FERTILITY PRESERVATION & POST TREATMENT CONCERNS IN HAEMATOLOGY PATIENTS Joanne McManus Consultant Gynaecologist Regional Fertility Centre Belfast Trust ----- Meeting Notes (12/10/17 19:42) ----- As my specialist areas in gynaecology are fertility & menopause I see a siginficant number of young women & men who have reproductive health problems as a result of haematological & other malignancies. It is therefore encouraging that we can now offer hope not only for men but also for girls by being able to store eggs as well as sperm. While this is described using the umbrella term of fertility preservation it is really only offering them one or maybe 2 chances to conceive using hteir own gametes. Nevertheless it offers some hope & optimism for a future that otherwise may look very bleak It is

FERTILITY SPECIALISTS QUESTIONS HAEMATOLOGISTS WHO should be referred? WHAT can be done? What procedures are involved? How long will it take Is fertility preservation worthwhile? FERTILITY SPECIALISTS WHAT is the disease? WHAT are the chances of treatment affecting fertility? What are the chances of survival ? How much time is there? Is is Safe to do procedures? The initial title for this talk that was suggested by Laura was ‘fertility Myth busting’ which suggests that you as haematology nurses & doctors know that you should be referring patients for fertility preservation but probably cant answer the patients questions as to what it involves or if they are suitable candidates for it. But I can reassure you that you proabably know a lot more about fertility than we as gynaecologists know about haematological disorders. The first patient I ever saw for fertility preservation had myelodysplasia which I dint think I had even heard of.

What conditions affect patients of reproductive age? Hodgkins Lymphoma 64% <50years Acute Lymphoblastic Leukaemia 75% < 50years Acute Myeloid Leukaemia 20% <50years NHL 17% <50years At time of Stem cell transplant 40% allogenic survivors < 40y 25% autologous recipients <40y The main conditions that affect men & women of reproductive age are Hodgkins & the acute leukaemias While we have seen lots of men over the years for sperm storage with HL we are now getting young women coming along for egg storage pre chemo. Obviously we don’t see the acute leukaoemia patients as they are too unwell & don’t have time to go through egg harvesting but I see these women after transplants for treamtent of their POF HL – 80% cure rate, 40-90% for ALL HCT= haemopoietic stem cell transplant

EFFECT ON FERTILITY HIGH DOSE CHEMO OR RADIOTHERAPY – gonadal failure Lower doses – may not stop periods but will reduce ovarian reserve – accelerate menopause- cause a fertility deficit Risk of ovarian failure increases with age at treatment Can start off as as less gonadotoxic therapy but change if relapse MEN – sperm may recover –opposite for women Although lower dose

Effect on Female Fertility 1-2x 106 eggs at birth 25,000 age 37 1000 age 51 Chemotherapy destroys mature ovarian follicles- induces apoptosis in Granulosa cells Also harms dormant primordial follicles – ‘ovarian reserve’ via other mechanisms Radiation directly toxic to oocytes both in dormant primordial & larger antral follicles Dose 2Gy – 50 % reduction in ovarian reserve Chemo – damages dormant primordial follicles by poorly understood mechanisms ?apoptosis of primordial follicles themselves ? Cause fibrosis of stroma & reduced blood supply to primordial follicles Depletion of primordial follicles by premature activation triggered by a decrease in granulosa cells(which secrete factors that inhibit primordial cell recruitment Alison W. Loren Hematology 2015;2015:138-145

Fertility Preservation Options MALE Post puberty Sperm banking well established Pre–pubertal No options locally (research - Oxford) FEMALE IVF & embryo storage - if partner Egg harvesting & cryopreservation Pre-pubertal Ovarian tissue storage not available NI Can refer to Edinburgh or Oxford Although still regarded as experimental & research >100 babies have been born worldwide

Male Fertility Preservation Longstanding service in Regional Fertility Centre Non –invasive but patient must be relatively well One or more appointments – if time allows Sperm count often low in haematological malignancies Stored sperm usually only suitable for IVF/ICSI Most men don’t use their stored sperm Storage regulations - labour intensive for clinic

Guardian December 2015 Oxford – Dr Sheila Lane Work in mice has shown sperm production after transplantation of testicular tissue & in vitro maturation

