Slide 1: 4D-710 for Cystic Fibrosis: Phase 1 Interim Data
4D-710 for Cystic Fibrosis: Phase 1 Interim Data December 17, 2025 © 2025 4D Molecular © 2025 4DTherapeutics. Molecular Therapeutics. All Rights Reserved. All Rights Reserved.
Slide 2: Legal Disclaimer
Legal Disclaimer This Presentation contains forward looking statements that involve substantial risks and uncertainties. All statements, other than statements of historical facts, contained in this Presentation, including statements regarding our clinical development plans, strategy, future operatio
Slide 3: Today's Speakers
Today’s Speakers Jennifer L. Taylor-Cousar, Felix Ratjen, David Kirn, M.D. M.D., MSCS M.D., Ph.D, FRCP(C), FERS Co-founder and Chief Executive Officer Professor, Departments of Medicine and Professor of Pediatrics at the University of Pediatrics, and Co-Director, Adult Cystic Toronto, Program Head a
Slide 4: Phase 1 Interim Data: Key Takeaways from Lower Dose Cohorts 3 & 4
Phase 1 Interim Data: Key Takeaways from Lower Dose Cohorts 3 & 4 SAFETY DATA: Well Tolerated with 4 to 24 Months follow-up LUNG FUNCTION: Clinically Meaningful Activity (FEV1, LCI2.5) 4D-710 Durable, Redosable, Variant- QUALITY-OF-LIFE: Clinically Meaningful Activity (CFQ-R-R) Agnostic, Disease-Mod
Slide 5: Cystic Fibrosis Lung Disease: High Unmet Need Despite Modulators
Cystic Fibrosis Lung Disease: High Unmet Need Despite Modulators Lung Disease Burden CF Epidemiology2,3 Ineligible or Burdensome ~15k Intolerant Daily to CFTR Supportive Care: modulators ~20k Airway clearance ~70k Suboptimal responders4 (~100 mins), Responsive to CFTR modulators inhaled antibiotics
Slide 6: Conventional Gene Therapy Trials in CF Lung Failed Due to Lack of Effective Gene Delivery
Conventional Gene Therapy Trials in CF Lung Failed Due to Lack of Effective Gene Delivery & Expression ▪ Prior AAV Gene Therapy Study Design1,2 o AAV2-based CFTR gene therapy (tgAAVCF) o Randomized Phase 2 trial (n=51) o Repeat aerosol administration on days 1 & 30 ▪ Clinical Data Reports o Safe & w
Slide 7: 4D-710 Design: Aerosol Delivery of Novel A101 Vector Invented with Directed Evolution
4D-710 Design: Aerosol Delivery of Novel A101 Vector Invented with Directed Evolution Novel AAV Vector: A101 Overcome Potential Barriers Enhanced Transduction of Airway Cells ✓ Mucus penetrant Airway ✓ Transduction of delivery multiple airway cell types AeroEclipse® II ✓ Resistance to pre- Breath-ac
Slide 8: 4D-710 Design: Highly Functional CFTR Transgene Payload
4D-710 Design: Highly Functional CFTR Transgene Payload Novel AAV Vector: A101 Overcome Potential Barriers Enhanced Transduction of Airway Cells ✓ Mucus penetrant Airway ✓ Transduction of delivery multiple airway cell types AeroEclipse® II ✓ Resistance to pre- Breath-actuated nebulizer (FDA approved
Slide 9: 4D-710 Design: Durable, Redosable, Variant-agnostic Disease-modifying Therapy
4D-710 Design: Durable, Redosable,Variant-agnostic Disease-modifying Therapy for CF Lung Disease Novel AAV Vector: A101 Overcome Potential Barriers Enhanced Transduction of Airway Cells ✓ Mucus penetrant ✓ Transduction of 4D-710 Airway delivery multiple airway cell CMV173 CFTR∆R types PROMOTER TRANS
Slide 10: 4D-710 Interim Phase 1 Data: Study Design, Baseline Characteristics, Safety
4D-710 Interim Phase 1 Data: Study Design, Baseline Characteristics, Safety Jennifer L. Taylor-Cousar, M.D., MSCS, National Jewish Health Data Cutoff: December 1, 2025 © 2025 4D Molecular © 2025 4DTherapeutics. Molecular Therapeutics. All Rights Reserved. All Rights Reserved.
