resistance.

3.3. Activity in PARP Inhibitor-Resistant Models

The ability of TNG-348 to overcome PARPi resistance was a key differentiator. Preclinical data showed that TNG-348, particularly when combined with a PARPi, was effective in human cancer models that had developed resistance to PARP inhibitors.[4] This suggested that TNG-348 could offer a therapeutic option for patients whose tumors had progressed on or after PARPi therapy, a population with limited treatment alternatives. This activity underpinned a significant part of the clinical development strategy for TNG-348.

4. Clinical Development Program: Trial NCT06065059

Based on the promising preclinical data, TNG-348 advanced into clinical development with the initiation of the NCT06065059 trial.

4.1. Study Title and Identifiers

The clinical trial was officially titled: "A Phase 1/2, Multi-Center, Open-Label Study to Evaluate the Safety, Tolerability, and Preliminary Antitumor Activity of TNG348 Single Agent and in Combination with a PARP Inhibitor in Patients with BRCA ½ Mutant or Other HRD+ Advanced or Metastatic Solid Tumors".19

The study is registered on ClinicalTrials.gov with the identifier NCT06065059.7 It is also referenced by other identifiers, including NCI-2023-10958 and TNG348-C101.21

4.2. Phase and Design

The trial was a Phase I/II study, employing an open-label, multi-center design.[2] The Phase 1 portion focused on dose escalation to determine safety and tolerability, while Phase 2 was intended for dose expansion to assess preliminary efficacy.[21] This integrated design is common in oncology to expedite early drug development.

4.3. Objectives

The primary objectives of the NCT06065059 trial were to evaluate the safety and tolerability of TNG-348, both as a single agent and in combination with the PARP inhibitor olaparib. A key goal was to determine the Maximum Tolerated Dose (MTD) and/or the Recommended Phase 2 Dose(s) (RP2D) for subsequent investigation.19

Secondary objectives included characterization of the pharmacokinetic (PK) profile of TNG-348 alone and in combination, and assessment of its preliminary antitumor activity.19

4.4. Target Patient Population and Key Eligibility Criteria

The study aimed to enroll adult patients (≥18 years of age) with advanced or metastatic solid tumors.19 A critical inclusion criterion was the documented presence of a BRCA1 or BRCA2 mutation, or other evidence of homologous recombination deficiency (HRD+), in the tumor, as identified by a validated sequencing test.4 While the trial was open to various HRD+ solid tumors, specific cancers targeted for recruitment included advanced breast, ovarian, pancreatic, and prostate cancer.19 This broad inclusion initially suggests a "basket trial" approach, common in early-phase studies of targeted therapies, to explore efficacy across different tumor types sharing a common molecular vulnerability.

Other key eligibility criteria included an Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1, measurable disease according to RECIST v1.1 criteria, and adequate organ and bone marrow function.19 Exclusion criteria encompassed uncontrolled intercurrent illnesses, active prior or concurrent malignancies (with some exceptions), CNS metastases associated with progressive neurological symptoms, myelodysplastic syndrome (MDS), clinically relevant cardiovascular disease, and active or chronic infections.19

4.5. Regulatory Milestones

The Investigational New Drug (IND) application for TNG-348 was cleared by the U.S. Food and Drug Administration (FDA), with the announcement made on September 06, 2023.[4] Following IND clearance, Tango Therapeutics planned to initiate the clinical trial in the first half of 2024.[4]

4.6. Intervention Details

TNG-348 was administered orally. The trial was designed to evaluate TNG-348 as a monotherapy and in combination with olaparib, an orally administered PARP inhibitor.[4] The Phase 1 portion of the study involved escalating doses of TNG-348 to establish the MTD/RP2D.[21] The Phase 2 portion would have utilized the determined RP2D(s).[21] The inclusion of both monotherapy and combination arms from early phases reflects a strategy to efficiently gather data on TNG-348's profile in different therapeutic contexts, likely driven by the strong preclinical rationale for both approaches.

4.7. Status at Discontinuation

The NCT06065059 trial was terminated in May 2024.[7] At the time of termination, the study was in the dose-escalation phase (Phase 1).[25] Crucially, no patients had yet received the combination therapy of TNG-348 and olaparib.[25] The discontinuation during this early stage, before the combination arm could be evaluated, represented a significant setback to the program, as the synergy with PARP inhibitors was a key component of its development strategy.

Table 2: Clinical Trial NCT06065059 Overview

Aspect	Details
Official Title	A Phase 1/2, Multi-Center, Open-Label Study to Evaluate the Safety, Tolerability, and Preliminary Antitumor Activity of TNG348 Single Agent and in Combination with a PARP Inhibitor in Patients with BRCA ½ Mutant or Other HRD+ Advanced or Metastatic Solid Tumors
Phase	Phase I/II
Status	Terminated (May 2024)
Primary Objectives	Evaluate safety and tolerability; Determine Maximum Tolerated Dose (MTD) / Recommended Phase 2 Dose (RP2D)
Key Eligibility Criteria	Adults (≥18 years) with advanced or metastatic solid tumors with documented BRCA1/2 mutation or other HRD+; ECOG performance status 0 or 1; Measurable disease (RECIST v1.1)
Intervention Arms Planned	TNG-348 monotherapy (oral, dose escalation); TNG-348 (oral, RP2D) + Olaparib (oral)

Data compiled from [4]

5. Discontinuation of Clinical Development

The clinical development of TNG-348 was halted prematurely due to safety concerns that emerged during the early stages of human testing.