Oocyte Cryopreservation (egg freezing/ vitrification) Funding for 12 cases per year in RFC (1st case 2013) Conditions requiring surgery/chemo-radio- therapy likely to affect fertility Unlikley to be beneficial >37-38 years old Need approx 20 eggs to achieve a live birth Possible for any post- pubertal girl – how young?? Published case of 13 year old pre-menarchal girl with Myleodysplasia – 18 mature eggs stored (Cornell) Fertility & Sterility 2012;98:1225-8 EGG FREEZING HAS REALLY ONLY BECOME SUCCESSFULA OVER THE LAST DECADE WITH THE DEVELOPMNT OF VITRIFICATION AS A TECHNIQUE FOR FREEZING - SIGNIFICANT ADVANCE IN FERTILITY PRESERVATION AS WOMEN & GIRLS WHO DON’T HAVE A PARTNER NOW HAVE A REALTIVELY EASY MEANS OF PRESERVING some FERTILITY OR AT LEAST HAVING A CHANCE OF HAVING THEIR OWN CHILD IN THE FUTURE

EGG FREEZING Time needed = 2-+weeks Ovarian stimulation - gonadotrophin injections usually 10-12 days GnRH Antagonist – prevents premature ovulation Ultrasound follicle tracking – ideally transvaginal Serum oestradiol measurements Transvaginal egg retrieval – GA in younger girls Egg freezing has become very

OVARIAN STIMULATION & EGG RETRIEVAL RISKS INFECTION BLEEDING NEEDLE INJURY – BOWEL, BLADDER, VASCULAR OVARIAN HYPERSTIMULATION SYNDROME POOR RESPONSE – NO EGGS OR LOW NUMBERS

Leah Age 15 Diagnosed with High Risk Myleodysplasia March 2013 Seen by adult & Paediatric Haematology Recommended treatment allogenic BMT - 13yr old brother Likely to induce ovarian failure – conditioning chemo & radiotherapy Mother enquired about fertility preservation

Leah Attended RFC with Mum 8th May 2013 Discussed the option of egg harvesting & freezing Leah understood that her treatment was likely to result in ovarian failure & infertility Processes involved in ovarian stimulation and egg collection explained Urgency of BMT discussed with Bristol who felt that there was time for egg freezing RISKS – BLEEDING, INFECTION

Oocyte Cryopreservation Monitoring by abdominal ultrasound & blood tests Straightforward trans-vaginal egg collection - GA Prophylactic ciprofloxacin 500mg twice daily & Fluconazole 100mg once daily x 5 days– commenced 48h before egg collection 21 eggs, 18 mature eggs vitrified Future chance of live birth ? 25year old, 6 eggs injected – 37% chance of a live birth Fertil Steril 2013;100,492-9 Tragically Leah died 7 months later

EGG FREEZING 5/6/13- 02/10/17 Date Age Eggs Diagnosis Alive / Deceased 05/06/13 15 18 Myleodysplasia D Jan ‘14 30/10/13 22 12 Adenoca. cervix D Oct ‘14 11/11/13 24 11 Hodgkins Alive 10/07/14 17 31 AML relapse (TBI) Alive (GvH) 01/09/14 16 Ewings + ALL D March ‘16 23/06/15 20 09/07/15 36 2 Hodgkins 4B 26/08/15 30 4 Metastatic SB ca. D May ‘17 12/10/15 23 8 Yolk sac tumour 09/11/15 19 6 2/11/16 21 9 Ovary teratoma 21/11/6 37 Breast cancer 22/05/17 25 Adenosquamous cx 19/07/17 10 Hodgkins 4a 20/07/2017 34 3 embryos 20 MDS (ALL) 31/08/2017 38 7 embryos Breast ca 02/10/2017 29 13 Hodgkins 2a 27 Glioma 16 patients have had egg freezing - A THIRD OF OUR EGG STORAGE PATIENTS HAVE HL 4 out of 16 have died = 25%

Hodgkins Lymphoma LD age 29, single, no children Hodgkins Lymphoma neck & Mediastinum Gd 2a ABVD – low risk to fertility If relapse -further treatment would result in ovarian failure Less eggs likely to be obtained for freezing after initial chemo Anti- Mullerian (AMH) Hormone = 12.3pmol/l (average ovarian reserve) Swerdlow et al J Natl Cancer Inst 2014;106

Patient LD Screening for Hepatitis B, C & HIV carried out in advance of appointment Started ovarian stimulation on the same day attended RFC 20th September – day 5 of cycle Egg collection 2/10/17 13 eggs vitrified Vitrification (rather than slow rate freezing) has improved the success of egg freezing

