Chronic Myelogenous Leukemia Relapse Presenting With Central Nervous System Blast Crisis and Bilateral Optic Nerve Infiltration

To describe the history, clinical findings, and possible pathogenic etiologies of the constellation of neuro-ophthalmic findings discovered in astronauts after long-duration space flight and to discuss the terrestrial implications of such findings.; ; Retrospective review of published observational, longitudinal examination of neuro-ophthalmic findings in astronauts after long-duration space flight; analysis of postflight questionnaires regarding in-flight vision changes in approximately 300 additional astronauts; and hypothesis generating for developing possible future countermeasures and potential implications for neuro-ophthalmic disorders on Earth. Astronauts with neuro-ophthalmic findings, which were not present at the start of a space flight mission and only seen on return from long-duration space missions to the International Space Station, will be discussed.; ; After 6 months of space flight, 7 astronauts had ophthalmic findings consisting of optic disc edema in 5, globe flattening in 5, choroidal folds in 5, cotton-wool spots in 3, nerve fiber layer thickening detected by optical coherence tomography in 6, and decreased near vision in 6. Five of 7 astronauts with near vision complaints had a hyperopic shift ≥+0.50 diopters (D) between pre-/post-mission spherical equivalent refraction in 1 or both eyes (range, +0.50 to +1.75 D). These 5 astronauts showed globe flattening on magnetic resonance imaging. A total of 6 lumbar punctures have been performed to date (4 in the originally described cohort) and documented opening pressures of 18, 22, 21, 21.5, 28, and 28.5 cm H2O. These were performed at 8, 66, 19, 7, 12, and 57 days after mission, respectively. The 300 postflight questionnaires documented that approximately 29% and 60% of astronauts on short-duration and long-duration missions, respectively, experienced a degradation in distant and near visual acuity. Some of these vision changes remain unresolved for years after flight. Several possible pathogenic mechanisms, as well as potential countermeasures and discussion of possible terrestrial implications, are described.; ; We previously hypothesized that the optic nerve and ocular changes that we described in astronauts may be the result of orbital and cranial cephalad fluid shifts brought about by prolonged microgravity exposure. The findings we reported previously and continue to see in astronauts may represent parts of a spectrum of ocular and cerebral responses to extended microgravity exposure. Future investigations hopefully will lead to countermeasures that can be used to eliminate or lessen the magnitude of these potentially harmful findings before long-duration space flight including the possibility of a manned mission to Mars.

Clinical Observation Chronic Myelogenous Leukemia Relapse Presenting With Central Nervous System Blast Crisis and Bilateral Optic Nerve Infiltration Joyce N. Mbekeani, MD, FRCS, FRCOphth, Maaly Abdel Fattah, MD, PhD, Randa M. Al Nounou, MD, Wahiba Chebbo, MD, Mohammed Asif Dogar, MD Abstract: Bilateral, simultaneous optic nerve sheath infiltration as a manifestation of leukemia relapse is very rare. A 45year-old woman with chronic myelogenous leukemia was successfully treated to cytogenetic bone marrow remission 1 year previously and maintained on imatinib. She developed total bilateral blindness with marked, bilateral optic disc edema and evidence of bilateral optic nerve infiltration on magnetic resonance imaging. Cerebrospinal fluid cytology confirmed central nervous system (CNS) blast crisis. She recovered visual acuity of 20/20 in the right eye, and 20/25 in the left eye with salvage systemic and intrathecal chemotherapy before radiation therapy. Our report underscores the importance of timely and aggressive intervention of blast crisis of the CNS and