Early Human Development 86 (2010) 619625

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Early Human Development

j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / e a r l h u m d ev

Retinoblastoma Current treatment and future direction

Manoj V. Parulekar
Birmingham Children's Hospital, Steelhouse Lane, Birmingham B4 6NH, United Kingdom

a r t i c l e
Keywords: Retinoblastoma Infant Tumour Ocular Mutation Heritable Chemotherapy Screening Intra-arterial Enucleation Cancer

i n f o

a b s t r a c t
Retinoblastoma is the commonest primary ocular malignancy of childhood. There are two forms heritable and non heritable. Heritable retinoblastoma is a cancer susceptibility syndrome. Presentation is in the rst few years of life, sometimes in the neonatal period. Early detection and prompt treatment can give cure rates up to 95% for intraocular tumours, but extraocular disease carries a very high mortality. The diagnosis is essentially clinical and biopsy is contraindicated due to the risk of extraocular spread. Treatment requires signicant multidisciplinary input, with local ophthalmic treatment, systemic chemotherapy and external beam or plaque radiotherapy, or surgery to remove the affected eye. Screening of family members is essential for early detection. Lifelong surveillance of mutation carriers is needed due to the risk of second cancers. Newer treatment modalities including intra-arterial chemotherapy have been added to the therapeutic armamentarium in recent years. 2010 Elsevier Ireland Ltd. All rights reserved.

1. Introduction Retinoblastoma is a malignant tumour arising from the developing retina. It typically presents in the rst 23 years of life, often during infancy. The tumour is conned to the eye in the early stages, and cure rates for intraocular retinoblastoma can be as high as 95%. Extraocular spread (Fig. 1) carries a very poor prognosis [1], with cure rates below 510%. Early diagnosis and prompt treatment is therefore crucial to save life and vision.

3. Genetics Understanding the genetics of retinoblastoma is important when planning management. The tumour arises from primitive cells of the developing retina with loss of function of the Rb tumour suppressor gene (Ch13q14). [6] The retinoblastoma gene was the rst oncogene to be cloned, and is the prototype genetic cancer syndrome [7]. Other genetic cancer syndromes include the LiFraumeni syndrome and Xeroderma pigmentosum. There are two copies (alleles) of the Rb gene in every cell in the body, and at least one functioning allele is required to prevent the development of retinoblastoma in a retinal cell. Loss of both alleles will result in retinoblastoma tumours. Interestingly, in 1973 Knudson, a statistician proposed that two separate mutational events M1 and M2 that result in loss or inactivation of both Rb gene copies are required to initiate the tumour on the basis of his epidemiological studies, and this came to be known as the Knudson two-hit hypothesis which has stood the test of time [8]. Recent evidence suggests that tumour initiation (retinoma) can occur with 2 hits, but subsequent mutations (M3Mn) are necessary for the tumour to grow into retinoblastoma [9]. Loss of segments of the chromosome carrying the Rb1 genetic material e.g. chromosome 13q14 deletion can also result in retinoblastoma, and are often associated with developmental delay and dysmorphic features. The two hits can occur in one of two situations. In the genetic (also referred to as germline or heritable) form, every cell in the body is missing one copy of the Rb gene, the mutation occurring at the zygote stage. Every retinal cell is missing one copy of the gene, and can potentially give rise to a tumour. The other situation is the somatic

2. Epidemiology Retinoblastoma is the commonest primary malignant intraocular tumour of childhood accounting for up to 1% of all tumours in infancy. The age adjusted annual incidence is 12 per million population [2] under the age of 4 years. The incidence of sporadic retinoblastoma is 1 in 15,00020,000 live births, with no gender or racial predilection. As expected, the burden of disease is highest in populous developing countries in proportion to the birth rate [3]. The median age at presentation is under 12 months in heritable cases, and closer to 24 months in sporadic cases [4]. Presentation after the age of 6 years is extremely rare, although there are isolated reports of cases presenting as late as 26 years [5].