5.1. Timeline and Announcement

Tango Therapeutics publicly announced the discontinuation of the TNG-348 program on May 23, 2024.[7] This decision was based on emerging safety data from the ongoing Phase 1/2 dose-escalation study (NCT06065059).[7] The termination occurred relatively early in the clinical trial process, highlighting the rapid identification of unacceptable toxicity.

5.2. Rationale for Termination: Liver Toxicity

The definitive reason for terminating the TNG-348 clinical program was the observation of unexpected liver toxicity in treated patients.[7] Specifically, Grade 3/4 liver function abnormalities were reported in patients who had remained on the study for longer than eight weeks.[7] Grade 3 and 4 adverse events are considered severe and potentially life-threatening, respectively, often leading to dose modification, interruption, or discontinuation of therapy. The manifestation of such significant liver toxicity in patients after more than two months of treatment suggested a potential cumulative or delayed adverse effect profile that was deemed unacceptable for continued development. This "unexpected" nature of the liver toxicity, despite the preclinical optimization efforts to mitigate other liabilities like high microsomal clearance and hERG effects [2], underscores the inherent challenges in predicting human-specific toxicities from animal models or in vitro systems.

5.3. Impact on Development Program and Stakeholders

The discontinuation of TNG-348 had immediate repercussions. Tango Therapeutics' stock price experienced a notable decline of 9.2% at market open on the day following the announcement, reflecting investor concern.7 Medivir AB, the company from which Tango Therapeutics had in-licensed the preclinical USP1 program, also acknowledged the termination.7

In response to this setback, Tango Therapeutics announced a strategic shift to focus its resources and capital on its other pipeline assets, with a particular emphasis on its PRMT5 inhibitor programs (TNG908 and TNG462).5 This refocusing involved a reduction in spending on its preclinical pipeline and the deferral of certain other clinical combination studies, aiming to extend its cash runway.5

Table 3: Summary of Liver Toxicity Findings Leading to Discontinuation of TNG-348

Aspect	Details
Clinical Trial	NCT06065059 (Phase 1/2 Dose Escalation)
Nature of Toxicity	Grade 3/4 liver function abnormalities
Affected Patient Cohort	Patients remaining on study longer than eight weeks
Timing of Observation	Data emerging from the dose escalation study, leading to program discontinuation announced on May 23, 2024
Company Statement	Decision based on emerging data and prioritization of patient safety; resources to be reallocated to other programs (e.g., PRMT5).

Data compiled from [5]

The observation that liver abnormalities became apparent in patients on study for longer than eight weeks [25] is critical. It suggests a potential for cumulative toxicity or a delayed onset that might not be captured in shorter-term preclinical toxicology studies or in early clinical cohorts if patients discontinue treatment rapidly due to disease progression. This highlights the importance of careful long-term safety monitoring even in early-phase oncology trials.

6. Patent Landscape

The intellectual property surrounding TNG-348 and the broader class of USP1 inhibitors involves multiple entities, reflecting a competitive research environment.

6.1. TNG-348 Specific Patents

Given that TNG-348 was developed by Tango Therapeutics from a program in-licensed from Medivir, patents covering this specific molecule or its near analogues would likely list these entities as assignees.

Patent application WO2022197892A1, titled "Purine Derivatives as Anticancer Agents," lists Tango Therapeutics, Inc. and Medivir AB as applicants.4 The international filing date was March 17, 2022, with a priority date of March 17, 2021, and it was published on September 22, 2022.18 The abstract describes compounds of "Formula (I)" and their pharmaceutically acceptable forms, intended for the prevention and treatment of various conditions, including cancer, likely through USP1 inhibition given the inventors and assignees involved with TNG-348.18 TNG-348 itself is a purine derivative, aligning with the subject matter of this patent.4

6.2. Broader USP1 Inhibitor Patents

The field of USP1 inhibition is active, with other companies also pursuing this target. For instance, patent WO2020132269A1, titled "Substituted Pyrazolopyrimidines and Substituted Purines and Their Use as Ubiquitin-specific-processing Protease 1 (usp1) Inhibitors," is assigned to KSQ Therapeutics, Inc. and has a priority date of December 20, 2018.28 This patent covers a different chemical class of USP1 inhibitors, including KSQ-4279 (also known as RO7623066 or example 143 from the patent), and their use in treating cancer.30

Other entities such as Forma Therapeutics Inc., Insilico Medicine IP Ltd., and Impact Therapeutics Inc. also hold patents related to USP1 inhibitors, indicating a diverse range of chemical scaffolds being explored to target USP1.28 This competitive patenting activity underscores the perceived therapeutic value of USP1 inhibition. The variety of patented chemical structures suggests that the optimal scaffold for USP1 inhibition, balancing potency, selectivity, and safety, may still be under investigation.

6.3. TNG-348 Chemical Information in Relation to Patents

TNG-348 is identified by CAS number 2839740-79-1.[2] Its chemical structure, characterized as a purine derivative, aligns with the scope of patent WO2022197892A1 assigned to Tango Therapeutics and Medivir.[4] The co-assignment of this patent reflects the collaborative origin of the TNG-348 program, stemming from Medivir's initial preclinical work and Tango's subsequent development efforts.[4]

7. Discussion and Future Perspectives

The development and subsequent discontinuation of TNG-348 offer valuable lessons for the field of oncology drug development, particularly for USP1 inhibitors and therapies targeting the DNA damage response.

