Bone marrow transplant and gene therapy in cerebral ALD

Bone marrow transplant and gene therapy in cerebral ALD Dr Robert Chiesa Consultant in Bone Marrow Transplantation Great Ormond Street Hospital London 10:15 Dr Robert Chiesa- BMT/ Gene Therapy 1

Bone marrow transplant why the bone marrow (BM)? BM contains different cells including STEM CELLS and immune cells (T-cells); Stem cells have the ability to transform into different blood cells / cells forming the immune system / enzyme replacement; STEM CELL Stem cells from healthy donors can be collected (harvested) and transplanted (BMT) in patients with defective haematology/immunology/metabolics Immune cells prevent rejection & infections (viral) Stem cells can transform into different blood cells, and can be taken from bone marrow, peripheral blood or umbilical cord blood. Bone marrow transplants are used to treat blood conditions such as leukaemia. Over 2000 bone marrow transplants took place between 1980-2013. 2

BMT sources of stem cells BONE MARROW: requires harvest in theatre/general anaesthesia for donor PERIPHERAL BLOOD: stimulation with G-CSF, collection of stem cells UMBILICAL CORD BLOOD: used since 1988. 500.000 stored worldwide. 3

Indications to BMT in children Leukaemia Immunodeficiency Metabolic Bone marrow failure Inflammatory Bowel Diseases Rheumatoid Dermatology (EB) Thalassaemia Sickle cell disease 4

BMT in metabolic disorders (1980-2013) Worldwide >2000 BMTs have been performed in IEM; 4 main diseases: MPS type I X-ALD MLD GLD MLD 5% 9% X-ALD 17% Other 13% MPS I Hurler syndrome 28% Osteopetrosis 18% Other MPS 10% Osteopetrosis CIBMTR database 2013 5

BMT in lysosomal storage diseases (LSD) Lack of lysosomal enzymes; Enzyme is secreted by donor-derived (normal) cells and taken up by deficient cells; Cross correction of host cells. 6

How does BMT correct some LSD? 1 Chemo to eliminate patient s own stem cells 4 CNS Penetration donor cells 2 BMT Stem cell infusion 3 ENGRAFTMENT Migration and distribution donor derived-cells into host tissues The process is a combination of chemotherapy (eliminating the patient s own stem cells), infusing donor stem cells and grafting these on to the patient. A soluble enzyme secreted by the donor cells corrects cells in the patient. 7

Rationale for BMT in ALD Mechanism of action in ALD not entirely clear; ALDP is not a soluble enzyme (membrane-bound protein); Accumulation VLCFA may destabilize/damage myelin structure trigger inflammation; Donor-derived cells migration in the brain (microglia); Decrease oxidative stress/damage in tissues reduce inflammation; Immuno-modulatory properties of BMT The mechanism of how bone marrow transplants work for ALD is not clear, as ALD is not related to a soluble enzyme. A bone marrow transplant reduces inflammation, stress and damage, but it is not clear how this works. 8

Bone marrow transplantation steps DONOR SELECTION CONDITIONING REGIMEN STEM CELL INFUSION MANAGEMENT POST-BMT COMPLICATION & FOLLOW-UP Steps of a bone marrow transplant: Donor selection: tissue match needed between patient and donor. Preparation/Conditioning Regimen: Chemotherapy drugs are given to children to prepare them for transplant over the course of the 7-9 days before infusion of stem cells. Stem cell infusion Long follow up to monitor and tackle the consequences of conditioning 9

DONOR SEARCH tissue typing Blood tests to verify the presence of a tissue-matched donor (in the family or unrelated); PATIENT DONOR 1 A fully matched family donor is found in 25-30% patients; The chance of finding a fully matched unrelated donor depends upon the recipient s ethnic background (70% Caucasians vs 10% non-caucasians); = 10/10 25% find a match within their family. The chances of finding a fully matched unknown donor depend on race: Caucasians have a 70% chance of finding a match, while other races and those of mixed race will have much lower chances. 10

DONOR SEARCH tissue typing Blood tests to verify the presence of a tissue-matched donor (in the family or unrelated); PATIENT DONOR 2 A fully matched family donor is found in 25-30% patients; The chance of finding a fully matched unrelated donor depends upon the recipient s ethnic background (70% Caucasians vs 10% non-caucasians); The use of mismatched donors increases transplant-related toxicity; = 09/10 The use of mismatched donors increases the risk of transplant related toxicity. 11