Y T - attends HRT Clinic Age 36, Premature Ovarian failure HL diagnosed 2008 Age 27, married, 1st child 15mths old Relapse during 2nd pregnancy (2010) Treatment started when 2nd child 5 weeks old No periods after treatment for relapse Married – could have been referred for IVF & embryo freezing

Jane AML age 16 4 courses chemo Relapse June 2014 – referred to RFC Ovarian reserve (AMH) excellent Offered egg harvesting & storage 31 eggs stored Myeloablative chemo & sibling SCT Acute leukaemia – so no time for egg storage 4TH patient we stored eggs for in RFC

Jane Ovarian failure - referred to HRT clinic Graft v Host Disease including vagina Hormone replacement therapy – combined contraceptive pill Future fertility Options? Use frozen eggs ? Periods on Qlaira ? Dose of radiation

TOTAL BODY IRRADIATION 12GY – increased risk of Miscarriage Pre-term labour Low birth weight babies Very low chance of natural conception IVF – cryopreserved eggs, donor eggs

EDINBURGH OVARIAN CRYOPRESERVATION MRC study – 1st patient 1993 PROVISIONAL CRITERIA ≤ 30 years old No existing children Reasonable chance of surviving 5 years >50% chance of having ovarian function destroyed by the therapy If age >15: no previous chemo/radiotherapy If age <15: eligible if previous ‘mild’ chemotherapy 90 PATIENTS SO FAR – TISSUE REPLACED IN 3 50% <16 YEARS YOUNGEST 1.3

RISK OF MALIGNANT CONTAMINATION Ovary from ALL patients tested with PCR transplanted into mice showed malignant transformation in ovarian tissue if positive with PCR but not if negative. Gold standard should be transplantation of ovarain tissue into host mice for 20 weeks first. No risk of transmission if cryopreservation carried out after complete remission J Assist Reprod Genet. 2013 Jan; 30(1): 11–24

POST TREATMENT FERTILITY CONCERNS Young adults referred from Late Effects clinic to Fertility Clinic or Adolescent Gynae clinic MALES Semen Analysis FEMALES Can have assessment of ovarian reserve AMH measurement & ultrasound scan of ovaries Reasssuring if good result If AMH low may cause unnecessary distress

Patient ZF Junior doctor age 26 ALL age 14 Concerned re irregular periods – no period for 5 months Worried that chemo had caused premature menopause Going to Australia Wanted to discuss possibility of egg freezing.

Patient ZF Pelvic ultrasound Ovaries looked polycystic – lots of antral follicles Dominant follicle – imminent ovulation Hormone levels were ‘ovulatory’ Anti Mullerian Hormone – 64.1pmol/l – in keeping with excellent ovarian reserve & PCOS

POST TREATMENT - FEMALES May have reduced ovarian reserve & shorter reproductive window IF EARLY menopause – refer to SPECIALIST HRT CLINIC Need Hormone Replacement - optimised G v H can affect vagina If ovarian failure & no eggs or embryos stored can have donor eggs – refer to FERTILITY CLINIC HRT CLINIC MATER HOSPITAL, REGIONAL FERTILITY CENTRE - RVH

POST TREATMENT -MALES Childhood Cancer Often have anxiety re fertility May have very low sperm count or no sperm Adolescents /adults should have had sperm stored If low sperm count will need IVF/ICSI If need IVF/ICSI for male – or female – problem ? Use stored or fresh sperm High rate of aneuploidy & DNA damage in sperm after chemotherapy – but no increase in congenital abnormalities Concern that ICSI maybe bypass natural selection

Post Chemotherapy MEN – 2 YEARS – but what if female partner is 38+? How long to wait before trying for a pregnancy? MEN – 2 YEARS – but what if female partner is 38+? could consider using stored sperm? WOMEN - 6 MONTHS ? longer because of risk of relapse But may not have time if reduced ovarian reserve IVF SUCCESS AFTER CHEMO No evidence of reduced success – but women may have reduced ovarian reserve & hence less eggs Increased abnormalities – aneuploidy & DNA damage has been found in sperm of men post chemotherapy – many years later. But no increase in congenital abnormalities in babies – probably as ‘normal’ sperm naturally selected. Concern re ICSI bypassing natural selection hence some suggest using stored sperm

The Pain of Infertility Letter from patient’s mother Conor ALL 1979, age 3y 8mths. Relapse 1982- chemo finished 1984

