Retinoblastoma: Comprehensive Analysis of the Disease and Treatments

This report provides a comprehensive overview of Retinoblastoma, an intraocular cancer primarily affecting young children, caused by mutations in the RB1 gene. It delves into the background of the disorder, its inheritance patterns, and the pathology of the tumor, including its growth and characteristics. The report explains the role of the defective RB1 gene, the normal function of the wild-type protein, and the signaling pathways involved in cell cycle regulation. It also discusses the latest research, including epigenetic mechanisms and the role of SYK, along with current treatment management strategies such as chemotherapy, cryotherapy, and enucleation. Furthermore, it touches upon future therapeutic approaches and concludes with a discussion of the disease's impact and ongoing efforts to improve outcomes for affected individuals.

Running head: RETINOBLASTOMA
Retinoblastoma
Name of the Student
Name of the University
Author Note

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Abstract
Retinoblastoma is an intraocular cancer that is caused by the loss of either single or both the
copies of Rb gene. Retinoblastoma (Rb) protein was first identified as the product of the
prototype tumour-suppressor gene (RB).The loss of the normal function of the RB gene
arising out of the mutation is associated with the disease retinoblastoma, which is hereditary
in nature.The disease primarily affects children who are below 5 years of age. Rb gene is
normally involved in the cell cycle progression and faults in the phosphorylation leads to the
tumour development. Retinoblastoma is extracted from the cells, which are from
neuroectodermal origin and is present in the inner layer of the optic cup. The tumour usually
grows toward the subretinal space or vitreous via forming a multilobulated white mass.
At present treatment of retinoblastoma leads to either enucleation or chemotherapy or
cryotherapy. However, latest research is being carried out to elucidate novel therapeutic
approach.

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Table of Contents
Background of the disorder........................................................................................................3
Current knowledge of inheritance pattern..................................................................................3
Pathology....................................................................................................................................5
Information on defective gene...................................................................................................6
Normal role of wide-type protein...............................................................................................7
Signalling pathway of wild type protein....................................................................................8
Defect in Rb1 gene and problem phenotype..............................................................................9
Latest research..........................................................................................................................10
Treatment management........................................................................................................11
Future therapies....................................................................................................................12
Conclusion................................................................................................................................12
References................................................................................................................................14

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Background of the disorder
Retinoblastoma (Rb) protein was first identified as the product of the prototype
tumour-suppressor gene (RB). The proteins arising out of the tumour-suppressor genes
function in different ways. This inhibits the faulty progression through the cell cycle. The loss
of the normal function of the RB gene arising out of the mutation is associated with the
disease retinoblastoma, which is hereditary in nature. A child suffering from retinoblastoma
inherits one normal RB+ allele from one parent and another mutant allele, RB- from another
parent. If the RB+ allele is mutated to RB- allele, then the functional protein is not expressed
and the cell becomes cancerous. This mutation from RB+ to RB- occurs in the retinal cells,
leading to the formation of the retinal tumours and thus the name of the disease,
retinoblastoma. The mutation or the inactivation of RB gene occurs via phosphorylation
(Alberts et al. 2013).
Current knowledge of inheritance pattern
A hereditary predisposition for certain types of cancers is prevalent among the
individuals who are suffering from inherited mutations in the tumour suppressor genes.
Susceptible individuals inherit germ-line mutation in a single allele of one gene and the
somatic mutation in the corresponding allele promotes tumour progression. Retinoblastoma is
the first tumour suppressor gene identified in the signalling pathway. It is a classic case of
loss of function of the RB gene. Children who are suffering from hereditary retinoblastoma
inherit a single copy of the defective gene RB from one parent. Defects in the retinoblastoma
arise out of the deletion in the one of the gene residing in chromosome 13. The children
suffering from hereditary retinoblastoma develop retinal tumours in both the eyes during the
early stages of life, leading to blindness. Individuals who are suffering from the sporadic
retinoblastoma inherit two normal RB alleles from both the parents. This normal allele

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undergoes a loss of function somatic mutation in the single retinal cell. In sporadic
retinoblastoma, two separate mutations occur in a particular retinal call or its progeny
produce homozygous recessive allele (RB-/RB-). However, losing both the copies of the RB
gene is less than loosing single copy, the conditions like sporadic retinoblastoma, is rare and
develops during the later stages of life and usually affects one eye(Alberts et al. 2013).
Figure: Gene mutation and transmission in Retinoblastoma
(Source: Alberts et al. 2013)
Removal of the retinal tumours before the achievement of the malignancy helps in
cutting down the severity of the disease. Such treatment when conducted in children who are
suffering from retinoblastoma helps them to survive up to their adulthood. However, since