Slide 11: Phase 1: Dose Selection Framework for Further Development of 4D-710
Phase 1: Dose Selection Framework for Further Development of 4D-710 Focus of Phase 1: Focus of Phase 2: Dose Finding Characterize Clinical Activity Safety Stability or improvement in large-/mid- airway disease (ppFEV1) Physiologically relevant Phase 2 Improvement in small airway disease (LC12.5) CFT
Slide 12: Initial Phase 1 Protocol: Designed to Characterize Safety & Expression
Initial Phase 1 Protocol: Designed to Characterize Safety & Expression to Inform Dose Selection Phase 1 (Dose Ranging) Key Endpoints: Cohort 2 2E15 vg ▪ Safety & tolerability (n=4) Cohort 1 ▪ Lung biomarkers 1E15 vg (bronchoscopy): (n=3) o Weeks 4-8 ▪ Clinical activity (lung function, quality of lif
Slide 13: Protocol Amendment: Endpoints Added for Pulmonary Clinical Activity Assessments
Protocol Amendment: Endpoints Added for Pulmonary Clinical Activity Assessments, 2nd Biopsy to Assess Durability at 1 Yr & Beyond Phase 1 (Dose Ranging) Key Endpoints: Cohort 2 2E15 vg ▪ Safety & tolerability (n=4) Cohort 1 ▪ Lung biomarkers 1E15 vg (bronchoscopy): (n=3) o Weeks 4-8 Cohort 3 o ≥1 ye
Slide 14: AEROW Clinical Trial: Lower Dose Cohorts 3 & 4 (N=9)
AEROW Clinical Trial: Lower Dose Cohorts 3 & 4 (N=9) Demographics & Baseline Characteristics Cohort 3: 5E14 vg Cohort 4: 2.5E14 vg Participant 1 2 3 1 2 3 4 5 6 Number Age, y 42 40 34 26 54 37 56 33 37 Sex Female Female Male Male Female Female Male Male Male CFTR mod. status Intolerant Ineligible In
Slide 15: 4D-710 Safety & Tolerability Day 1-14: Well-Tolerated in Lower Dose Cohorts
4D-710 Safety & Tolerability Day 1-14: Well-Tolerated in Lower Dose Cohorts with Transient & Generally Mild AEs Typical of Nebulized Therapies Day 1 Dosing Activities ppFEV1 4D-710–Related AE Routine ACT ▪ Participant 1: throat irritation 1 (Historical) after dosing (Grade 1, ~5 sec), increased prod
Slide 16: 4D-710 Safety & Tolerability: No Pulmonary Related AEs After Day 14
4D-710 Safety & Tolerability: Well-Tolerated in Lower Dose Cohorts No Pulmonary 4D-710–related AEs After Day 14 4D-710–Related Pulmonary Adverse Events None Grade 1 Grade 2 Grade 3 Grade 4 1 5E14 vg 2 3 1 2 3 2.5E14 vg 4 5 6 D14- Month (post dose): M1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
Slide 17: 4D-710 Interim Phase 1 Data: Transgene Expression
4D-710 Interim Phase 1 Data: Transgene Expression Jennifer L.Taylor-Cousar, M.D., MSCS National Jewish Health Data Cutoff: December 1, 2025 © 2025 4D Molecular © 2025 4DTherapeutics. Molecular Therapeutics. All Rights Reserved. All Rights Reserved.