Tel.: +44 121 333 9465; fax: +44 121 333 9461. E-mail addresses: manojparulekar@aol.com, manoj.parulekar@bch.nhs.uk. 0378-3782/$ see front matter 2010 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.earlhumdev.2010.08.022

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Fig. 1. Advanced retinoblastoma with fungating orbital mass.

(non heritable) type where a single developing retinal cell loses one copy of the Rb gene during retinal development the rest of the body cells are normal. This constitutes the rst hit. The second hit is a random event, and gives rise to a tumour. This makes it easier to understand why genetic cases often have multiple tumours in one or both eyes (bilateral or unilateral multifocal), while somatic cases are always unilateral and unifocal.

4. Inheritance patterns Retinoblastoma may be inherited or occur sporadically. Over 90% of cases are sporadic (with no family history). In most of these cases the mutation is somatic and gives rise to isolated unilateral disease. A third of the sporadic cases arise from new germline mutations which are heritable (can be passed to offspring but not inherited from the parent). 40% of all cases are bilateral (and necessarily germline), and 60% are unilateral (which could be somatic or germline). Of the unilateral cases, 15% are germline, and carry the same risks as bilateral germline cases. Germline mutations are highly penetrant. Over 90% of children carrying the Rb gene defect will develop retinoblastoma. It has been suggested that there is an increased incidence of retinoblastoma in babies born of assisted conception. There is however insufcient evidence in the literature to support or refute this theory [10,11].

(Fig. 3). Early presentation within the rst few months of life is usual for bilateral cases, and even at birth in some cases. Unilateral cases (most of which are somatic mutations) usually present later, often at the age of 2 to 3 years or older. However, approximately 15% of unilateral cases carry the germline mutation, and such cases can present early, often in infancy. The commonest presenting features in unilateral cases are leukocoria (60%) and squint (20%). Less common modes of presentation are reduced vision detected at school screening, acute red eye, orbital inammation and excess watering. If a child presents with unilateral retinoblastoma early in life, it is more likely to be germline (heritable) as described earlier, with a high risk of developing more tumours in the same or fellow eye. Careful screening of the fellow eye is essential in all such cases through childhood. Advanced cases are rarely seen in economically advanced countries. This is more common with unilateral cases. Anterior segment involvement may result in raised pressure in the eye (glaucoma), enlargement of the globe or change in iris color (heterochromia). Less common presenting features include phthisis bulbi (shrinkage of the globe), pseudohypopyon (cells in the anterior chamber appearing as a layered white material) rubeosis (dilatation of iris vessels) and hyphaema (bleeding into the anterior chamber). Some cases may present with periocular inammation. Late presentation with proptosis (forward displacement of the eye) or fungating orbital mass is not unusual in countries with limited access to healthcare and carries a very poor prognosis (Fig. 1). In advanced cases, the tumour can spread along the optic nerve to involve the central nervous system, or through the choroid resulting in haematogenous spread to bones, lungs and abdominal solid organs. Digital ash photography can highlight the white reex (Fig. 3) in some cases, and lead to earlier detection, and this continues to be the subject of campaigns to raise awareness. Only a minority of cases with a white reex on photographs will turn out to have retinoblastoma however. Squint, refractive errors and photographs taken from an angle (off axis) can similarly produce a white reex diminishing the specicity of this technique as a screening test [12]. However, the Honduran retinoblastoma campaign was associated with a reduction in the proportion presenting with advanced disease [13].

6. Diagnosis The diagnosis of retinoblastoma is essentially clinical, and may be supported by imaging in some cases. Diagnostic biopsy is contraindicated because of the risk of extraocular spread [14,15]. The typical ndings are one or more round white retinal mass(es) growing into the vitreous cavity (endophytic) or growing into the sub retinal space (exophytic). Fragments of the tumour can spread as vitreous or sub retinal seeds (Fig. 2). A characteristic feature highly supportive of the diagnosis is visible calcication within the tumour on ophthalmoscopy. This might also be detectable on ultrasonography [16].

