Brian D. Smith

Effects of the calcium-mediated enzymatic cross-linking of membrane proteins on cellular deformability.

SummaryExcess calcium binding affects the shape and dynamics of cellular deformation of human erythrocytes. It may be hypothesized that incorporation of calcium may modify cellular deformability by processes which include specific cross-linking of membrane proteins with resultant changes in cell shape and deformability. Since previous studies indicate that accumulation of calcium ions causes development of γ-glutamyl-ɛ-lysine bridges in membrane proteins, under control of a membrane transamidating enzyme which specifically requires calcium ions for activation, experiments were devised to examine the relationship between cross-linking and deformability and to determine the effects of specific inhibitor of membrane protein cross-linking on the calcium-dependent modification of erythrocyte to the echinocytic shape. The elastic shear modulus of the membrane was not significantly affected by calcium-induced cross-linking, indicating that induced shape change, not altered elasticity, causes the observed reduction in cellular deformability. These findings support the interpretation that Ca++-induced and transamidase-catalyzed cross-linking of membrane proteins contributes to fixation of altered cellular shape and decreased cellular deformability.

Ocular side effects following intravitreal injection therapy for retinoblastoma: a systematic review

Purpose To describe the ocular side effects in patients receiving intravitreal injection therapy (IViT) for retinoblastoma. Methods PubMed (1946–present), Scopus (all years), Science Citation Index (1900–present) and Conference Proceedings Citation Index—Science (1990–present) electronic databases were searched to identify all published reports of therapeutic intravitreal injections for retinoblastoma in humans. Results Ten studies with original IViT ocular side effect data were included in this systematic review. In these combined reports, a total of 1287 intravitreal injections were given to 306 eyes of 295 patients, with a mean follow-up of 74.1 months. Two hundred sixty-one (88.5%) patients received comparatively standard melphalan IViT doses (8–30 mcg). Ocular side effects occurred in 38 patients (17 significant, 21 minor). The proportion of patients experiencing potentially significant ocular side effects following standard melphalan IViT regimens was 0.031 (8/261; 95% CI 0.013 to 0.06). The side effects of these eight included iris atrophy in three, two each with chorioretinal atrophy and vitreous haemorrhage and one with retinal detachment. Of the other nine patients with significant complications, five experienced sight-threatening complications following dramatic dose escalations (four with melphalan, one with thiotepa), three experienced complications that are commonly associated with concurrent therapies given to these patients and one had a retinal detachment. Of the 61 patients receiving IViT via safety-enhancing injection techniques, all six significant side effects were either attributed to the therapeutic dose or confounded by concurrent treatments. Conclusions Significant ocular complications following IViT for retinoblastoma are uncommon, and this risk may be reduced further by the use of careful injection technique and standard dosing regimens. Care must be taken in the dosing of intravitreal treatments to avoid potentially irreversible vision loss.

Evaluating the risk of extraocular tumour spread following intravitreal injection therapy for retinoblastoma: a systematic review

Background Intravitreal injection therapy (IViT) for retinoblastoma has shown promise in the treatment of vitreous seeds; however, the potential for tumour dissemination following intravitreal penetration has limited its use. This review evaluates the risk of extraocular tumour spread in patients receiving therapeutic intravitreal injections for retinoblastoma. Methods PUBMED (1946–present), SCOPUS (all years), Science Citation Index (1900–present) and Conference Proceedings Citation Index—Science (1990–present) electronic databases were searched to identify all published reports of IViT for retinoblastoma in humans. Results 14 studies with original IViT data were included in this review. A total of 1304 intravitreal injections were given in 315 eyes of 304 patients, with one report of extraocular tumour spread and one patient in whom intravitreal treatment could not be excluded as a contributor to metastatic disease. The proportion of subjects with extraocular tumour spread potentially due to IViT in these combined reports was 0.007 (95% CI 0.0008 to 0.0236), with a mean follow-up of 72.1 months. In a subset of 61 patients receiving IViT via safety enhancing injection techniques (347 injections, 19.6 months mean follow-up), there were no reports of tumour spread. Conclusions Local and systemic tumour spread following IViT in cases of retinoblastoma is rare, and this risk is potentially reduced by the use of safety enhancing injection techniques. These results suggest that the risk of tumour spread should not preclude IViT use for carefully selected patients as part of multi-modal globe salvaging therapy.