Slide 18: Lung Biopsy Analysis Methods: Rationale & Endpoints
Lung Biopsy Analysis Methods: Rationale & Endpoints CFTR/∆R mRNA Expression: CFTR/∆R Protein Expression: ISH IHC Highly Sensitive Strength Highly Specific Intracellular Localization % of airway cell (+) Intensity Readouts % of airway cell (+) Localization Cell types Qualitative Quantitative Evaluati
Slide 19: Target Expression Profile for 4D-710 CFTR: Phase 2 Dose Selection
Target Expression Profile for 4D-710 CFTR: Phase 2 Dose Selection Transgene Expression Analyses from Biopsies & Brushings ~1 Month Post-Dose Target Airway Target Airway Dose Selection Criteria: BIOPSY Profile BRUSHING Profile CFTR∆R RNA expression (ISH) 100% of samples 10-25% airway cells1,2 positiv
Slide 20: CFTR Transgene RNA Expression in Airway Biopsies (ISH): Controls
CFTR Transgene RNA Expression in Airway Biopsies (ISH): Non-CF Lung & Non-Treated CF Biopsy Controls CFTR RNA (ISH): Airway Epithelial Cells 80 ▪ Commercial non-CF samples Mean % (+) epithelial cells 52.7 60 positive for endogenous CFTR 39.0 mRNA expression, used to identify 40 31.2 Endogenous targe
Slide 21: Dose-dependent CFTR Transgene RNA Expression: Cohort 4 Meets Target Profile
Dose-dependent CFTR Transgene RNA Expression Following 4D-710 Administration: Cohort 4 (2.5E14 vg) Meets Target Expression Profile CFTR RNA (ISH): Airway Epithelial Cells 80 2.5E14 vg Meets Target of ▪ Dose-dependent CFTRR mRNA Mean % (+) epithelial cells 52.7 ~10-25% 60 expression in airway cells
Slide 22: CFTR Transgene Protein Expression Pattern in Airway Biopsies (IHC): Controls
CFTR Transgene Protein Expression Pattern in Airway Biopsies (IHC): Non-CF Lung & Non-treated CF Biopsy Controls Non-CF Lung Control ▪ Commercial non-CF tissues are used as benchmark for target tissue expression quantity & level1, 2 ▪ CFTR overexpression may lead to protein mis-sorting to the basola
Slide 23: CFTR Protein Expression (IHC): Over-Expression Pattern at Highest Dose
CFTR Transgene Protein Expression Pattern in Airway Biopsies (IHC): Over-Expression Pattern at Highest Dose (Cohort 2) 2E15 vg Cohort 4 (2.5E14 vg) Participant 2 Participant 3 Participant 3 Participant 4 Non-CF Lung Control ▪ CFTR protein IHC staining pattern improved at 2.5E14 dose: o Abnormal cell
Slide 24: CFTR Protein Expression Meets Target Pattern at 2.5E14 vg Dose
CFTR Protein Expression Meets Target Expression Pattern in Airway & Interstitial Areas at 2.5E14 vg Dose (Cohort 4) Cohort 2 (2E15 vg) 2.5E14 vg Participant 2 Participant 3 Participant 3 Participant 4 Non-CF Lung Control ▪ CFTR protein IHC staining pattern improved at 2.5E14 dose: o Normal cellular
Slide 25: Dose-Related CFTR IHC Staining Patterns
Dose-Related CFTR IHC Staining Patterns: 2.5E14 vg Dose Meets Target Protein Expression Pattern 2E15 vg 2.5E14 vg (Cohort 4) Participant 2 Participant 3 Participant 3 Participant 4 Non-CF Lung Control Selected Phase 2 Dose © 2025 4D Molecular Therapeutics. All Rights Reserved. 25
Slide 26: Target Expression Profile Met at 2.5E14 vg Dose: Phase 2 Selection
Target Expression Profile Met at 2.5E14 vg Dose: Phase 2 Selection Dose Selection 2E15 vg 1E15 vg 5E14 vg 2.5E14 vg Target Profile Criteria: (n=4) (n=3) (n=3) (n=6) CFTR∆R RNA expression (ISH) 10-25% cells1,2 53% 39% 31% 17% CFTR protein expression (IHC) Positive Upper Airway Basal cells
Slide 27: 4D-710 Interim Phase 1 Data: Clinical Activity