the need for CNS penetrating induction and maintenance therapy. Journal of Neuro-Ophthalmology 2016;36:73-77 doi: 10.1097/WNO.0000000000000326 © 2015 by North American Neuro-Ophthalmology Society L eukemic infiltration of the optic nerves, whether as a manifestation of primary or relapsing disease, occurs infrequently. Indeed, extramedullary relapse with meningeal infiltration and central nervous system (CNS) blast crisis presenting principally with isolated ophthalmic signs is rare, with few reported cases in the literature (1-3). This manifestation Department of Surgery (JNM), North Bronx Health Network, Bronx, New York; Department of Ophthalmology & Visual Sciences (JNM), Albert Einstein College of Medicine of Yeshiva University, New York; Department of Ophthalmology (MAF), King Faisal Specialist Hospital & Research Center, Riyadh; Department of Ophthalmology (MAF), Cairo University Medical School, Cairo, Egypt; Department of Hematologic Pathology (RMAN), King Faisal Specialist Hospital & Research Center, Riyadh, Saudi Arabia; Department of Hematology-Oncology (WC), King Faisal Specialist Hospital & Research Center, Riyadh, Saudi Arabia; and Department of Neuro-Radiology (MAD), King Faisal Specialist Hospital & Research Center, Riyadh, Saudi Arabia. The authors report no conflicts of interest. Address correspondence to Joyce N. Mbekeani, MD, FRCS, FRCOphth, North Bronx Health Network, Department of Surgery, 1400 Pelham Parkway, Bronx NY 10461; E-mail: [e-mail] Mbekeani et al: J Neuro-Ophthalmol 2016; 36: 73-77 leads to visual compromise from direct neural compression, intraneural infiltration, vascular disruption, intracranial hypertension, or a combination of these mechanisms. The current preference of radiotherapy to treat optic nerve infiltration stems from experience with poor CNS penetration by most systemic chemotherapeutic agents. The CNS is thus a sanctuary site for neoplastic cells of chronic myelogenous leukemia (CML). Imatinib, a potent, selective, first-generation tyrosine kinase inhibitor, is used to treat CML in its chronic and blast phases. Tyrosine kinase is produced by an aberrant Philadelphia chromosome present in almost all CML patients (4) that causes uncontrolled growth and survival of hematopoietic progenitor cells. This aberrant chromosome results from chromosome 9q34 (bcr) to chromosome 22q11.2 (abl) segment translocation, creating the CML (bcr/abl) oncogene (4). Reverse transcriptase polymerase chain reaction (PCR) for this gene can quantify bcr/abl and is used to diagnose and monitor CML response to tyrosine kinase inhibitors. Although imatinib, currently a first-line therapy for CML, is very effective in inducing hematopoietic and cytogenetic remission, like most other chemotherapeutic agents, it fails to penetrate the blood- brain barrier, and incidents of CNS blast crisis while on imatinib are increasingly being reported (5-11). We describe the clinical presentation and imaging features of a patient with CML and proven hematologic and cytogenetic remission on maintenance imatinib, who developed CML relapse and CNS blast crisis manifesting as isolated bilateral blindness from optic nerve infiltration. Recovery of visual function without evidence of structural damage was achieved with intravenous and intrathecal chemotherapy. Our case demonstrates both a rare ophthalmic manifestation of CML relapse and recovery without radiation therapy. CASE REPORT A 45-year-old woman with a history of CML underwent neuro-ophthalmic evaluation for profound bilateral vision 73 Copyright © North American Neuro-Ophthalmology Society. Unauthorized reproduction of this article is prohibited. Clinical Observation loss. She was diagnosed with CML 1 year earlier and presented with a 3-month history of 5 kg weight loss, lethargy, dizziness, and epigastric discomfort. On examination, she appeared pale and had hepatosplenomegaly, without ascites or lymphadenopathy. All