5. Presentation Unilateral and bilateral cases differ in the manner and timing of presentation. Bilateral cases often present with poor vision, and nystagmus or searching eye movements as a result of the tumour involving the central part of the retina (macula) in both eyes (Fig. 2), or leukocoria (white reex in the pupil) noticed by the parents in one or both eyes

Fig. 2. Multiple tumours in both eyes in a 4 month old with germline retinoblastoma.

Fig. 3. White reex on digital photography leucocoria.

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The diagnosis can be conrmed histologically if the eye is removed surgically (enucleation see below). Characteristic histological features include abnormal patterns of retinoblasts such as the FlexnerWintersteiner rosettes, Homer Wright rosettes, and eurettes. 7. Investigations 7.1. Imaging B scan ultrasonography: conrms the presence of masses in the posterior segment of the eye. Characteristic ndings are intra-lesional calcication with high internal reectivity and acoustic shadowing. CT/MRI scan: apart from ultrasonography, imaging is not routinely indicated. MRI may be useful if there is suspicion of extraocular (particularly intracranial) spread, if the child presents with signs of raised intracranial pressure (to look for pinealblastomatrilateral retinoblastoma) or if the diagnosis is in doubt [18]. X-ray and CT scan are best avoided as any dose of radiation magnies the risk of secondary malignancies in germline cases. [19]. 7.2. Genetic testing Mutation testing: is an essential investigation, and can be performed on peripheral blood, and tumour tissue if available (from the enucleated eye). This often helps distinguish between germline and somatic cases, which has major implications for determining the risk to unaffected relatives, future siblings and offspring. 8. Staging The new international classication of intraocular retinoblastoma has 5 groups A to E of increasing severity. There is also a staging system for extraocular disease [20]. 8.1. Differential diagnosis

These can easily be distinguished from retinoblastoma by ophthalmologists with experience in retinoblastoma management [17]. 9. Treatment Retinoblastoma has evolved from a deadly childhood cancer to a largely curable cancer within the past 40 years. Current treatment strategies aim to salvage the eye and provide the best visual outcome possible. This requires signicant multidisciplinary input and should be coordinated by a specialised centre. The various modalities of treatment are: Laser treatment: laser treatment is suitable for primary treatment of smaller tumours, or larger tumours after they have been shrunk to a treatable size with chemotherapy (chemoreduction). Laser treatment is however not effective for vitreous seeds. Laser is delivered through dilated pupils using the indirect ophthalmoscope or microscope. The two common laser wavelengths are 532 nM green light and 810 nM infrared light. Large spot 810 nM treatment heats the tumour slowly (thermotherapy) to produce necrosis and is preferred by many centres [21]. Cryotherapy: using a special probe applied through the sclera to produce temperatures as low as 60 to 80 C results in cryonecrosis of tumours. This treatment is suitable for larger, peripheral tumours, or localized vitreous disease close to the retina. Radiotherapy: this could be in the form of external beam radiotherapy (EBRT, teletherapy) or using a radioactive plaque (brachytherapy). Once the mainstay of treatment, EBRT is now reserved for diffuse disease in the only remaining eye, or recurrent disease not responsive to all other forms of treatment. External beam radiotherapy has signicant risks including induced secondary malignancies (major risk in germline cases) within the radiation eld, cataracts, dry eyes and soft tissue and bony atrophy. In contrast, plaque brachytherapy involves attaching a radioactive plaque of Iodine 131I or Ruthenium 106Ru onto the sclera for a specied period (hours to days) to deliver a high dose of radiation to a localized area without the risks of EBRT. It is highly effective against localized vitreous disease and for elevated tumours where laser is ineffective [22,23]. Chemotherapy: a very valuable addition to our armamentarium over the past 2 decades, its main role is to shrink the tumours to a size where laser treatment can be effective (chemoreduction). It is also very effective against vitreous and sub-retinal diseases, and invaluable for extraocular involvement and metastases. Common regimens include carboplatin, etoposide, and vincristine (referred to as JOE or CEV chemotherapy). There are signicant short and long term side effects of chemotherapy, including hearing loss with carboplatin, and nephrotoxicity [24,25]. Typically, this involves 46 cycles at 3 weekly intervals. Enucleation: is the oldest treatment, and is curative for intraocular retinoblastoma. It is the treatment of choice for advanced uniocular disease or the worse eye of bilateral cases [26]. The eye is removed with a long segment of optic nerve and sent for histology, and tumour DNA studies to identify the mutations. These mutations are then tested against peripheral blood to establish if it is a germline or somatic case. It is usual to replace the lost volume of the enucleated globe with a spherical implant of appropriate size depending on the age of the child. The implant material may be porous such as porous polyethylene (Medpor) or hydroxyapatite, or bioceramic, or non porous such as silicone or acrylic. A prosthetic shell painted to match the other eye is tted in due course for cosmesis. Newer treatment modalities: over the past few years, research has focused on developing techniques for local delivery of chemotherapy to minimize systemic adverse effects. This includes i. Periocular injections of carboplatin and other agents to increase the intraocular levels of the drug and enhance efcacy.