7.1. Key Learnings from the TNG-348 Program

The TNG-348 program successfully demonstrated strong preclinical proof-of-concept for USP1 inhibition as a therapeutic strategy in HRD+ cancers. The observed synergy with PARP inhibitors and its activity in PARPi-resistant models were particularly compelling, addressing a significant unmet medical need.[4] However, the emergence of "unexpected liver toxicity" in the Phase I/II trial, leading to the program's termination, underscores the critical challenge of translating preclinical safety profiles into human clinical outcomes.[7] This occurred despite earlier medicinal chemistry efforts that successfully addressed other common drug development hurdles like high microsomal clearance and hERG liability for the initial lead compound.[2] This highlights the unpredictability of idiosyncratic or delayed-onset human toxicities that may not be apparent in standard preclinical toxicology packages. The discontinuation of TNG-348, despite its strong efficacy rationale, reiterates that safety remains paramount in drug development.

7.2. The Evolving Landscape of USP1 Inhibitors: Challenges and Opportunities

The journey of TNG-348 is not isolated; other USP1 inhibitors have also encountered developmental challenges. For instance, KSQ-4279 (RO7623066) reportedly showed limited efficacy and was associated with hematologic toxicity, with anemia being a common side effect.16 The differing primary toxicities observed between TNG-348 (liver) and KSQ-4279 (hematologic) may suggest that these are off-target effects related to their distinct chemical scaffolds rather than an inherent, unavoidable on-target toxicity of USP1 inhibition itself. This distinction is crucial because if the toxicities are scaffold-dependent, there remains a significant opportunity for developing safer USP1 inhibitors through novel chemical designs.

The primary challenge for the field is to definitively understand whether these observed toxicities are an on-target consequence of USP1 inhibition in specific tissues or off-target effects. If they are on-target, the therapeutic window for USP1 inhibitors might be narrow, necessitating careful patient selection, dose optimization, or perhaps intermittent dosing schedules.

The opportunity lies in leveraging the mechanistic understanding gained from TNG-348 and other early USP1 inhibitors to design next-generation compounds with improved safety profiles. An example is VRN19, which is reportedly being developed with the specific aim of avoiding the heme and liver toxicities seen with earlier USP1 inhibitors, by optimizing target binding affinity and solubility while minimizing toxicity.16

7.3. Considerations for Future Development of USP1-Targeted Therapies

The path forward for USP1-targeted therapies requires a multi-pronged approach. Firstly, there is a need for more predictive preclinical toxicology models, particularly those that can better anticipate human-specific liver and hematologic adverse events. Secondly, if on-target toxicities are a class effect, strategies to mitigate them, such as alternative dosing regimens (e.g., intermittent schedules) or co-administration with cytoprotective agents, may need to be explored. Thirdly, a more refined understanding of patient stratification beyond general HRD+ status or BRCA1/2 mutations could help identify subpopulations most likely to benefit with an acceptable risk profile. Finally, continued exploration of diverse chemical scaffolds is essential to identify USP1 inhibitors that retain potent efficacy but possess a more favorable therapeutic index, as exemplified by the aims for VRN19.[16] The discontinuation of TNG348 will likely intensify scrutiny on the safety profiles of new USP1 inhibitors entering clinical trials, potentially leading to more cautious dose-escalation strategies and enhanced monitoring for liver and hematologic function.

8. Conclusion

8.1. Concise Summary

TNG-348, an orally bioavailable, allosteric, and reversible inhibitor of USP1, was developed by Tango Therapeutics from a preclinical program in-licensed from Medivir. It was designed to induce synthetic lethality in cancers with homologous recombination deficiency (HRD+), such as those with BRCA1/2 mutations. Preclinical studies demonstrated promising efficacy, particularly in combination with PARP inhibitors and in models resistant to PARPi, by disrupting DNA damage tolerance pathways.

Following FDA IND clearance, TNG-348 entered a Phase I/II clinical trial (NCT06065059) to evaluate its safety and efficacy as a single agent and in combination with olaparib in patients with HRD+ solid tumors. However, in May 2024, the TNG-348 program was discontinued due to the observation of unexpected Grade 3/4 liver function abnormalities in patients who had been on study for longer than eight weeks.

8.2. Implications

The TNG-348 development story serves as a stark reminder of the complexities and high attrition rates inherent in oncology drug development. Even with a strong mechanistic rationale, promising preclinical data, and optimization against early ADME-Tox liabilities, unforeseen human-specific toxicities can derail a program.