other systems were normal. Complete blood count revealed decreased hemoglobin of 9.5 gm/dL (normal 12-15 gm/dL), platelets of 316 · 109/L (normal: 150-400 · 109/L), and white blood cells of 73.2 · 109/L (normal: 4-11 · 109/L). The peripheral smear showed 40% polymorphonuclear leukocytes, 6% lymphocytes, 25% myelocytes, 2% promyelocytes, 10% metamyelocytes, and 2% blast cells. The patient was successfully treated with 2 months of hydroxyurea and allopurinol after which imatinib was commenced. She was evaluated at our hospital for further management. Peripheral blood real-time PCR bcr/abl oncogene levels were 8.758% (international scale [IS]), and bone marrow aspiration showed changes consistent with partially treated CML. Subsequent regular PCR measurements revealed gradually declining bcr/abl levels, and after 5 months reached 0.018% (IS), with a log reduction of 2.7, representing major molecular response (MMR #0.1% IS or log reduction of $3.0). When the patient developed rapidly progressive, bilateral, simultaneous visual failure, she also complained of severe headaches. At the time, her only other medication was atenolol for intercurrent hypertension. Visual acuity was counting fingers at 1 foot, right eye, and no light perception, left eye. Pupils were round and sluggishly reactive to light, with a left relative afferent pupillary defect. The anterior segments were normal, eye movements were intact, and ophthalmoscopy revealed severe bilateral optic disc edema (Fig. 1A). General physical and neurologic examinations were unremarkable. Brain and orbital magnetic resonance imaging (MRI) disclosed thickening and enhancement of both optic nerves and similar changes along the tentorium (Fig. 2). There was no evidence of intracranial hypertension. Lumbar puncture revealed bloody cerebrospinal fluid (CSF), with 200 · 106/L red blood cells, 390 · 106/L white blood cells (normal: 0-5 cells), glucose of 3.75 · 106/L (normal: 2.8-4.4 · 106/L), protein of 83.6 mg/dL (normal 15-50 mg/dL), and 25% myeloblasts (Fig. 3A). Fluorescence in situ hybridization analysis was positive for bcr-abl gene mutation in 100% of nuclei of the 22 cells examined. Peripheral blood had normal cellularity, with unremarkable morphology and no blast cells. Cytogenetic testing revealed bcr-abl levels of 0.179% IS (log reduction = 1.743). Bone marrow studies similarly showed normal trilineage hematopoiesis, consistent with controlled or treated CML and hematologic remission (Figs. 3B, 3C). Reticulin stain used to assess the degree of bone marrow fibrosis, a prognostic index in CML, revealed normal reticulin levels (Fig. 3D). CSF and peripheral blood cultures and virology workup were negative. CNS blast crisis was diagnosed, and salvage therapy was commenced with intravenous fludarabine and cytarabine and four cycles of intrathecal cytarabine, methotrexate, and hydrocortisone. One month later, the patient had resolution of headaches, visual acuity of 20/20 in right eye, and 20/25 in left eye, intact color vision, and reactive pupils, with no relative afferent pupillary defect. There was marked improvement in bilateral optic disc edema (Fig. 1B) and no evidence to suggest optic atrophy on optical coherence tomography, with average retinal nerve fiber layer thickness of 115 mm, right eye, and 111 mm, left eye. There was mild bilateral visual field loss, greater in the left eye (Fig. 4). Lumbar puncture revealed no blast cells, and MRI showed significant reduction of optic nerve thickening and enhancement. The patient was discharged 6 weeks after admission in stable condition. Subsequent peripheral blood and bone marrow PCR revealed undetectable bcr/abl oncogene levels, representing complete cytogenetic response. Subsequently, the patient underwent allogeneic bone marrow FIG. 1. A. At presentation, there is bilateral optic disc edema with retinal vascular engorgement and tortuosity. B. Almost complete resolution of disc edema after 1 month of intravenous and intrathecal chemotherapy. FIG. 2. Contrast-enhanced axial (A) and coronal (B) T1 magnetic resonance imaging shows enlargement and enhancement of both prechiasmal optic nerves (arrows) and enhancement of the tentorium (arrowheads). 