1. Congenital cataracts present with a white pupil (leukocoria) 2. Persistent fetal vasculature PFV (previously called persistent hyperplastic primary vitreous PHPV) results from failure of regression of fetal vessels in the vitreous 3. Retinal dysplasia seen in Norrie's disease, Patau's syndrome, Edward's syndrome, Walker Warburg and other neuronal migration disorders 4. Early onset Coat's disease- a vascular retinopathy resulting in exudation of lipid under the retina (Fig. 4). 5. Congenital infections like Toxocara affecting the eye 6. Congenital glaucoma presenting with a cloudy, red and watery eye

Fig. 4. Lesions simulating retinoblastoma 4 week old boy with Coat's disease.

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ii. Direct injection of drugs into the vitreous cavity has been attempted by some centres. However, this technique has not been adopted widely due to the fear of producing extraocular spread of the tumour. iii. The most signicant advance in recent years has been the interventional radiological technique of intra-arterial chemotherapy (IAC). This involves transfemoral artery canulation to allow delivery of a high dose of the chemotherapeutic agent Melphalan into the ophthalmic artery, avoiding systemic side effects. Although long term experience with this technique is lacking, the initial results are encouraging in selected cases [27]. 10. Research directions The relatively small numbers of cases seen by each centre, lack of a satisfactory animal model, and numerous non-comparable staging systems have hindered retinoblastoma research in the past. The widely accepted new international classication system, and multicentre studies currently underway will pave the way for a more evidence based approach to retinoblastoma management. Current research is directed towards 1. A better understanding of genotypephenotype relationships in retinoblastoma that will be useful in the multidisciplinary management of this disease. 2. Improvements in local drug delivery methods will address the problem of systemic toxicity from existing chemotherapy regimens. 3. Minimising the side effects of treatment 4. Developing better animal models 5. Exploring biologic treatment e.g. anti angiogenic agents, growth factors etc. It is likely future research will be directed towards targeted molecular therapy to individualize treatment, and gene therapy to prevent tumour formation. 11. Treatment principles Retinoblastoma is a unique cancer by virtue of its connement within the scleral envelope, and has a 95% cure rate with appropriate treatment. The diagnosis is clinical, and it is important to avoid breaching this envelope with an intraocular procedure such as diagnostic biopsy to avoid the mortality associated with extraocular spread. A combination of treatment modalities e.g. chemotherapy with laser, cryotherapy or plaque brachytherapy helps minimize adverse effects [28]. Close monitoring with examinations under anaesthesia (EUA) at decreasing frequency as the child grows older is important for early detection of recurrent or new tumours, with examinations without anaesthesia for older children. During active treatment, chemotherapy is given over 46 cycles at 3 weekly intervals, with EUAs before each cycle to monitor response and apply local treatment (laser or cryotherapy). Local treatment may be continued at further EUAs until all tumours are inactive [29,30]. A typical care pathway is shown in Table 1. 12. Supportive treatment 1. Prosthesis tting for enucleated eyes is an important part of rehabilitation, usually a few weeks after surgery. 2. Psychological support for children and families to deal with loss of eye, vision and a chronic illness 3. Protective eye wear for the better/remaining eye during contact sport