While TNG-348 will not reach patients, the scientific knowledge generated regarding its mechanism of action, its interaction with USP1 (including structural data from cryo-EM studies 13), and its effects on DNA repair pathways remains valuable. This information contributes to the broader understanding of USP1 biology and the DNA damage response, potentially informing future drug discovery efforts targeting USP1 or related pathways. The experience with TNG-348, alongside other USP1 inhibitors like KSQ-4279, underscores the critical need to differentiate between on-target and off-target toxicities to guide the successful development of this class of therapeutic agents. The continued pursuit of USP1 inhibitors with improved safety profiles, such as VRN19 16, suggests that the therapeutic potential of targeting USP1 in HRD+ cancers is still considered significant, provided the safety challenges can be overcome.# TNG-348: A Comprehensive Review of a USP1 Inhibitor for HRD+ Cancers

1. Introduction to TNG-348

TNG-348 emerged as an orally bioavailable, small molecule inhibitor designed to target the ubiquitin-specific protease 1 (USP1).[1] Its development was rooted in the principle of synthetic lethality, aiming to exploit specific vulnerabilities in cancer cells with deficient DNA repair mechanisms.[2]

1.1. Chemical Identity and Basic Properties

TNG-348 is chemically identified by the CAS Number 2839740-79-1.[2] Its molecular formula is C27​H23​F6​N9​O, corresponding to a molecular weight of 603.5 g/mol.[2] For computational drug-like property assessment, its CLogP (a measure of lipophilicity) is 3, and its Topological Polar Surface Area (TPSA) is 104 Ų.[2] These parameters are crucial in early drug development for predicting pharmacokinetic properties such as absorption and membrane permeability. The SMILES (Simplified Molecular Input Line Entry System) string for TNG-348 is CN1C=C(N=C1C2=CC=C(C=C2)CN3C4=NC(=NC=C4N(C3=N)CC(F)(F)F)C5=C(N=CN=C5OC)C6CC6)C(F)(F)F.[2]

1.2. Developer and Origin

TNG-348 was developed by Tango Therapeutics, a clinical-stage biotechnology company specializing in precision oncology and synthetic lethality.[2] The foundational preclinical USP1 inhibitor program that led to the discovery and development of TNG-348 was in-licensed by Tango Therapeutics from Medivir AB in 2020.[4] This collaboration highlights a common strategy in the pharmaceutical industry where specialized biotechnology companies advance promising preclinical assets originating from other research entities. Tango Therapeutics' focus on synthetic lethality suggests that the precursor program from Medivir demonstrated significant potential in selectively targeting cancer cells with specific DNA repair deficiencies, aligning with Tango's core research and development strategy.

1.3. Therapeutic Class

TNG-348 is classified as a Ubiquitin-Specific Protease 1 (USP1) inhibitor.[1] More specifically, it functions as an allosteric, selective, and reversible inhibitor of the USP1 enzyme.[12] USP1 is a deubiquitinating enzyme (DUB) that plays a critical role in DNA damage repair processes. The development of an allosteric, selective, and reversible inhibitor like TNG-348 likely represented a deliberate drug design approach. This strategy may have been aimed at achieving greater target specificity and potentially a more favorable safety profile compared to earlier or non-allosteric USP1 inhibitors, although, as later discussed, safety objectives were not ultimately met.

Table 1: TNG-348 Key Characteristics

Characteristic	Details
Name	TNG-348
Alternative Designations	Company Code: TNG348
Developer	Tango Therapeutics
Origin	Preclinical program in-licensed from Medivir
CAS Number	2839740-79-1
Molecular Formula	C27​H23​F6​N9​O
Molecular Weight	603.5 g/mol
Therapeutic Class	USP1 inhibitor (allosteric, selective, reversible)
SMILES String	CN1C=C(N=C1C2=CC=C(C=C2)CN3C4=NC(=NC=C4N(C3=N)CC(F)(F)F)C5=C(N=CN=C5OC)C6CC6)C(F)(F)F
CLogP	3
TPSA	104 Ų

Data compiled from [1]

2. Mechanism of Action

The therapeutic rationale for TNG-348 revolves around the inhibition of USP1, a critical enzyme in the DNA damage response (DDR) network. Understanding USP1's role provides context for its selection as an anti-cancer target.

2.1. The Role of USP1 in DNA Damage Repair and Cancer

Ubiquitin-Specific Protease 1 (USP1) is a deubiquitinating enzyme (DUB) that is found to be overexpressed in a variety of tumor cell types.[1] Its primary function involves the regulation of key DNA repair pathways, notably the Fanconi Anemia (FA) pathway and the translesion synthesis (TLS) pathway.[1] USP1 achieves this by deubiquitinating critical protein substrates, including Proliferating Cell Nuclear Antigen (PCNA) and the FANCD2-FANCI complex.[3] Deubiquitination of these proteins is essential for the proper progression of DNA replication and repair, particularly in response to DNA damage induced by genotoxic agents like interstrand crosslinking (ICL) agents.[18] Beyond its direct role in DNA repair, USP1 also contributes to cancer cell survival and aggressiveness by stabilizing proteins such as ID1 (Inhibitor of DNA binding 1) and CHEK1 (Checkpoint Kinase 1), thereby promoting cancer stem cell maintenance and radioresistance, particularly observed in glioblastoma.[18]

2.2. TNG-348 as a Selective, Allosteric, and Reversible USP1 Inhibitor

TNG-348 was engineered as a small molecule that specifically targets and inhibits USP1 through an allosteric mechanism.[1] This means TNG-348 binds to a site on the USP1 enzyme distinct from its catalytic active site, inducing a conformational change that inhibits its deubiquitinating activity. The interaction is characterized as selective for USP1 and reversible.[12] The precise nature of this allosteric binding was further elucidated by cryo-electron microscopy (cryo-EM) studies, which provided structural insights into the USP1-TNG348 complex.[13] Such allosteric inhibitors can offer advantages in terms of selectivity over active-site inhibitors, which often face challenges with off-target effects due to conserved active site motifs across enzyme families.