4D-710 Interim Phase 1 Data: Clinical Activity Jennifer L.Taylor-Cousar, M.D., MSCS National Jewish Health Felix Ratjen, M.D., Ph.D, FRCP(C), FERS University of Toronto, Program Head, SickKids Research Institute, and Co- Head of the Cystic Fibrosis Center at SickKids Data Cutoff: December 1, 2025 ©
Slide 28: Integrated Biomarker & Clinical Endpoint Strategy
Integrated Biomarker & Clinical Endpoint Strategy to Demonstrate Mechanism of Action & Clinical Activity in AEROW Phase 1 Trial Lung Tissue Biomarkers Pulmonary Clinical Activity Endpoints Biopsies Larger Airways Pulmonary Symptoms ▪ ppFEV1 ▪ CFQ-R-R Brushings Structural/Functional Changes Small Air
Slide 29: ppFEV1 & LCI2.5: Complementary Measures of Lung Function
ppFEV1 & LCI2.5: Complementary Measures of Lung Function Measure / Endpoint Spirometry: ppFEV1 Lung restriction/obstruction, Measures marker of larger airway disease Effort Dependent Yes Sensitivity to Early Disease Low Medium Responsiveness to Intervention may miss subtle improvement in early/mild
Slide 30: ppFEV1 & LCI2.5: Complementary Measures of Lung Function
ppFEV1 & LCI2.5: Complementary Measures of Lung Function Measure / Endpoint Spirometry: ppFEV1 Multiple Breath Washout: LCI2.5 Lung restriction/obstruction, Ventilation inhomogeneity, Measures marker of larger airway disease marker of small airway disease Effort Dependent Yes No Sensitivity to Early
Slide 31: LCI2.5 Score Severity Correlates Significantly with Survival (N=237)
LCI2.5 Score Severity Correlates Significantly with Survival (N=237) Kurz JM et al., Eur Respir J 2022 © 2025 4D Molecular Therapeutics. All Rights Reserved. 31
Slide 32: LCI2.5 Demonstrated Greater Sensitivity than ppFEV1 for Treatment Effect
LCI2.5 Demonstrated Significantly Greater Sensitivity than ppFEV1 for Treatment Effect: Required <19 Subjects vs ~350 Inactive Control HS vs. IS Required Sample Outcome Analysis Treatment Effect* Size† Spirometry: ppFEV1 1.8 ± 12.0 351 Treatment ppFEF25-75 5.3 ± 22.3 141 effect size: 1.16 ± 0.94 CFQ
Slide 33: In Orkambi Trial in Children, LCI2.5 Demonstrated Robust Treatment Response
In Orkambi® Trial in Children, LCI2.5 Demonstrated Robust Treatment Response at Every Timepoint vs. Placebo in Contrast to ppFEV1 Primary Endpoint: LCI2.5 Treatment Placebo LUM/IVA Difference Endpoint (n=101) (n=103) vs Placebo Absolute change in 0.08 -1.01 -1.09 LCI2.5 through (-0.18 to 0.34) (-1.2
Slide 34: 4D-710 Interim Phase 1 Clinical Activity Data: Lower Dose Cohorts
4D-710 Interim Phase 1 Clinical Activity Data: Lower Dose Cohorts Jennifer L.Taylor-Cousar, M.D., MSCS National Jewish Health Data Cutoff: December 1, 2025 © 2025 4D Molecular © 2025 4DTherapeutics. Molecular Therapeutics. All Rights Reserved. All Rights Reserved.
Slide 35: Pulmonary Clinical Activity: Mean Change in ppFEV1, LCI2.5 & CFQ-R-R
Pulmonary Clinical Activity: Mean Change in ppFEV1, LCI2.5 & CFQ-R-R in Lower Dose Cohorts From Baseline Through 1 Year of Follow-up (Months 3 to 12*) ppFEV1 LCI2.5 CFQ-R-R Minimal Clinically Important Difference: +4 points Phase 2 Dose Phase 2 Dose Phase 2 Dose *Post-Hoc analysis of evaluable data:
Slide 36: Participant-level Clinical Activity Data Following 5E14 vg Dosing (Cohort 3)
Participant-level Clinical Activity Data Following 5E14 vg Dosing (Cohort 3) Jennifer L.Taylor-Cousar, M.D., MSCS National Jewish Health Data Cutoff: December 1, 2025 © 2025 4D Molecular © 2025 4DTherapeutics. Molecular Therapeutics. All Rights Reserved. All Rights Reserved.