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their germ cells contain one mutant RB allele and one normal allele, these individuals on an
average pass the mutant allele to half of their upcoming generation and normal allele to
another half of the generation. Children who inherit both the normal allele leads a normal life,
the condition is relevant only when both their parents also have two normal RB alleles.
However, the children inheriting the mutant allele have a chance of developing retinal
tumours like their affected parents, even if they have inherited one normal allele from one of
their normal parent. Thus, the tendency of development of the retinoblastoma is inherited as a
dominant trait.
Pathology
The cell in the inner layers of the optic cup, which comes from the neuroectodermal
origin is the site of occurrence of Retinoblastoma. The tumour expands toward the subretinal
space or vitreous via forming a multilobulated white mass. This white mass is associated with
vitreous or subretinal seeds. This expansion of tumour occurs under the influence of the
vascular supply while keeping a distance of 90 to 110 micro-meters from the central vascular
channel. The tumour is targeted through ischemic necrosis. At times, it constitutes areas
highlighting dystrophic calcification and thereby generating a visible cottage cheese or
chalky appearance. Tumour also grows in a diffuse state inside the retina via extracting its
nutrition from the source of retinal vasculature. Both the subretinal and vitreous seeds of
retinoblastoma are avascular in nature. It constitutes diverse areas of necrosis located at the
centre of the seed that is away from the subretinal fluid and vitreous (Grossniklaus 2014).

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(Source: Grossniklaus 2014)
Information on defective gene
RB1 gene is the first ever tumour suppressor gene identified. The mutational
inactivation of RB1 gene causes human cancer, retinoblastoma which is common among the
children (Chinnam and Goodrich 2011). Apart from causing retinoblastoma, the loss of
function of RB1 gene is also related with several other cancer, which arise molly during the
later stages in life like carcinoma of lungs, breast cancer and bladder cancer (Alberts et al.
2013).

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Normal role of wide-type protein
Retinoblastoma protein (RB) is associated with several cellular processes like
regulation of cell cycle, regulation of DNA-damage responses, DNA repair mechanism, DNA
replication and shield against premature apoptosis and cell differentiation. RB exerts its affect
during the first 2/3rd of the G1 phase of the cell cycle. RB performs a notable alteration
during the R (restriction) point of the cell cycle. Through the ongoing or the early hours of
the G1 phase of the cell cycle, RB remains de-phosphorylated however, a bulk of RB
produced in the cell during the last hour of the G1 phase of the cell cycle remains
hyperphosphorylated. RB abides the hyperphosphorylated form throughout the remainder of
the cell cycle, while only losing its multiple phosphorylated groups only during the
emergence of the M phase of the mitosis phase of the cell cycle. This phosphorylation of the
RB leads to the inactivation of the growth inhibitory functions of RB.
Figure: Cell signaling via Rb

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Source: Bertoli et al. 2013
Signalling pathway of wild type protein
RB protein helps in the cell cycle progression from G1 to the S phase of the cell cycle.
The activity of the RB gene is regulated via phosphorylation as the cell gradually progresses
through different stages of cell cycle. To be precise, RB is phosphorylated via the action the
of cyclin dependent protein kinase of cyclin D, Cdk4/6 and this phosphorylation events take
place as the cell progress through the restriction point in G1. While unphosphorylated during
the G0 or early G1 phase of the cell cycle, Rb binds to transcription factor, E2F. This
transcription factor in turn regulates the activity of other cell cycle modulating proteins. E2F
binds to its target sequence both in the presence or absence of the Rb. However, when E2F
remains in a bonded form with Rb, it fails to undergo the transcription of its targeted gene
sequence. Thus, Rb acting as a suppressor, suppressing the transcription of the E2F mediated
genes. The phosphorylation of Rb under the action of Cyclin D/Cdk 4/6 leads to structural
changes in Rb, promoting the release of Rb from E2F and facilitating the transcription of E2F
regulated genes and thereby leading to progression in cell cycle. Thus, it concluded that Rb
acts as a molecular switch that convert E2F from a repressor to an activator upon
phosphorylation, leading to cell cycle progression. The regulation of Rb via Cyclin D/Cdk4/6
in turn is regulated via the presence of growth factors that provides positive signals in the cell
cycle progression (Cooper and Hausman2000).