2.3. Molecular Interactions and Impact on Cellular Pathways

The inhibition of USP1's deubiquitinating function by TNG-348 has direct consequences on downstream cellular processes. A key outcome is the dose-dependent accumulation of ubiquitinated forms of USP1 substrates, particularly mono- and poly-ubiquitinated PCNA (ub-PCNA) and ubiquitinated FANCD2.[3] This accumulation, observable both in vitro and in vivo, serves as a pharmacodynamic biomarker indicating target engagement by TNG-348.

The persistence of ubiquitinated PCNA, due to USP1 inhibition, disrupts the normal progression of the TLS pathway. TLS is a DNA damage tolerance mechanism that allows DNA replication to proceed past DNA lesions that would otherwise stall the replication fork. TNG-348's interference with TLS occurs via the RAD18-dependent induction of ub-PCNA.[12] The dysregulation of these DNA repair and tolerance mechanisms leads to increased DNA damage, replication fork degradation, inhibition of DNA replication, and ultimately, apoptosis and decreased survival of tumor cells.[1] This cytotoxic effect is particularly pronounced in cancer cells with pre-existing deficiencies in homologous recombination (HRD), such as those harboring BRCA1 or BRCA2 mutations.

2.4. Rationale for Synthetic Lethality in Homologous Recombination Deficient (HRD+) Cancers

The primary therapeutic strategy for TNG-348 was based on the concept of synthetic lethality. Cancer cells with HRD, commonly found in a significant percentage of ovarian (up to 50%), breast (up to 25%), prostate (up to 10%), and pancreatic cancers (up to 5%), are unable to efficiently repair DNA double-strand breaks (DSBs) via the high-fidelity HR pathway.[4] Consequently, these cells become heavily reliant on alternative DNA repair and damage tolerance pathways, such as TLS, in which USP1 plays a critical role.[13] By inhibiting USP1, TNG-348 aims to cripple this compensatory pathway, leading to an overwhelming accumulation of DNA damage and selective cell death in HRD+ cancer cells, while largely sparing normal cells with intact HR function.[12] The targeting of DUBs like USP1 represents an emerging field in oncology. The development of TNG-348, despite its eventual discontinuation, has contributed to the validation of USP1 as a druggable target within this complex DDR network. The specific reliance of HRD+ cells on USP1-mediated TLS provided a strong rationale for the clinical investigation of TNG-348 in this patient population.

2.5. Comparison and Synergistic Potential with PARP Inhibitors (PARPi)

Poly(ADP-ribose) polymerase inhibitors (PARPi) are an established class of drugs that also exploit synthetic lethality in HRD+ cancers. While both TNG-348 and PARPi selectively target HRD tumor cells, their mechanisms of action are distinct.[4] PARPi primarily function by inhibiting PARP enzymes involved in single-strand break repair and by trapping PARP on DNA, leading to the formation of DSBs during replication, which are lethal in HR-deficient cells.

A crucial finding from preclinical studies was that the inhibition of USP1 by TNG348 acts synergistically with PARPi, such as olaparib.[4] This synergy results in enhanced anti-tumor effects in HRD tumor models. Furthermore, and of significant clinical relevance, TNG348 demonstrated the ability to overcome acquired resistance to PARPi in vivo.[12] CRISPR screen data indicated that knockout of PARP1, a mechanism of PARPi resistance, actually sensitizes cells to TNG348 treatment.[12] This suggests non-overlapping resistance mechanisms and a potential sequential vulnerability, where cells that become resistant to PARPi might become even more susceptible to USP1 inhibition. This mechanistic distinction and synergistic interaction formed a strong basis for evaluating TNG-348 both as a monotherapy in PARPi-resistant settings and in combination with PARPi.

3. Preclinical Evaluation

The preclinical development of TNG-348 provided the foundational evidence for its advancement into human clinical trials. These studies focused on demonstrating its efficacy, target engagement, and synergistic potential, particularly in the context of HRD+ cancers.

3.1. Summary of in vitro and in vivo Efficacy Findings

In vitro, TNG-348 demonstrated selective cytotoxicity against cancer cell lines harboring BRCA1/2 mutations, which are a hallmark of HRD.[3] This selective killing of HRD+ cells aligns with the intended synthetic lethal mechanism of action. A key pharmacodynamic effect observed both in vitro and in vivo was the dose-dependent accumulation of ubiquitinated USP1 substrates, such as ub-PCNA and ub-FANCD2.[12] This served as a critical biomarker confirming that TNG-348 was engaging its target and modulating the USP1 pathway as intended.

In vivo, TNG-348 showed activity in xenograft models derived from human cancers. Notably, it was effective in models that exhibited primary resistance to PARP inhibitors and also in models that had acquired resistance to PARP inhibitors after prior treatment.[4] This activity in PARPi-resistant settings was a particularly significant finding, as PARPi resistance is a major clinical challenge. The ability of Tango Therapeutics to overcome high microsomal clearance and hERG (human Ether-à-go-go-Related Gene) liabilities, which often plague early drug candidates, during the discovery of TNG348 suggests significant medicinal chemistry efforts were undertaken to optimize its drug-like properties before these efficacy studies.[2] This makes the eventual clinical failure due to a different toxicity (liver toxicity) more notable, emphasizing the unpredictable nature of drug development.