DONOR SEARCH tissue typing Blood tests to verify the presence of a tissue-matched donor (in the family or unrelated); PATIENT DONOR 3 A fully matched family donor is found in 25-30% patients; The chance of finding a fully matched unrelated donor depends upon the recipient s ethnic background (70% Caucasians vs 10% non-caucasians); The use of mismatched donors increases transplant-related toxicity; = 05/10 Parents can be donors, but they will only have a 50% match 12

PREPARATIVE REGIMEN Chemotherapy (rarely radiotherapy) to prepare the patient to receive the transplant; Given over 7-9 days prior to the transplant; Two main components of the preparative regimen: - myeloablation (create space in the bone marrow) - immunosuppression (prevent graft rejection) Preparation/Conditioning Regimen: Chemotherapy drugs are given to children to prepare them for transplant over the course of the 7-9 days before infusion of stem cells. Myeloablation is used to create space in the bone marrow for new cells Immunosuppressants prevent graft rejection 13

PREPARATIVE REGIMEN intensity hierarchy Drugs are used in different combinations to produce a range of conditioning from mild to strong Minimal Intensity Conditioning ( MIC ) Reduced Intensity Conditioning ( RIC ) Myeloablative Intensity Conditioning ( MAC ) Different combinations of drugs are needed for different conditions- bone marrow transplants for ALD require the strongest combination- MAC. 14

PREPARATIVE REGIMEN toxicity Depends on the intensity of conditioning (RIC vs MAC) Acute (I): nausea, vomit, lethargy Acute (II): inflammation/pain mouth & gut, transient hair loss, organ toxcity (liver, kidneys, lungs), low immunity and infections Effective supportive treatment provided Late : possible effects on fertility / other.. The toxicity of drugs can cause nausea, vomiting and lethargy, with longer term possible side effects including inflammation, pain in the mouth, hair loss, low immunity causing infections, organ toxicity and potential infertility. 15

GRAFT REJECTION When the patient s residual immune system does not recognize the graft and fights it / rejects it; The preparative regimen weakens the patients immune system, so that the patient is more tolerant ; The immunosuppressants (like cyclosporine) have to continue for a few months after BMT In a fully matched graft, there is a 5% chance that the immune system will fight and reject the graft. This chance is higher in a mismatched graft. Preparation and immunosuppressants are used to prevent graft rejection 16

GRAFT VERSUS HOST DISEASE (GvHD) When the immune cells (lymphocytes) infused with the bone marrow do not recognize the patient and fight; Acute GvHD: commonly skin rash. Less commonly diarrhea or liver/lung disease; Treatment: more immunosuppression (drugs): resolution in 80% cases. There is also a risk of Graft Vs Host Disease (GVHD), which is the opposite of graft rejection. In GVHD, the graft does not recognise the patient and fights back against host cells. Immunosuppressants resolve this in 80% of cases. 17

Bone marrow transplantation Preparative regimen 7-9 days 18

Bone marrow transplantation BMT Preparative regimen 7-9 days 19

Bone marrow transplantation Preparative regimen BMT Acute toxicity Bacterial/ fungal infections Low counts 7-9 days 10-21 days 2-3 weeks after transplant, children experience acute toxicity of the chemotherapy, including immunodeficiencies leaving them vulnerable to bacterial, fungal and viral infections 20

FIRST RESULT AFTER BMT ENGRAFTMENT / CHIMERISM SCENARIO N.1 SCENARIO N.2 SCENARIO N.3 100% donor engraftment mixed chimerism rejection PATIENT CELLS FULL ENGRAFTMENT DONOR CELLS After 2-3 weeks, white blood cells will have been produced by the new cells. Blood samples are sent to a laboratory to give the first indications as to the success of the procedure. Results will be one of 3 options: All white blood cells are from the donor, showing a full and successful engraftment Mixed chimerism: some white blood cells from the host have survived, alongside donor cells. Many diseases can still be cured even when some patient white blood cells have survived Rejection: no white blood cells are from the donor. There is a 5% chance of this happening. 21