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Figure: Cell cycle regulation via Rb and E2F
Source: Created by author
Defect in Rb1 gene and problem phenotype
During the disease condition, The RB gene gets phosphorylated even under the
absence of the growth factor response and this leads to the progress through the cell cycle
even under faulty condition, leading to the generation of the disease. The developmental
defects, which are manifested upon the loss of Rb1 is attributed to both non-cell autonomous
and cell autonomous mechanisms. For example, in the absence of Rb1, there ocuurs a failure
of proper placental development and macrophage differentiation of the liver. This contributes
to failure in proper erythrocyte maturation. On the other hand, hematopoietic defects arising
out of Rb1 is expressed in the Central Nervous System (CNS). Thus loss of function of Rb1
gene generated both indirect and direct effects on the cell division that contributes the
generation of faulty phenotypes (Chinnam and Goodrich 2011).

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Latest research
The sequencing of genome of retinoblastoma revealed that it has relatively stable
genome sequence (Zhang et al. 2012). Thus, epigenetic mechanism is responsible for the
tumour genesis of retinoblastoma. Numerous known oncogenes along with tumour
suppressor genes that have specific regions of histone modifications along with altered DNA
methylation, correlate the overall change in the gene expression. According to Zhang et al.
2012, Spleen tyrosin kinase (SYK) is an important determinant in retinoblastoma. SPK is a
progenitoronco-gene. The progenitor cells of retina and the neurons of the retina express no
or rather say little SYK. Moreover, SYK has no reported function in the process of
development of the visual system. However, the epigenetic analysis showed SYK as an
important oncogene behind the development of retinoblastoma (Zhang et al. 2012).
The diagnosis of the retinoblastoma is done via fundoscopy under general anaesthesia.
The entire test procedure is performed via an ophthalmologist. This particular test helps in the
elucidation of several parameters related to tumour like number, laterality, size, tumour
seeding, sub retinal space and the specific anterior segment of the tumour. Further diagnostic
imaging in this field assists in unfolding other associated brain abnormalities like intracranial
tumour extension, midline intracranial primitiveneuroectodermal tumour (PNET) and other
related brain malformations in retinoblastoma patients. The malformations are mostly severe
in patients who are suffering from 13q deletion syndrome (de Graaf et al. 2012).
Conservative treatment strategies are found to be providing successful results if
conducted during the early stages of the disease. The eye preservation therapy has been
foundto provide positive results in recent years. This therapy is majorly based on the motive
of tumour reduction and is given in collaboration with the chemotherapy, laser coagulation,
radioactive plaque or cryotherapy. Recently, in order to treat the additional stages of the
disease, selective ophthalmic artery infusion of a potential chemotherapeutic agent is made

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available. However, imaging is extremely important at local staging. A standardized MRI
protocol is used to the imaging of the retinoblastoma patient before the onset of the treatment.
The imaging technique helps in the process of lesion characterization and detection of the
tumour extent (de Graaf et al. 2012).
Treatment management
At present, the cure rate for retinoblastoma is 85%, which is significant in the field of
positive treatment but this cure of the disease usually occurs at cost of partial or complete loss
of vision. Moreover, there is also an estimated mortality count of 50% in the developing
countries. the patients who are suffering from RB lies at an increased risk of developing three
of the major life-threatening conditions namely metastasis arising out of RB, brain tumour
pinealoblastoma and primary level of cancer. At present the widely used treatment for RB,
include cryotherapy, laser treatment and radiotherapy. Recently chemotherapy is systemic in
nature and the main drugs used for this purpose include carboplatin, vincrisine and etoposide.
These are all broad-spectrum antibiotics with dose limiting side effects like loss of hearing
and other associated secondary cancers (Basavarajappa and Corson 2012).
Retinoblastoma is curable if there are no associated metastatic risk factors. The
restoration of the visual function is dependent on the process of ocular preservation,
anatomical relationships between the tumours of the macula and optic disk and the initial
volume of the tumour. In the presence of the metastatic risk factors, adjuvant treatment
therapy is prescribed in order to restrict a life-threatening relapse (de Graaf et al. 2012).
Retinoblastoma is regarded as the primary cause of the neoplasm in the pair of eyes
during the early childhood. This debilitating disease in the industrialized sectors affects more
than 95% of the children. Survival rate of the metastatic retinoblastoma was initially poor but