3.2. Demonstration of Synergy with PARP Inhibitors

A cornerstone of the preclinical rationale for TNG-348 was its synergistic interaction with PARP inhibitors. Multiple studies demonstrated that combining TNG-348 with PARPi, such as olaparib, resulted in significantly enhanced anti-tumor effects compared to either agent alone in HRD tumor models.[4] This synergy was observed as increased tumor growth inhibition and, in some cases, tumor regression in various mouse xenograft models.[12] The strong preclinical synergy provided a compelling argument for clinical investigation of TNG-348 in combination with PARPi, either to enhance first-line efficacy or to overcome resistance.

3.3. Activity in PARP Inhibitor-Resistant Models

The ability of TNG-348 to overcome PARPi resistance was a key differentiator. Preclinical data showed that TNG-348, particularly when combined with a PARPi, was effective in human cancer models that had developed resistance to PARP inhibitors.[4] This suggested that TNG-348 could offer a therapeutic option for patients whose tumors had progressed on or after PARPi therapy, a population with limited treatment alternatives. This activity underpinned a significant part of the clinical development strategy for TNG-348.

4. Clinical Development Program: Trial NCT06065059

Based on the promising preclinical data, TNG-348 advanced into clinical development with the initiation of the NCT06065059 trial.

4.1. Study Title and Identifiers

The clinical trial was officially titled: "A Phase 1/2, Multi-Center, Open-Label Study to Evaluate the Safety, Tolerability, and Preliminary Antitumor Activity of TNG348 Single Agent and in Combination with a PARP Inhibitor in Patients with BRCA ½ Mutant or Other HRD+ Advanced or Metastatic Solid Tumors".19

The study is registered on ClinicalTrials.gov with the identifier NCT06065059.7 It is also referenced by other identifiers, including NCI-2023-10958 and TNG348-C101.21

4.2. Phase and Design

The trial was a Phase I/II study, employing an open-label, multi-center design.[2] The Phase 1 portion focused on dose escalation to determine safety and tolerability, while Phase 2 was intended for dose expansion to assess preliminary efficacy.[21] This integrated design is common in oncology to expedite early drug development.

4.3. Objectives

The primary objectives of the NCT06065059 trial were to evaluate the safety and tolerability of TNG-348, both as a single agent and in combination with the PARP inhibitor olaparib. A key goal was to determine the Maximum Tolerated Dose (MTD) and/or the Recommended Phase 2 Dose(s) (RP2D) for subsequent investigation.19

Secondary objectives included characterization of the pharmacokinetic (PK) profile of TNG-348 alone and in combination, and assessment of its preliminary antitumor activity.19

4.4. Target Patient Population and Key Eligibility Criteria

The study aimed to enroll adult patients (≥18 years of age) with advanced or metastatic solid tumors.19 A critical inclusion criterion was the documented presence of a BRCA1 or BRCA2 mutation, or other evidence of homologous recombination deficiency (HRD+), in the tumor, as identified by a validated sequencing test.4 While the trial was open to various HRD+ solid tumors, specific cancers targeted for recruitment included advanced breast, ovarian, pancreatic, and prostate cancer.19 This broad inclusion initially suggests a "basket trial" approach, common in early-phase studies of targeted therapies, to explore efficacy across different tumor types sharing a common molecular vulnerability.

Other key eligibility criteria included an Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1, measurable disease according to RECIST v1.1 criteria, and adequate organ and bone marrow function.19 Exclusion criteria encompassed uncontrolled intercurrent illnesses, active prior or concurrent malignancies (with some exceptions), CNS metastases associated with progressive neurological symptoms, myelodysplastic syndrome (MDS), clinically relevant cardiovascular disease, and active or chronic infections.19

4.5. Regulatory Milestones

The Investigational New Drug (IND) application for TNG-348 was cleared by the U.S. Food and Drug Administration (FDA), with the announcement made on September 06, 2023.[4] Following IND clearance, Tango Therapeutics planned to initiate the clinical trial in the first half of 2024.[4]

4.6. Intervention Details

TNG-348 was administered orally. The trial was designed to evaluate TNG-348 as a monotherapy and in combination with olaparib, an orally administered PARP inhibitor.[4] The Phase 1 portion of the study involved escalating doses of TNG-348 to establish the MTD/RP2D.[21] The Phase 2 portion would have utilized the determined RP2D(s).[21] The inclusion of both monotherapy and combination arms from early phases reflects a strategy to efficiently gather data on TNG-348's profile in different therapeutic contexts, likely driven by the strong preclinical rationale for both approaches.

4.7. Status at Discontinuation

The NCT06065059 trial was terminated in May 2024.[7] At the time of termination, the study was in the dose-escalation phase (Phase 1).[25] Crucially, no patients had yet received the combination therapy of TNG-348 and olaparib.[25] The discontinuation during this early stage, before the combination arm could be evaluated, represented a significant setback to the program, as the synergy with PARP inhibitors was a key component of its development strategy.