Combined intravitreal and subconjunctival carboplatin for retinoblastoma with vitreous seeds

Background To describe the technique of intravitreal chemotherapy preceded by subconjunctival chemotherapy for the treatment of vitreous seeds in advanced stage retinoblastoma. Methods This non-comparative interventional case series retrospectively reviewed the medical records and postenucleation histopathological findings of two patients who presented within weeks of each other with bilateral retinoblastoma, Reese–Ellsworth (R-E) stage Vb in the worse eye. Both patients had failed systemic chemotherapy prior to receiving a single treatment of 0.5 ml (5 mg per 0.5 ml) of subconjunctival carboplatin, through which 0.05 ml (3 mcg per 0.05 ml) of carboplatin was injected into the vitreous (Case 2 received 0.1 ml of intravitreal carboplatin). The subconjunctival chemotherapy was given to reduce the risk of orbital tumour seeding following intravitreal injection. Following enucleation, ocular toxicity and the presence or absence of viable tumour cells at the intravitreal injection site were recorded. Results Histopathological examination did not reveal patency of the pars plana intravitreal penetration site in either case at 6 weeks post-treatment nor was malignant seeding detected in the area of injection. Examination of the two enucleated eyes did not demonstrate structural toxicity to the cornea, anterior segment, iris or retina. Additionally, both cases were followed for over 37 months post-treatment, without the occurrence of orbital malignancy. Conclusions Injecting a bleb of subconjunctival chemotherapy prior to intravitreal drug delivery appeared to mitigate the risk of orbital tumour seeding in two patients with advanced stage retinoblastoma. Incorporating this technique may allow further investigation of intravitreal chemotherapy for the treatment of vitreous seeds in retinoblastoma.

Urine catecholamine levels as diagnostic markers for neuroblastoma in a defined population: implications for ophthalmic practice

PurposeAlthough elevated urinary catecholamine levels have been reported in 90–95% of patients with neuroblastoma, more recent studies of pediatric Horner syndrome caused by an underling neuroblastoma have reported normal values at presentation. The purpose of this population-based study is to report the percentage of cases of neuroblastoma with elevated urinary catecholamine levels at presentation and to suggest a recommended work-up for cases of idiopathic pediatric Horner syndrome.MethodsThe medical records of all pediatric (<19 years) residents of Olmsted County, Minnesota diagnosed with neuroblastoma from 1 January 1969 through 31 December 2008 were retrospectively reviewed.ResultsA total of 14 patients <19 years of age were diagnosed with neuroblastoma as residents of Olmsted County, Minnesota, during the 40-year study period. A total of 10 (71%) of the 14 cases manifested elevated urinary catecholamine metabolites at the initial presentation. Urinary vanillylmandelic acid (VMA) levels were greater than twice the upper limit of normal in eight (57%) of 14 cases, whereas homovanillic acid (HVA) levels were greater than two times the upper limit of normal in 10 (71%) of the 14 cases. Three (75%) of the four cases without significantly elevated urinary VMA or HVA levels were diagnosed with stage IV disease, whereas one (25%) had stage II neuroblastoma.ConclusionUrinary catecholamine levels were significantly elevated at presentation in 10 (71%) of the 14 neuroblastoma cases during the 40-year study period, suggesting that greater emphasis be placed on performing a thorough physical examination and obtaining warranted imaging studies in cases of idiopathic pediatric Horner syndrome.

Incidence, Ocular Manifestations, and Survival in Children with Neuroblastoma: A Population-Based Study

PURPOSE To determine the incidence, ophthalmic manifestations, and survival among children with neuroblastoma in a defined population. DESIGN Population-based retrospective cohort. METHODS The medical records of all pediatric (<19 years) residents of Olmsted County, Minnesota, diagnosed with neuroblastoma from January 1, 1969, through December 31, 2008, were retrospectively reviewed. RESULTS Fourteen children were diagnosed with neuroblastoma as residents of Olmstead County, Minnesota, during the 40-year period, yielding an age- and gender-adjusted incidence of 11.8 (95% confidence interval [CI]: 5.6-18.0) per million patients <15 years of age. The calculated incidence for patients presenting before the age of 5 in this cohort was 1 in 5970 children (95% CI: 3920-12 580 children). The mean age at diagnosis for the 14 study patients was 22.5 months (range, 10.4-42.6 months). Six of the 14 (43%; 95% CI: 18%-71%) had ocular manifestations, including orbital metastasis in 6 (100%), proptosis and ecchymosis in 4 (67%), ptosis in 2 (33%), and strabismus in 1 (17%). The Kaplan-Meier rate of survival for all 14 children was 57% at 1 year (95% CI: 36%-90%) and 50% at 5 years (95% CI: 30%-84%), while the 6 with eye findings had a survival rate of 17% at 9 months (95% CI: 3%-100%). CONCLUSIONS The incidence of neuroblastoma in this population was 11.8 per million patients <15 years, with ophthalmic involvement observed in 6 of the 14 study patients (43%). Orbital metastasis in the 6 children in this cohort was associated with poor prognosis.