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with the advent of the high-dose chemotherapeutic agents followed by the autologous stem
cell rescue (ASCR), the mortality rate of retinoblastoma decreased (Palma et al. 2012).
Retinoblastoma tumour has a heterogeneousmicro-environment that is made of high
angiogenic activity along with low areas of oxygen tensed conditions. This oxygen-tensed
condition becomes more pronounced during the advanced stage of the disease. The tumour
cells, which grown within this hypoxic conditions are found resistant towards
chemotherapeutic radiation. Treatment with glycolytic inhibitor 2-deoxy-2-fluro-D-glucose
(2-FG) helps to reduce the load of tumour burden along with hypoxia in the retinal tumours.
2-FG is more potent glycolytic inhibitor under low oxygen concentration and thus helping to
reduce the tumour in eyes arising out of retinoblastoma (Pina et al. 2012).
Future therapies
Despite significant advancement in the field of RB pathology and disease biology,
there are till date no targeted chemotherapies available for the disease treatment. At present,
efforts are being made to use KIF14, an oncogene for retinoblastoma as a novel target for
therapeutics. Identification of the 1q31 region of the RB gene led to the discovery of KIF14.
KIF14 is over-expressed at the mRNA level with excess genomic gain. Moreover, the over-
expression of KIF14 has been confirmed via carrying a study in an independent RB cohort.
These information are now being significantly utilized by Basavarajappa and Corson
(2012)inorder to develop novel therapeutic approach for RB.
Conclusion
Thus from the above discussion it can be concluded that retinoblastoma is an
intraocular cancer that is caused by the loss of either single or both the copies of Rb gene.
The disease primarily affects children who are below 5 years of age. Rb gene is normal
involved in the cell cycle progression and faults in the phosphorylation leads to the tumour

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development. At present treatment of retinoblastoma leads to either enucleation or
chemotherapy or cryotherapy. However, latest research is being carried out to elucidate novel
therapeutic approach.

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References
Alberts, B., Bray, D., Hopkin, K., Johnson, A., Lewis, J., Raff, M., Roberts, K. and Walter,
P., 2013. Essential cell biology. Garland Science.
Basavarajappa, H.D. and Corson, T.W., 2012. KIF14 as an oncogene in retinoblastoma: a
target for novel therapeutics?. Future medicinal chemistry, 4(17), pp.2149-2152.
Bertoli, C., Skotheim, J.M. and De Bruin, R.A., 2013. Control of cell cycle transcription
during G1 and S phases. Nature reviews Molecular cell biology, 14(8), pp.518-528.
Chinnam, M. and Goodrich, D.W., 2011. RB1, development, and cancer. Current topics in
developmental biology, 94, p.129.
Cooper, G.M. and Hausman, R.E., 2010. The cell (Vol. 85).Sunderland: Sinauer Associates.
deGraaf, P., Göricke, S., Rodjan, F., Galluzzi, P., Maeder, P., Castelijns, J.A., Brisse, H.J.
and European Retinoblastoma Imaging Collaboration, 2012. Guidelines for imaging
retinoblastoma: imaging principles and MRI standardization. Pediatric radiology, 42(1),
pp.2-14.
Grossniklaus, H.E., 2014. Retinoblastoma.Fifty years of progress.The LXXI Edward Jackson
memorial lecture. American journal of ophthalmology, 158(5), pp.875-891.
Palma, J., Sasso, D.F., Dufort, G., Koop, K., Sampor, C., Diez, B., Richard, L., Castillo, L.
and Chantada, G.L., 2012. Successful treatment of metastatic retinoblastoma with high-dose
chemotherapy and autologous stem cell rescue in South America. Bone marrow
transplantation, 47(4), pp.522-527.
Pina, Y., Decatur, C., Murray, T.G., Houston, S.K., Lopez-Cavalcante, M., Hernandez, E.,
Celdran, M., Shah, N., Feuer, W. and Lampidis, T., 2012.Retinoblastoma treatment:

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utilization of the glycolytic inhibitor, 2-deoxy-2-fluoro-D-glucose (2-FG), to target the
chemoresistant hypoxic regions in LHBETATAG retinal tumours. Investigative
ophthalmology & visual science, 53(2), pp.996-1002.
Zhang, J., Benavente, C.A., McEvoy, J., Flores-Otero, J., Ding, L., Chen, X., Ulyanov, A.,
Wu, G., Wilson, M., Wang, J. and Brennan, R., 2012.A novel retinoblastoma therapy from
genomic and epigenetic analyses. Nature, 481(7381), pp.329-334.