Table 2: Clinical Trial NCT06065059 Overview

Aspect	Details
Official Title	A Phase 1/2, Multi-Center, Open-Label Study to Evaluate the Safety, Tolerability, and Preliminary Antitumor Activity of TNG348 Single Agent and in Combination with a PARP Inhibitor in Patients with BRCA ½ Mutant or Other HRD+ Advanced or Metastatic Solid Tumors
Phase	Phase I/II
Status	Terminated (May 2024)
Primary Objectives	Evaluate safety and tolerability; Determine Maximum Tolerated Dose (MTD) / Recommended Phase 2 Dose (RP2D)
Key Eligibility Criteria	Adults (≥18 years) with advanced or metastatic solid tumors with documented BRCA1/2 mutation or other HRD+; ECOG performance status 0 or 1; Measurable disease (RECIST v1.1)
Intervention Arms Planned	TNG-348 monotherapy (oral, dose escalation); TNG-348 (oral, RP2D) + Olaparib (oral)

Data compiled from [4]

5. Discontinuation of Clinical Development

The clinical development of TNG-348 was halted prematurely due to safety concerns that emerged during the early stages of human testing.

5.1. Timeline and Announcement

Tango Therapeutics publicly announced the discontinuation of the TNG-348 program on May 23, 2024.[7] This decision was based on emerging safety data from the ongoing Phase 1/2 dose-escalation study (NCT06065059).[7] The termination occurred relatively early in the clinical trial process, highlighting the rapid identification of unacceptable toxicity.

5.2. Rationale for Termination: Liver Toxicity

The definitive reason for terminating the TNG-348 clinical program was the observation of unexpected liver toxicity in treated patients.[7] Specifically, Grade 3/4 liver function abnormalities were reported in patients who had remained on the study for longer than eight weeks.[7] Grade 3 and 4 adverse events are considered severe and potentially life-threatening, respectively, often leading to dose modification, interruption, or discontinuation of therapy. The manifestation of such significant liver toxicity in patients after more than two months of treatment suggested a potential cumulative or delayed adverse effect profile that was deemed unacceptable for continued development. This "unexpected" nature of the liver toxicity, despite the preclinical optimization efforts to mitigate other liabilities like high microsomal clearance and hERG effects [2], underscores the inherent challenges in predicting human-specific toxicities from animal models or in vitro systems.

5.3. Impact on Development Program and Stakeholders

The discontinuation of TNG-348 had immediate repercussions. Tango Therapeutics' stock price experienced a notable decline of 9.2% at market open on the day following the announcement, reflecting investor concern.7 Medivir AB, the company from which Tango Therapeutics had in-licensed the preclinical USP1 program, also acknowledged the discontinuation.7

In response to this setback, Tango Therapeutics announced a strategic shift to focus its resources and capital on its other pipeline assets, with a particular emphasis on its PRMT5 inhibitor programs (TNG908 and TNG462).5 This refocusing involved a reduction in spending on its preclinical pipeline and the deferral of certain other clinical combination studies, aiming to extend its cash runway.5

Table 3: Summary of Liver Toxicity Findings Leading to Discontinuation of TNG-348

Aspect	Details
Clinical Trial	NCT06065059 (Phase 1/2 Dose Escalation)
Nature of Toxicity	Grade 3/4 liver function abnormalities
Affected Patient Cohort	Patients remaining on study longer than eight weeks
Timing of Observation	Data emerging from the dose escalation study, leading to program discontinuation announced on May 23, 2024
Company Statement	Decision based on emerging data and prioritization of patient safety; resources to be reallocated to other programs (e.g., PRMT5).

Data compiled from [5]

The observation that liver abnormalities became apparent in patients on study for longer than eight weeks [25] is critical. It suggests a potential for cumulative toxicity or a delayed onset that might not be captured in shorter-term preclinical toxicology studies or in early clinical cohorts if patients discontinue treatment rapidly due to disease progression. This highlights the importance of careful long-term safety monitoring even in early-phase oncology trials.

6. Patent Landscape

The intellectual property surrounding TNG-348 and the broader class of USP1 inhibitors involves multiple entities, reflecting a competitive research environment.

6.1. TNG-348 Specific Patents

Given that TNG-348 was developed by Tango Therapeutics from a program in-licensed from Medivir, patents covering this specific molecule or its near analogues would likely list these entities as assignees.

Patent application WO2022197892A1, titled "Purine Derivatives as Anticancer Agents," lists Tango Therapeutics, Inc. and Medivir AB as applicants.4 The international filing date was March 17, 2022, with a priority date of March 17, 2021, and it was published on September 22, 2022.18 The abstract describes compounds of "Formula (I)" and their pharmaceutically acceptable forms, intended for the prevention and treatment of various conditions, including cancer, likely through USP1 inhibition given the inventors and assignees involved with TNG-348.18 TNG-348 itself is a purine derivative, aligning with the subject matter of this patent.4

6.2. Broader USP1 Inhibitor Patents

The field of USP1 inhibition is active, with other companies also pursuing this target. For instance, patent WO2020132269A1, titled "Substituted Pyrazolopyrimidines and Substituted Purines and Their Use as Ubiquitin-specific-processing Protease 1 (usp1) Inhibitors," is assigned to KSQ Therapeutics, Inc. and has a priority date of December 20, 2018.28 This patent covers a different chemical class of USP1 inhibitors, including KSQ-4279 (also known as RO7623066 or example 143 from the patent), and their use in treating cancer.30

Other entities such as Forma Therapeutics Inc., Insilico Medicine IP Ltd., and Impact Therapeutics Inc. also hold patents related to USP1 inhibitors, indicating a diverse range of chemical scaffolds being explored to target USP1.28 This competitive patenting activity underscores the perceived therapeutic value of USP1 inhibition. The variety of patented chemical structures suggests that the optimal scaffold for USP1 inhibition, balancing potency, selectivity, and safety, may still be under investigation.