74 Mbekeani et al: J Neuro-Ophthalmol 2016; 36: 73-77 Copyright © North American Neuro-Ophthalmology Society. Unauthorized reproduction of this article is prohibited. Clinical Observation DISCUSSION FIG. 3. Histopathology of cerebrospinal fluid (CSF) smear and bone marrow. A. CSF smear reveals marked leukocytosis with mostly blast cells (arrows) and basophils (MayGrünwald-Giemsa ·100). B. Bone marrow aspirate shows active hematopoiesis with a normal maturation sequence (Wright-Giemsa, ·100). C. Bone marrow biopsy demonstrates normocellular marrow with active trilineage hematopoiesis (hematoxylin & eosin x50). D. Bone marrow trephine biopsy stained with reticulin/silver shows normal reticulin deposition (·40). transplantation and cranial and spinal radiation therapy to treat possible residual tumor in sanctuary sites, and imatinib was replaced with a second-generation tyrosine kinase inhibitor, dasatinib. Extramedullary relapse presenting as CNS blast crisis with isolated blindness from bilateral optic nerve infiltration is extremely rare, occurring more frequently in children with acute lymphoblastic leukemia than in adults with chronic myeloid leukemia (1). Radiotherapy has been the treatment modality of choice for the more common blast crises of acute leukemia with some reports of dramatic recovery (12,13). However, extrapolation from protocols developed for acute leukemia might not be appropriate for CML since acute forms, although rapidly progressive, often can be cured. Chronic leukemia tends to persist, rarely achieving full cure, requiring maintenance therapy for containment (4). Because CML accounts for only 15% of newly diagnosed leukemia in adults (4), an evidence-based protocol for CNS blast crises with optic nerve infiltration has yet to be established. Visual recovery is variable in reported cases when treated with the common protocol of intrathecal cytosine arabinoside or cytarabine, methotrexate, and a corticosteroid, coupled with external beam radiation (Table 1) (2,3,14-18). Our patient demonstrated the very rare manifestation of CML relapse with CNS blast crisis and bilateral optic nerve infiltration after proven cytogenetic response or MMR and while on the current first-line therapy, imatinib. Once managed with systemic and intrathecal chemotherapy, imatinib was replaced on a second-generation tyrosine kinase inhibitor, dasatinib, which has been shown to have greater potency and better CNS penetration (19-21). Although FIG. 4. Octopus automated visual fields reveal mild visual field loss, more prominent in the left eye. OD, right eye; OS, left eye. Mbekeani et al: J Neuro-Ophthalmol 2016; 36: 73-77 75 Copyright © North American Neuro-Ophthalmology Society. Unauthorized reproduction of this article is prohibited. Report/Reference Presentation Confirmatory Tests Initial VA CBC: [ WBC; LP: [ICP and blast cells; Not recorded BM: CML; autopsy: optic nerve head myeloid blast cells; also found at other ocular sites *Ellis and Little (14) Fever of unknown origin and abdominal pain; disc edema OU Hendrick et al. (15) Decreased vision/headaches; CBC: [WBC; LP: [ICP and blast cells; 20/30 OD; 20/200 OS disc edema and retinopathy OU BM: [myeloid blasts; No CT/MRI recorded Asymptomatic; mild disc CBC: [WBC (immature forms); BM: 20/20 OU edema OS immature leukocytes-CML; VEP: delayed P100 latency and Y frequency OS NLP OU MRI: normal optic nerves and Decreased vision; normal enhancement appearing optic discs / optic of VIIn and VIIIn (leptomeninges); LP: atrophy lymphoblasts (previously diagnosed CML) 20/60 OD; Decreased vision; disc edema MRI: enhancing left optic nerve, VIIn 20/200 OS and retinopathy