Biochemical factors influencing erythrocyte deformability and capillary entrance phenomena

Evaluation of the effects of biochemical modification of erythrocytes by low pressure filtration through Nuclepore filters of 2.8 and 3.0 micrometer pore diameter and glass micropipettes of 2.7 and 3.0 micrometer internal diameter and channel length greater than 100 micrometer indicates the relative sensitivity of the micropipette in detection of reduction of cellular deformability. In dynamic studies of erythrocyte entrance behaviour on micropipettes, the significance of induced relative sphericity on cellular deformability was evident. Reduction of intracellular ATP, cellular pH, sphere induction by bifunctional membrane protein cross-linking reagents and lead caused diminution in cellular deformability, attributable to the more spherical shape. Similarly echinocyte formation by effects of aged plasma, oleate and senscence, and stomatocyte induction by chlorpromazine caused marked decrease in capillary entry of cells having flow velocities of approximately 100 micrometer/s in contrast inhibitors of the glucose carrier, of calmodulin and manipulated decrease in membrane lipid of bilayer fluidity did not cause detectable change in cellular deformability. These data emphasize the relative importance of erythrocytic shape as a determinant of cellular deformability and suggest that necessity for adequate control of shape change is necessary when examining effects of biochemical manipulations on membrane elasticity and cellular deformability. The preliminary date from capillary entrance experiment indicate the importance of dynamic experiments in in vitro studies of microcirculation.

Viscoelastic Properties of Normal and Pathologic Human Granulocytes and Lymphocytes

Leukocytes differ significantly from normal erythrocytes in cytostructure, characteristics of plasma membrane, relative sphericity in vivo, and in phagocytic types, the energy dependent mechanisms which permit motility and phaqocytosis. Their rheologic behavior in vivo differs importantly from that of erythrocytes (1–3), flow is significantly slower (2) and the interactions of leukocytes with endothelium and erythrocytes influence blood flow (2,3). Granulocytes and lymphocytes can traverse capillaries of 4–8 µm dia. at relatively low pressures (3) however individual cells may obstruct flow at forces exceeding those in capillaries, with resulting obstruction presumed to be due to the leukocytes’ relative sphericity, surface properties in relation to endothelial cells, positioning in the flow and unknown factors. Pathologic cells from leukemic states are presumed to affect blood rheology adversely because of abnormalities of cell deformability associated with atypical cell development (4).

Preclinical acute ocular safety study of combined intravitreal carboplatin and etoposide phosphate for retinoblastoma

BACKGROUND AND OBJECTIVE To describe the ocular toxicity of intravitreal carboplatin and etoposide phosphate (VP16P) in Dutch-Belted rabbits. MATERIALS AND METHODS Twenty-two adult male Dutch-Belted rabbits (Cohort 1) each received a single, bilateral intravitreal injection (0.05 mL). For Cohort 1, safety was assessed via electroretinograms (ERGs) and ocular examination. Of nine total groups in Cohort 1, the first five received the following single agents: Group 1: normal saline; Group 2: VP16P 75 µg; Group 3: VP16P 100 µg; Group 4: carboplatin 4 µg; and Group 5: carboplatin 8 µg. Groups 6 through 9 received the following combination of carboplatin/ VP16P, respectively: Group 6: 8 µg/75 µg, Group 7: 8 µg/50 µg, Group 8: 4 µg/50 µg, and Group 9: 2 µg/25 µg. Cohort 2 consisted of 15 Dutch-Belted rabbits in seven groups (Groups 10 through 16), each receiving a single, bilateral intravitreal injection. For Cohort 2, safety was assessed via histopathology. RESULTS Groups 2 through 8 demonstrated a statistically significant decrease (relative to Group 1) in at least one ERG waveform amplitude obtained 4 weeks postinjection (P < .05). Group 9 (carbo 2 µg/VP16P 25 µg) did not manifest ERG toxicity. Fundoscopic toxicity consisted of slight-to-moderate attenuation of vessels in rabbits receiving doses above carbo 4 µg/VP16P 50 µg. Histopathologic retinal toxicity (Cohort 2) was dose-dependent, ranging from full-thickness atrophy in rabbits receiving the highest dose to normal in rabbits receiving carbo 2 µg/VP16P 25 µg. CONCLUSIONS Combined carboplatin and VP16P may be compatible for intravitreal injection therapy, and a single dose of 2 µg/25 µg appears to be safe in a rabbit model. These agents may be a safer alternative to intravitreal melphalan (Alkeran; GlaxoSmithKline, Brentford, United Kingdom) for the treatment of vitreous seeds in retinoblastoma. [Ophthalmic Surg Lasers Imaging Retina. 2017;48:151-159.].

Chemotherapy for Adult Tumors

Chemotherapeutic agents continue to play an important role in the treatment of orbital disease in adult patients. The last decade has seen significant changes in both agents used and methods of delivery, with new, effective therapies now available in clinical practice. Local therapy has changed the treatment paradigm for ocular surface squamous neoplasia and adjuvant intra-arterial chemotherapy has shown promising results in the treatment of adenoid cystic carcinoma. Novel systemic therapies are available for basal cell nevus syndrome, malignant melanoma, and more, with numerous drugs undergoing phase III clinical trials, several with promising initial results. The breadth of pathology involving the orbit necessitates a broad range of chemotherapeutic agents and delivery methods. This chapter reviews historical chemotherapeutic treatments while focusing on newer therapies that are changing the treatment paradigm of adult orbital disease.