6.3. TNG-348 Chemical Information in Relation to Patents

TNG-348 is identified by CAS number 2839740-79-1.[2] Its chemical structure, characterized as a purine derivative, aligns with the scope of patent WO2022197892A1 assigned to Tango Therapeutics and Medivir.[4] The co-assignment of this patent reflects the collaborative origin of the TNG-348 program, stemming from Medivir's initial preclinical work and Tango's subsequent development efforts.[4]

7. Discussion and Future Perspectives

The development and subsequent discontinuation of TNG-348 offer valuable lessons for the field of oncology drug development, particularly for USP1 inhibitors and therapies targeting the DNA damage response.

7.1. Key Learnings from the TNG-348 Program

The TNG-348 program successfully demonstrated strong preclinical proof-of-concept for USP1 inhibition as a therapeutic strategy in HRD+ cancers. The observed synergy with PARP inhibitors and its activity in PARPi-resistant models were particularly compelling, addressing a significant unmet medical need.[4] However, the emergence of "unexpected liver toxicity" in the Phase I/II trial, leading to the program's termination, underscores the critical challenge of translating preclinical safety profiles into human clinical outcomes.[7] This occurred despite earlier medicinal chemistry efforts that successfully addressed other common drug development hurdles like high microsomal clearance and hERG liability for the initial lead compound.[2] This highlights the unpredictability of idiosyncratic or delayed-onset human toxicities that may not be apparent in standard preclinical toxicology packages. The discontinuation of TNG-348, despite its strong efficacy rationale, reiterates that safety remains paramount in drug development.

7.2. The Evolving Landscape of USP1 Inhibitors: Challenges and Opportunities

The journey of TNG-348 is not isolated; other USP1 inhibitors have also encountered developmental challenges. For instance, KSQ-4279 (RO7623066) reportedly showed limited efficacy and was associated with hematologic toxicity, with anemia being a common side effect.16 The differing primary toxicities observed between TNG-348 (liver) and KSQ-4279 (hematologic) may suggest that these are off-target effects related to their distinct chemical scaffolds rather than an inherent, unavoidable on-target toxicity of USP1 inhibition itself. This distinction is crucial because if the toxicities are scaffold-dependent, there remains a significant opportunity for developing safer USP1 inhibitors through novel chemical designs.

The primary challenge for the field is to definitively understand whether these observed toxicities are an on-target consequence of USP1 inhibition in specific tissues or off-target effects. If they are on-target, the therapeutic window for USP1 inhibitors might be narrow, necessitating careful patient selection, dose optimization, or perhaps intermittent dosing schedules.

The opportunity lies in leveraging the mechanistic understanding gained from TNG-348 and other early USP1 inhibitors to design next-generation compounds with improved safety profiles. An example is VRN19, which is reportedly being developed with the specific aim of avoiding the heme and liver toxicities seen with earlier USP1 inhibitors, by optimizing target binding affinity and solubility while minimizing toxicity.16

7.3. Considerations for Future Development of USP1-Targeted Therapies

The path forward for USP1-targeted therapies requires a multi-pronged approach. Firstly, there is a need for more predictive preclinical toxicology models, particularly those that can better anticipate human-specific liver and hematologic adverse events. Secondly, if on-target toxicities are a class effect, strategies to mitigate them, such as alternative dosing regimens (e.g., intermittent schedules) or co-administration with cytoprotective agents, may need to be explored. Thirdly, a more refined understanding of patient stratification beyond general HRD+ status or BRCA1/2 mutations could help identify subpopulations most likely to benefit with an acceptable risk profile. Finally, continued exploration of diverse chemical scaffolds is essential to identify USP1 inhibitors that retain potent efficacy but possess a more favorable therapeutic index, as exemplified by the aims for VRN19.[16] The discontinuation of TNG348 will likely intensify scrutiny on the safety profiles of new USP1 inhibitors entering clinical trials, potentially leading to more cautious dose-escalation strategies and enhanced monitoring for liver and hematologic function.

8. Conclusion

8.1. Concise Summary

TNG-348, an orally bioavailable, allosteric USP1 inhibitor developed by Tango Therapeutics, showed strong preclinical promise for treating HRD+ cancers, particularly in combination with PARP inhibitors and in PARPi-resistant settings. Its mechanism centered on inducing synthetic lethality by disrupting DNA damage tolerance pathways in cancer cells with deficient homologous recombination.

Despite FDA IND clearance and initiation of a Phase I/II clinical trial (NCT06065059), the TNG-348 program was discontinued in May 2024 due to unexpected Grade 3/4 liver toxicity observed in patients.

8.2. Implications

The TNG-348 story underscores the complexities of translating preclinical findings into clinical success and the critical importance of human safety data. While TNG-348 itself will not reach patients, the research invested in it (elucidating its mechanism, synergy with PARPi, structural biology [13]) still contributes valuable knowledge to the field of DNA damage response and USP1 biology. This information may aid future drug discovery efforts for this target or related pathways. The experience with TNG-348, alongside other USP1 inhibitors, highlights both the therapeutic potential of targeting USP1 and the significant safety challenges that need to be overcome for this drug class to succeed.

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