Slide 37: 5E14 vg Participant 1: 42 y.o. Female with Normal Baseline Lung Function
5E14 vg Participant 1: 42 y.o. Female with Normal Baseline Lung Function by ppFEV1; LCI2.5 Not Available CFTR∆R Lung Function Pulmonary Symptoms Expression mRNA (ISH) Larger Airway: ppFEV1 Quality of Life: CFQ-R-R (MCID: +4) Protein (IHC) Small Airway: LCI2.5 Not available (enrolled prior to LCI ame
Slide 38: 5E14 vg Participant 2: 40 y.o. Female with Moderate Lung Disease
5E14 vg Participant 2: 40 y.o. Female with Moderate Lung Disease, Positive 4D-710 Response by Transgene Expression, LCI2.5, ppFEV1 & CFQ-R-R CFTR∆R Lung Function Pulmonary Symptoms Expression mRNA (ISH) Larger Airway: ppFEV1 Quality of Life: CFQ-R-R (MCID: +4) Protein (IHC) Small Airway: LCI2.5 Numb
Slide 39: 5E14 vg Participant 3: 34 y.o. Male with Moderate Lung Disease
5E14 vg Participant 3: 34 y.o. Male with Moderate Lung Disease, Positive 4D-710 Response by Transgene Expression, LCI2.5, ppFEV1 & CFQ-R-R CFTR∆R Lung Function Pulmonary Symptoms Expression mRNA (ISH) Larger Airway: ppFEV1 Quality of Life: CFQ-R-R (MCID: +4) Protein (IHC) Small Airway: LCI2.5 Number
Slide 40: Participant Clinical Activity Data Following 2.5E14 vg Dosing (Cohort 4)
Participant Clinical Activity Data Following 2.5E14 vg Dosing (Cohort 4, Phase 2 Dose) Jennifer L.Taylor-Cousar, M.D., MSCS National Jewish Health Data Cutoff: December 1, 2025 © 2025 4D Molecular © 2025 4DTherapeutics. Molecular Therapeutics. All Rights Reserved. All Rights Reserved.
Slide 41: 2.5E14 vg Participant 1: 26 y.o. Male with Moderately Severe Lung Disease
2.5E14 vg Participant 1: 26 y.o. Male with Moderately Severe Lung Disease Positive 4D-710 Response by Transgene Expression, LCI2.5, ppFEV1 & CFQ-R-R CFTR∆R Lung Function Pulmonary Symptoms Expression mRNA (ISH) Larger Airway: ppFEV1 Quality of Life: CFQ-R-R (MCID: +4) Protein (IHC) Small Airway: LCI
Slide 42: 2.5E14 vg Participant 2: 54 y.o. Female with Mild Lung Disease
2.5E14 vg Participant 2: 54 y.o. Female with Mild Lung Disease Positive 4D-710 Response by Transgene Expression, LCI2.5, ppFEV1 & CFQ-R-R CFTR∆R Lung Function Pulmonary Symptoms Expression mRNA (ISH) Larger Airway: ppFEV1 Quality of Life: CFQ-R-R (MCID: +4) Protein (IHC) Small Airway: LCI2.5 Numbers
Slide 43: 2.5E14 vg Participant 3: 37 y.o. Female with Moderately Severe Lung Disease
2.5E14 vg Participant 3: 37 y.o. Female with Moderately Severe Lung Disease, Positive 4D-710 Response by Transgene Expression CFTR∆R Lung Function Pulmonary Symptoms Expression mRNA (ISH) Larger Airway: ppFEV1 Quality of Life: CFQ-R-R (MCID: +4) Protein (IHC) Small Airway: LCI2.5 Not available (Site
Slide 44: 2.5E14 vg Participant 3 Addition of Modulator at 8 Months
2.5E14 vg Participant 3 Addition of Modulator at 8 Months: Positive 4D-710 + Modulator Safety (Clinical Activity Not Evaluable) CFTR∆R Lung Function Pulmonary Symptoms Expression mRNA (ISH) Larger Airway: ppFEV1 Quality of Life: CFQ-R-R (MCID: +4) Protein (IHC) Small Airway: LCI2.5 Not available (Si
Slide 45: Pulmonary Clinical Activity: Mean Change Through One Year of Follow-up
Pulmonary Clinical Activity: Mean Change in ppFEV1, LCI2.5 & CFQ-R-R in Lower Dose Cohorts From Baseline Through One Year of Follow-up (Months 3 to 12*) ppFEV1 LCI2.5 CFQ-R-R Phase 2 Dose Phase 2 Dose Phase 2 Dose *Post-Hoc analysis of evaluable data: Mean values for each participant calculated from
Slide 46: Dose Selection Framework for Further Development of 4D-710
Dose Selection Framework for Further Development of 4D-710 Focus of Phase 1: Focus of Phase 2 (Enrolling): Dose Finding Characterize Clinical Activity Safety Stability or improvement in large-/mid- airway disease (ppFEV1) Phase 2 Physiologically relevant Dose: Improvement in small airway disease C
Slide 47: Transgene Expression Durability Data & Next Steps
Transgene Expression Durability Data & Next Steps David Kirn, M.D. Co-founder and Chief Executive Officer Data Cutoff: December 1, 2025 © 2025 4D Molecular © 2025 4DTherapeutics. Molecular Therapeutics. All Rights Reserved. All Rights Reserved.