Ist ICSI June 2007 SELF - FUNDED 11 eggs very few sperm 10 injected only 2 fertilised 2 good quality embryos - but not pregnant Sperm storage suggested for next treatment 3 x SAs Sept & Oct ‘07 – NO SPERM 2 samples stored Oct /Nov (10-30 sperm only)

2nd ICSI February 2008 NHS FUNDED 20 eggs, 15 injected, 3 fertilised 2 reasonable quality embryos transferred Clare admitted with Ovarian hyperstimulation Pregnant - baby boy 8/11/2008, 2.8kg @ 40 weeks gestation

ETHICAL ISSUES EQUALITY What is available ? where? to whom? FUNDING ASSISTED CONCEPTION WHO MAKES DECISIONS FOR CHILDREN/ADOLESCENTS ? Should funding for assisted conception for cancer survivors be separated from what might be considered less medical causes of infertility. Currently in NI there is the same allocation of one stimulated cycle & another embryo transfer if there are frozen embryos – for all couples. This is the same for a couple who have had 2 children naturally and fancy a 3rd but it isnt happening as it is for a young couple who need IVF because they have to use stored sperm or the man has a very low sperm count after chemo therapy

SUMMARY Low risk of ovarian failure with initial chemo for HL Treatment for relapse likley to induce ovarian failure & infertility ‘Egg Freezing’ should be considered for women of reproductive age (<38) especially if no children Lower egg yield even after ‘mild’ chemo – but still worth considering especially if young Male sperm storage easy & should be routine Women with ovarian failure need specialist HRT advice & can have fertility treatment with egg donation

Cytotoxic agents according to degree of toxicity High risk Intermediate risk Low risk Cyclophosphamide Doxorubicin Methotrexate Busulphan Cisplatin Bleomycin Melphalan Carboplatin 5-Fluorouracil Dacarbazine Actinomycin-D Procarbazine Mercaptopurine Ifosfamide Vincristine Thiotepa The ovaries are every sensitive to cytotoxic treatment, especially to alkylating agent eg cycle / ifos / busulphan / melphalan which are classified as high risk for gonadal dysfunction. Cyclophospahmide is the agent most commonly implicated in causing damage to oocytes and granulosa cells in a dose-dependent manner [meirow 1999]. Follicular destruction induced by alkylating agents generally results in loss of both endocrine and reproductive function depending on th edose and age of the patient. Indeed Larsen et al [JCEM 2003] reported a fourfold increased risk of POF in teenagers treated for cancer, and a risk increased by a factor of 27 in women aged 21-25yrs. This is a dose and dependent phenomena. For example, complete amenorrhoea (absence of periods) was reported after a dose of 5g of cyclo in women over 40yrs, and after doses of 9 and 20g in women 30-40 and 20-30 respectively [Shalet 1980]. A combination of chemotherapy agents increases risk. MOPP/ABVD – amenorrhoea developed in 89% and 20% over and under 25yrs at the age of treatment. Procarbazine has recently been described as high risk for inducing premature menopause [de bruin 2008]. 10yrs after treatment – 84% after high cumulative doses, compared to 15% after low dose (<=4.2g/m2)

Anti-Mullerian Hormone (AMH) AMH is a product of the granulosa cells in the pre antral & small antral follicles. It regulates folliculogenesis by inhibiting recruitment of resting follicles from the resting pool in order to select the dominant follicle

OVARIAN TISSUE TRANSPLANTATION >100 births worldwide several case series Van en Ver et al 2016 – 95 transplants in 74 women Average age 30y at cryopreservation, 34y at transplant Most common diagnosis – Breast ca & Hodgkins 1 year post transplant 62.5% - ovarian activity pregnancy rate = 27.5%, delivery 22.5% Mostly natural conception, some IVF CONCERN RE RISK OF MALIGNANT CONTAMINATION – OVARIAN TISSUE CAN BE BIOPSIED PRE STORAGE BUT CANT BIOPSY TISSUE ACTUALLY GOING BACK. CONCERN particularly in leukaemias Solid tumours – assumption that ovary should be free if no metastases

Age at ovarian cryopreservation: Edinburgh experience since 1993 Median age 18.4yr 50% aged 16 or less 90 patients so far, ovarian tissue replaced in 3

Ovarian or Oocyte preservation ? OR BOTH? EGG FREEZING OVARIAN FREEZING 2-4 week window required Puberty Lots of babies born Only one or 2 chances of conception – COST! Invasive procedure Limited centres Risk of disease transmission Success rates unknown ? Cure for menopause natural conception may avoid cost of IVF