4. Long term oncological surveillance especially for germline cases this is best undertaken by oncologists. 5. Counseling parents should be counseled soon after diagnosis, and the patients when they reach adolescence about the inheritance of retinoblastoma, and risk to siblings and offspring. The risk of secondary malignancies, advice about risk factors like smoking and how to look out for early warning signs should be discussed with adolescent patients. 6. Cataract surgery if needed should be delayed for at least 12 years after active treatment 13. Prenatal diagnosis the role of imaging and tissue sampling If there is a family history of retinoblastoma and the mutation in the affected parent is known, there are several options to prevent retinoblastoma or enable early detection i. Pre-implantation genetic diagnosis (PIGD) involves screening embryos at the blastocyst stage. One cell is removed from the embryo to look for the mutation. Unaffected embryos are selectively implanted ensuring the fetus is born free of the retinoblastoma mutation and does not require screening. Additionally, there is no risk of second cancers, and no risk to future generations. The obvious disadvantage of this technique is the need for in-vitro fertilization (IVF) to produce and then select embryos [31]. ii. Chorion villous sampling (CVS) or amniocentesis are interventional techniques to obtain fetal tissue samples prenatally and test for the RB mutation [32]. iii. Prenatal ultrasound in cases with a family history, and particularly if the child is shown to carry the mutation on CVS or amniocentesis, B scan ultrasonography may be advisable in the later stages of pregnancy to look for development of tumours. Although the sensitivity of this technique is quite low, in experienced hands, it may be possible to detect large tumours developing in the last few weeks of pregnancy. Some centres advocate induction of labour at 36 weeks to enable treatment in such cases. There can however be some difculty giving chemotherapy to preterm babies, and treatment may have to be delayed until term. There are also increased risks of surgical intervention such as caesarean section and forceps in induced births. As a result, there is no consensus on this subject. iv. Cord blood testing. The obstetric team can collect cord blood and send it to a reference laboratory to test for the Rb mutation. v. Free fetal DNA testing. This exciting new technique involves testing free (extracellular) fetal DNA which is known to cross the placental barrier. In cases where the mother is healthy and the father is the mutation carrier, maternal blood is tested using DNA amplication. If the mutation is found in maternal blood, one can deduce that it has come from the fetus which must be carrying the mutation. This can then be conrmed at birth with cord blood testing. 14. Screening for retinoblastoma Screening close relatives of retinoblastoma patients is invaluable in early detection and treatment, saving eyes and lives. If the mutation for the index case is known, testing can be offered to relatives to determine if they are at risk of suffering/passing on the disease. Screening is offered if mutation positive or if the mutation is not known for the index case, and risk cannot be excluded. Most centres will have protocols for screening. A suggested protocol is shown in Table 2. Screening is not needed if the relative does not carry

M.V. Parulekar / Early Human Development 86 (2010) 619625 Table 1 Care pathway for management of retinoblastoma.

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Leucocoria / strabismus

Paediatrician / family physician

Ophthalmologist

Optometrist / orthoptic screening

Clinic exam / EUA B Scan, Retcam

Retinoblastoma service

Oncologist

Ophthalmologist

Geneticist

Clinical examination Blood tests

VA, EUA, B Scan, Retcam, NO CT SCAN MRI only if suspicion of extra-ocular extension or hydrocephalus

Family history blood for mutation studies from proband

Multi-disciplinary team meeting

Multi-disciplinary team meeting

Unilateral

Bilateral

E/E A-D E Chemotherapy + laser/cryo Primary enucleation Failure Plaque or intraarterial chemotherapy Failure