Bone marrow transplantation Acute GvHD chronic GvHD Preparative regimen BMT Acute toxicity Bacterial/ fungal infections Low counts Viral reactivations Immuno suppression Count recovery Gradual weaning immunosuppressants Gradual immune reconstitution 7-9 days 10-21 days 21-100 days 3-12 months 11 months after the transplant, children can still have weak immune systems and be at risk of viral infection. The immune system should begin to strengthen after around 5 months. After 12 months, the child s care is handed over to the metabolic team. 22

BMT in X-ALD History 1990: first successful BMT in 8y old boy with cald: normal VLCFA levels and stabilization disease; 1997: 3 Swedish boys after BMT poor outcome for 2 symptomatic patients, suggesting early intervention; 1998: first successful use of cord blood transplant advantage as rapid stem cell source; 2004: Report on 126 boys with cald post-bmt(peters et al) main cause of death was progression of ALD. Higher risk if symptomatic pre-bmt. The first successful bone marrow transplant for ALD took place on an 8 year old in 1990. Bone marrow transplants on 3 Swedish boys in 1997 were successful in just 1/3 cases, as the condition of the boys was too advanced. This highlighted the need for early intervention for bone marrow transplants for ALD. From 1998, stem cells from umbilical cord blood have been used for bone marrow transplants, as these are a rapid stem cell source In 2004, an international study was conducted on 126 boys with ALD diagnoses. This again showed a higher risk in transplants occurring after symptoms arose. Progression of ALD was the main cause of death for those for whom the transplant was not successful 23

Measuring Neurologic and Radiographic Progression in CALD Neurologic Function Score (NFS) 1 Component Hearing/auditory processing problems Score Aphasia/apraxia 1 Loss of communication 3 Vision impairment 1 Cortical blindness 2 Swallowing dysfunctions 2 Tube feeding 2 Running difficulties 1 Walking difficulties/ spasticity 1 Spastic gait (need assistance) 2 Wheelchair dependence 2 No voluntary movement 3 Episodes of incontinence 1 Total incontinence 2 Non-febrile seizures 1 Possible Total 25 1. Moser et al. Neuropediatrics 2000;31:227-9. 2. Loes et al. AJNR Am J Neuroradiol 1994;15:1761-6 1 Loes MRI severity score 2 : Measurement of white matter changes by degree and extent of pathological hyperintense regions (0-34) Loes score = 1 Loes score = 15 Gadolinium enhancement: Indicator of active inflammation A scoring system called NFS was set up to establish children s neurological conditions for prognostics prior to BMT. This is combined with a clinical Loes score, established from MRI scans, to determine probability of success of bone marrow transplant. Timing of the bone marrow transplant is therefore critical. GdE + 24

BMT in X-ALD Results 60 consecutive boys with X-ALD in single Institution; Analysis risk factors; 70% MAC, 30% RIC protocol; 50% UCBT, 50% BM; 70% early stage, 30% advanced disease. 25

BMT in X-ALD Results Survival depends on MRI changes and neurology at BMT; Timing BMT crucial; No difference EFS according to stem cell source. 26

BMT in X-ALD Conclusions Comparison of transplanted patients with historical untreated patients show striking survival advantage for BMT patients (Mahmood et al) ; Best neurological outcome in patients with NFS=0 and Loes <10 at time of BMT; Limited role of BMT in advanced disease;? Role BMT in prevention of AMN (3/5 adults with AMN post BMT, Geel et al); The role of transplants in the prevention of AMN is not well studied; transplants may not prevent AMN. 27

Who, when & how to transplant Early diagnosis ALD (newborn screening); MRI monitoring for detection cald; Urgent BMT if gad +ve changes or expanding white matter changes on sequential scans, before neurological changes; Choice of best available donor (HLA matched); Busulfan-based conditioning regimen Early diagnosis is needed, through New Born Screening and monitoring of MRI scans. Transplants become urgent when changes start to appear on scans, before neurological changes take place. These should use the best available donor, fully matched if possible. 28