Slide 48: Durability of 4D-710-mediated CFTR Expression in Airway Biopsies
Durability of 4D-710–mediated CFTR Expression in Airway Biopsies: Persistent within Target Therapeutic Range through Over 1 Year Durability of 4D-710–Mediated CFTR∆R Expression (ISH): Mean % of Airway Epithelial Cells (+) in Individual Patients with Optional Paired Biopsies 40 Population & Methods 3
Slide 49: Advancing 4D-710 Together with CFF: 10 Year Collaboration
Advancing 4D-710 Together with CFF: 10 Year Collaboration & ~$31M in Financial Support From CFF Mechanism Financials Benefits ▪ JSC governance structure to coordinate strategic Research Grant $6.3M planning for 4D-710 development (2016) ▪ Senior development, biostats, and regulatory expertise ▪ Scie
Slide 50: 4D-710 Next Steps
4D-710 Next Steps Expected Milestones: ▪ H1 2026: Complete Phase 2 enrollment 4D-710 ▪ 2026: Advance redosing strategy Durable, Redosable, Variant- Agnostic, Disease-Modifying o Collect additional paired biopsy data in 2.5E14 vg dose Treatment Potential for People o Update redosing strategy based on
Slide 51: Thank You
THANK YOU 5858 Horton Street, Suite 455 | Emeryville, California 94608 (510) 505-2680 | Investor.Relations@4DMT.com IR.4DMT.com | LinkedIn © 2025 4D Molecular © 2025 4DTherapeutics. Molecular Therapeutics. All Rights Reserved. All Rights Reserved.
Slide 52: Protocol Amended with Novel Lung Endpoints to Enhance Clinical Activity Assessments
Protocol Amended with Novel Lung Endpoints to Enhance Clinical Activity Assessments, Additional Biopsy to Assess Durability Phase 2 Phase 1 (Dose Ranging) Key Endpoints: (Dose Expansion) Cohort 2 2E15 vg ▪ Safety & tolerability (n=4) Cohort 1 ▪ Lung biomarkers 1E15 vg (bronchoscopy): 2.5E14 vg (n=3)
Slide 53: 4D-710 Phase 1/2 Clinical Trial: Dose Exploration (Higher Doses, n=7)
4D-710 Phase 1/2 Clinical Trial: Dose Exploration Demographics and Baseline Characteristics (Higher Doses, n=7) Cohort 1: 1E15 vg Cohort 2: 2E15 vg Participant Number 1 2 3 1 2 3 4 Age, y 36 24 20 37 27 32 69 Sex Male Male Female Female Male Female Female CFTR modulator Intolerant Ineligible Ineligi
Slide 54: 4D-710 Phase 1/2 Clinical Trial: Dose Exploration (Lower Doses, n=9)
4D-710 Phase 1/2 Clinical Trial: Dose Exploration Demographics and Baseline Characteristics (Lower Doses, n=9) Cohort 3: 5E14 vg Cohort 4: 2.5E14 vg Participant 1 2 3 1 2 3 4 5 6 Number Age, y 42 39 34 25 54 36 56 32 37 Sex Female Female Male Male Female Female Male Male Male CFTR mod. status Intole
Slide 55: Biopsies & Brushings Collected in Multiple Lung Lobes Bilaterally
Biopsies & Brushings Collected in Multiple Lung Lobes Bilaterally For Evaluation of DNA, RNA & Protein Bronchoscopy: Week 4* Trachea Bronchoscopic Sampling Sites Right Left Endobronchial biopsy 1 Upper Lobe Apico-posterior 1 Right secondary carina Upper Lobe 3 Anterior Apical Superior-lingular Anter
Slide 56: 4DMT CFTR IHC Assay Development
4DMT CFTR IHC Assay Development Validated by Extensive Control Testing to Ensure Specificity to CFTR Epitope Test Control Cell/Tissue Result Specificity and Transfected vs. un-transfected HEK293T cells Confirmed Signal Differential Untreated HT29 vs. CFTR CRISPR-modified knockout HT29 cell lines Con
Slide 57: IHC & ISH Assay Specificity and Sensitivity