E/A-D

Chemotherapy and/or laser/cryotherapy

Chemotherapy + laser/cryo Failure Plaque or intraarterial chemotherapy Failure

Failure Plaque or intra-arterial chemotherapy or secondary enucleation

External beam radiotherapy or secondary enucleation

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the mutation. This approach helps avoid unnecessary screening, and focuses resources. Ideally the rst screening exam for neonates with a close family history of retinoblastoma is performed in the obstetric unit before discharge, and subsequent examinations can be arranged with ophthalmic units familiar with retinoblastoma management. 15. Prognosis 15.1. Prognosis for life Most untreated tumours proceed to local invasion and metastasis to cause death within 2 years. Occasionally however, the tumour may spontaneously stop growing to form a retinoma, or necrose to cause phthisis bulbi (shrunken globe). Most small/medium tumours without vitreous seeding can be successfully treated while preserving useful vision. Overall there is a 95% survival rate (in the developed world). Poor prognostic factors include: size of tumour, optic nerve involvement, extraocular spread and older age at presentation. 15.2. Prognosis for vision The prognosis for vision in retinoblastoma survivors is good, with better than 6/12 vision in the better seeing/remaining eye in over 80% of cases (personal data,). 16. Recurrence Recurrence can develop within the eye in previously treated tumours, and regular follow-up examinations are essential. Delayed extraocular recurrence in the orbit or distant metastases may be seen in a small number of advanced cases that appeared to be restricted to the eye at initial presentation, and/or were negative on metastatic workup. Metastases may occur several decades after the initial presentation. The management of these cases can be challenging [33]. 17. Risk of second cancers and the role of long term surveillance Patients with germline mutations are at increased risk of developing secondary malignancies such as pinealblastoma (trilateral retinoblastoma) [34] ectopic intracranial retinoblastoma, and osteogenic or soft tissue sarcomas, melanoma and bladder cancer [35]. The cumulative risk of second cancers has been reported between 20 and 48% over 50 years in various studies [36]. This risk is increased with radiation exposure [37]. The role of long term screening of retinoblastoma survivors is not clearly dened in the literature. Patient education and health awareness play a key role in minimising delay in diagnosis and treatment of second malignancies in these patients [38].

18. Summary There have been signicant advances in our understanding and management of retinoblastoma over the past few decades. Early detection to minimize visual morbidity remains the focus of research in countries with universal access to healthcare. Late presentation with life threatening disease is a major challenge in economically underprivileged countries, and strategies must focus on raising awareness and acceptance of modern treatment methods through health workers. Developing a global network of retinoblastoma treatment centres that share information and offer advice, and global funding initiatives will save lives and vision. Key guidelines If diagnosed early and treated appropriately, cure rate for intraocular retinoblastoma is over 95%. Prognosis for extraocular disease is very poor. There are 2 forms- genetic and non genetic (somatic). Genetic retinoblastoma often presents in infancy while somatic retinoblastoma usually presents later. Unilateral cases can be somatic or germline. Bilateral cases are alwaysgermline. 90% cases are sporadic. Prenatal diagnosis is available, including PIGD. Leucocoria and squint are the commonest presenting features. The diagnosis is essentially clinical. Ultrasound may be supportive. Diagnostic biopsy is strictly contra-indicated due to risk of extraocular spread. CT scan is best avoided due to the risk of inducing second cancers. Treatment involves the use of multiple modalities including systemic chemotherapy and local treatment such as laser, cryotherapy and plaque brachytherapy. External Beam Radiotherapy is reserved for resistant cases. Surgical removal of the affected eye (enucleation) is offered where there is little prospect of saving the eye. Screening of at risk relatives is recommended to enable early detection. There is an increased risk of second cancers in other organs in germline cases. There is a role for patient education and counseling regarding the risks, advocating a healthy lifestyle and to promptly seek medical advice. Acknowledgements My patients and their families who deal with the disease with such dignity and bravery, and my colleagues in the Birmingham Children's Hospital Retinoblastoma Unit who make my work enjoyable and from whom I learn so much. References

Table 2 Screening protocol. Screening of the immediate family Fundus examination of all siblings and both parents at the time of proband diagnosis Siblings of retinoblastoma sufferers or offspring of parents with retinoblastoma Dilated fundus examination within a week of birth, then Monthly to 6 months of age 2 monthly to 2 years of age 3 monthly to 3 years of age 4 monthly to 4 years of age 6 monthly to 10 years of age Screening to be discontinued only if genetic testing results show patient does not carry the RB mutation. Where Rb mutation is not proven in parent, patient should be advised to seek genetic counselling at age 16+.

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