Limitations of BMT Risk Treatment-related mortality % of patients 10% 1-4 Graft failure ~5% 1,4 Acute grade II-IV GVHD 10-40% 1-4 Chronic GVHD 10-20% 3,4 Full HLA donor match is required for best outcomes 1 1. Miller et al, Blood 2011; 2. Peters et al, Blood 2004; 3, Beam et al, Biol Blood Marrow Transplant. 2007; 4. Mitchell et al, Pediatr Transplant. 2012; 5. Martin et al, Biol Blood Marrow Transplant. 2006 Bone Marrow Transplants have limitations, as there are risks involved: Risk of death by infection ~10% Risk of graft failure ~5% Risk of Graft Vs Host Disease ~30% 29

Gene therapy DNA is introduced into a patient to treat a genetic disease; The new DNA usually contains a healthy gene to correct the effects of a disease-causing mutation; First clinical trials in 80 s; Extensive experience in primary immunodeficiencies; GT in other mono-genetic disorders (metabolic diseases) Gene therapy is an alternative approach, used to correct genetic diseases by treating the patient s own DNA. It can be used instead of a bone marrow transplant if there is no donor match available. The first trials of gene therapy took place in the 1980s, and this is now expanding to treat many metabolic diseases. 30

Starbeam Study: Open-label, Single-arm Phase 2/3 Study of Lenti-D in cald Key enrollment criteria: Age 17 years, evidence of active CALD (GdE+) with early disease (Loes score 0.5-9.0, NFS 1), no matched sibling donor for allo-hsct Primary efficacy outcome: Proportion of patients who are alive and MFD-free at 24 months Primary safety outcome: Proportion of patients who experience Grade 2+ acute GVHD or chronic GVHD at 24 months post-treatment Secondary and exploratory efficacy outcomes: Changes in NFS, GdE+ resolution, overall survival, and change in Loes score Additional key safety parameters: Engraftment failure, adverse events, detection of replication-competent lentivirus, insertional oncogenesis Eichler et al. NEJM 2017;377:1630-38. 31 The STARBEAM Study is a gene therapy trial, monitoring patients for 2 years after therapy to see changes in MRI, toxicity and so on. The study used a carefully selected group who showed limited signs of the disease when the process began. 31

Starbeam Study: Treatment Protocol Lenti-D Drug Product consists of an autologous CD34+ cell-enriched population that contains cells transduced with lentiviral vector that encodes an ABCD1 cdna for human 32 ALDP 32

Patient Enrollment and Baseline Characteristics Baseline Characteristics (N=21) Parameter Median (range) 21 subjects treated as of August 25, 2017 Median follow-up 30.2 months (1-46 months) 4 patients with 4 months of follow-up Age at consent (years) 6.0 (4-13) Time from CALD diagnosis to treatment (months) Time from consent to treatment (days) 4.7 (2.4-17.2) 66 (58.0-89.0) Baseline Loes score 2.0 (1.0-7.5) Baseline NFS score 0 (0-0) MFDs present at baseline None 33 33

Primary Endpoint Assessments Primary Endpoints Patient 2018 Patient 2016 Remaining Cohort Proportion of patients who are alive and MFD-free at 24 months Experienced rapid disease progression early on-study eventually resulting in MFDs and death Withdrew from study due to radiologic progression as evidenced by MRI Went on to receive an allo- HSCT and died due to transplant complications 15/17 (88%) remain MFD-free at 24 months 1 Historical data suggest ~75% of comparable patients receiving allo-hsct remain MFD-free at 2 years 2-5 % of patients who experience Grade 2+ agvhd or cgvhd at 24 months posttreatment All patients were free of GVHD 1. Eichler et al. NEJM 2017;377:1630-38. 2. Raymond et al. ALD-101 (in preparation); 3. Peters et al. Blood 2004;104:881-8. 4. Miller et al. Blood 2011;118:1971-8 2011. 5. Beam et 34 al. Biol Blood Marrow Transplant. 2007;13:665-74. 6. Baumann et al. Eur J Pediatr 2003;162:6-14. Of 17 patients involved, 15 had survived 2 years after the beginning of the process, with 1 of the 2 deaths occurring in surgery complications. The remaining 15 show no signs of major disability. Longer follow up is needed on this. 34

Secondary Efficacy Endpoint: NFS NFS was stable in 18 patients with evaluable data 1 Stable NFS: score of 4 without an increase in >3 points since baseline NFS consistent with those seen after allo-hsct in patients with similar baseline disease characteristics 2 1. Eichler F. Child Neurology Society, 2017. 2. Raymond et al. ALD-101 manuscript (in preparation) 35 35