IHC & ISH Assay Specificity and Sensitivity Superior Sensitivity of IHC Compared to ISH Confirmed in 4D-710–Transduced HT29 CFTR CRISPR KO Cells 4D-710–Transduced HT29 CFTR KO Cells HT29 HT29 Parental CFTR KO MOI 10 MOI 200 MOI 4000 MOI 10,000 MOI 20,000 MOI 40,000 CFTRR RNA (ISH) CFTR Protein (IHC
Slide 58: CFTR Protein Expression: Machine Learning-Assisted Image Analysis
CFTR Protein Expression: Machine Learning-Assisted Image Analysis (Quantitative & Qualitative Analyses) ▪ Clinical-grade image analysis software* Non-CF CF Lung Study Sample ▪ Holistic & objective whole-slide analysis: o 100% of airway epithelial cells analyzed o 100% manual QC to confirm accuracy ▪
Slide 59: 4D-710-Mediated CFTR Protein Expression Confirmed, Above Controls
4D-710–Mediated CFTR: Protein Expression Confirmed, Above Non-CF and CF Lung Controls CFTR % (+) Epithelial Cells (IHC) CFTR Staining Intensity (IHC)* 99.9 97.3 98.9 300 224.9 218.4 100 90.6 191.9 250 Mean % (+) epithelial cells Mean intensity score* 80 200 44.2 139.8 60 150 18.4 40 100 46.7 20 18.8
Slide 60: Pre-existing A101 Immunity Did Not Affect CFTR RNA or Protein Expression
Pre-existing A101 Immunity Did Not Affect CFTR∆R RNA or CFTR Protein Expression * Pre-existing Anti-A101 Capsid Antibodies ▪ 5/16 positive for pre-existing A101 capsid antibodies ▪ No significant difference in mRNA/protein expression between participants with (n=5) and without (n=11) pre-existing A1
Slide 61: LCI Detects Early Airway Disease in Children Before FEV1 Decline
LCI Detects Early Airway Disease in Children Before Noticeable FEV1 Decline 20 Normal FEV1* Cystic fibrosis Healthy controls 18 16 Patients with CF who have normal FEV1 but elevated LCI 14 Elevated LCI LCI 12 10 8 6 4 -5 -4 -3 -2 -1 0 1 2 3 FEV1 z-score • 50% of children (age 6 to 16 years) with CF
Slide 62: LCI2.5 Accepted by Regulatory Agency (Vertex Products)
LCI2.5 Accepted by Regulatory Agency (Vertex Products) In Pivotal Trial to Support Drug Approval Product Agency Primary/Secondary Endpoint Population Approval Primary in 1 trial CF 6–11 y Label extension to 6–11 y (efficacy and safety) EMA Secondary in 3 trials CF 2–5 y Label extension to 2–5 y (saf
Slide 63: Redosing Considerations: Data Support Feasibility of Repeat Dosing
Redosing Considerations: Biological Context and Data Support Feasibility of Repeat Dosing for Aerosolized 4D-710 Rationale 4DMT Results ▪ Lung epithelium expected, repeat dosing expected to ▪ NHP pre-existing anti-A101 immunity (cross- be required reactivity): Safe & equivalent transduction vs immun
Slide 64: First-time Durability in CF Lung with 4D-710-mediated CFTR Expression
First-time Durability in CF Lung with 4D-710–mediated CFTR Expression Persistent within Target Therapeutic Range through 1 to 3 Years Durability of 4D-710–Mediated CFTR∆R Expression (ISH): Mean % of Airway Epithelial Cells (+) in Individual Patients with Optional Paired Biopsies 50 Population & Meth
Slide 65: Strong Rationale for 4D-710 + Trikafta Combination
Strong Rationale for 4D-710 + Trikafta Combination Rationale In vitro: 4D-710+Trikafta = CFTR Function Improvement ▪ Unmet need remains F508del/G542X F508del/G542X o Trikafta Ph 3 results suggest population with 50 15 NT suboptimal response, remaining unmet need NT Trikafta Trikafta 4D-710 4D-710 4D