Exploratory Efficacy Endpoint: Loes Scores Loes scores have stabilised in 15/20 patients at last visit 1* Stable Loes score: change of 5 points or an absolute Loes score 9 Loes scores consistent with those seen after allo-hsct in patients with similar baseline disease characteristics 2 1. Eichler F. Child Neurology Society, 2017. 2. Raymond et al. ALD-101 manuscript (in preparation). *Excludes patient 2016 who had a stable Loes score at last visit 36 36

Subject ID Secondary Endpoint: gadolinium Enhancement GdE- GdE+ By Month 24, 11 of 15 patients were negative for gadolinium enhancement 1 Early results suggest no correlation between post-treatment gadolinium enhancement status and clinical outcomes 2 In all patients who had reemergence of gadolinium enhancement, the enhancement was less extensive than that seen at baseline 1. Eichler F. Child Neurology Society, 2017. 2. Eichler et al. N Engl J Med 2017;377:1630-1638. 37 37

Safety Data Profile consistent with myeloablative conditioning Parameter Drug Product Cell Dose (x10 6 /kg) Neutrophil engraftment 1 (n=21; with and without GCSF) Median (min, max) 10.5 (6.0, 19.4) Day +13 (11, 39) Platelet engraftment 2 (n=18) Day +28.5 (16, 55) 1. 3 consecutive absolute neutrophil count values of 0.5 x 10 9 /L obtained on different days. 2. 3 consecutive platelet values 20 x 10 9 /L obtained on different days (unsupported) No graft failure or GVHD No insertional oncogenesis No replication competent lentivirus detected to date AEs consistent with myeloablative conditioning Grade 3 non-lab AEs in >1 subject C < NE (N=21) NE to M24 (N=21) Febrile neutropenia 17 (81%) 0 Stomatitis 6 (29%) 0 Nausea 4 (19%) 0 Vomiting 2 (10%) 0 Decreased appetite 9 (43%) 0 Hypokalaemia 4 (19%) 0 Epistaxis 2 (10%) 0 C = date of the start of conditioning; < NE = the day before neutrophil engraftment; NE = neutrophil engraftment Three AEs considered related or possibly related to drug product: BKmediated viral cystitis (SAE, grade 3), tachycardia (grade 1), vomiting (grade 1) These AEs and SAEs resolved with standard measures Data as of August 25, 2017 38 38

Gene therapy in cald summary Of the 17 patients who completed 24 months of follow-up, 15 (88%) remain alive and free of major functional disabilities; No graft failure or GvHD reported to date Adverse events profile consistent with myeloablative conditioning with busulfan and cyclophosphamide Additional follow-up is ongoing to assess durability of effect and long-term safety 39 39

?THERAPEUTIC OPTIONS Sir Luke Fildes The doctor 40

THERAPEUTIC OPTIONS CONVENTIONAL BMT GENE THERAPY Established treatment Long term follow-up Excellent results with fully matched donor Reduced TRM No GVHD Good preliminary neurological outcome Mortality 10% Risk of rejection/gvhd Use of mismatched donor Limited follow-up? Role gad +ve MRI post-gt 41

CONCLUSIONS BMT is the standard of care for c-ald, with good outcomes when performed early in pre-symptomatic patients; Importance NBS/early diagnosis; Role of BMT in more advanced disease is uncertain; Unclear if BMT can prevent AMN (possibly not); Risk of TRM and GVHD, especially when mismatched donors are used; Gene therapy is a promising alternative to BMT when matched donor is not found GT: good preliminary neurological & safety data. No risk of GVHD; GT: needs long term follow up; Importance of counselling families regarding available strategies. Summary: There are 2 options available for those with ALD, as long as early diagnosis has occurred: bone marrow transplant and gene therapy. Bone marrow therapy is a more established approach, with a longer follow-up history, but this requires a matched donor: mortality is ~10% higher for transplants with mismatched donors. Longer follow up is needed on the effects of gene therapy on ALD patients. Counselling for families is essential in both cases. It is possible that bone marrow therapy cannot stop AMN, as there is limited evidence of this 42