OU and VIIIn lesions; CBC: [WBC; BM: [granulocytes-CML MRI: enhancing left optic nerve; LP: Light perception Decreased vision, pain, and myeloid blast cells OS exophthalmos OS; headaches and vomiting; disc edema OS Not recorded "Visual impairment" headaches, MRI: normal brain and optic nerves; CBC: [WBC; LP: blast cells nausea/vomiting, confusion, seizures, and nuchal rigidity; optic atrophy CF OD; light Decreased vision and headaches MRI: chiasm and bilateral optic nerve enhancement; LP: myeloid blast cells; perception OS BM: normal trilineage (treated CML) *Costagliola et al. (16) †Schocket et al. (2) *Mandic et al. (3) Mbekeani et al: J Neuro-Ophthalmol 2016; 36: 73-77 †Yamamoto et al. (17) †Gomez and Duenas (18) †Mbekeani et al (this case) Treatment Final VA IV daunomycin X2 cycles, PO busulfan, and I/T methotrexate 20/25 (+2.00) OU; (died with persistent pale papilledema, retinal edema, and hemorrhages) IV doxorubicin, 6-thioguanine, NLP OD; 20/200 OS (died with vitreous I/T cytosine arabinoside hemorrhages OU) methotrexate, and DXT Induction not documented Not documented I/T cytarabine, IV solumedrol, NLP OU and DXT Induction hydroxyurea and interferon 20/20 OD; 20/60 OS (premacular hemorrhage) I/T cytarabine, methotrexate, Light perception OS dexamethasone, and focal DXT No VA recovery I/T cytosine arabinoside, methotrexate, and dexamethasone I/T cytarabine, methotrexate, 20/20 OD; 20/25 OS and hydrocortisone; IV fludarabine and cytarabine *Initial manifestation of CML. † Patients on imatinib therapy. CML, chronic myelogenus leukemia; VA, visual acuity; BM, bone marrow examination; CBC complete blood count; CF, counting fingers; DXT, external beam radiation; I/T, intrathecal; ICP, intracranial pressure; LP, lumbar puncture; MRI, magnetic resonance imaging; NLP, no light perception; OD, right eye; OS, left eye. Copyright © North American Neuro-Ophthalmology Society. Unauthorized reproduction of this article is prohibited. Clinical Observation 76 TABLE 1. Visual recovery in reported cases of chronic myelogenous leukemia with optic nerve involvement Clinical Observation a dramatic response was achieved in our patient, with no further CSF blasts, we acknowledge that systemic and intrathecal chemotherapy might not be enough to eradicate all leukemic cells in the perineural space and other sanctuaries, even with institution of dasatinib. Thus, our patient had subsequent allogeneic bone marrow transplantation, a potentially curative intervention, and spinal and cranial radiation therapy. Cases of optic disc edema associated with imatinib therapy also have been reported (22,23). There was temporal association with the start of therapy with resolution on cessation. Disc edema also was documented within two months of starting therapy. Our patient had been on imatinib without incident for 1 year and demonstrated a significant MRI infiltration and CSF confirmation of CNS blast crisis to account for bilateral disc edema. She achieved dramatic recovery of vision after systemic and intrathecal chemotherapy before radiotherapy. Despite 3 weeks of severe visual disability from optic nerve dysfunction, no permanent structural damage was demonstrable by optical coherence tomography. This finding is encouraging for our patient in terms of her long-term visual prognosis. STATEMENT OF AUTHORSHIP Category 1: a. Conception and design: Joyce N Mbekeani and Maaly AbdelFattah; b. Acquisition of data: Joyce N Mbekeani, Maaly Abdel Fattah, Wahiba Chebbo, Randa Al Nounou, and Mohammed Asif Dogar; c. Analysis and interpretation of data: Joyce N Mbekeani, Wahibo Chebbo, Randa Al Nounou, and Mohammed Asif Dogar. Category 2: a. Drafting the manuscript: Joyce N Mbekeani, Maaly Abdel Fattah, Wahibo Chebbo, Randa Al Nounou, and Mohammed Asif Dogar; b. Revising it for intellectual content: Joyce N Mbekeani, Randa Al Nounou, and Mohammed Asif Dogar. Category 3: a. Final approval of the completed manuscript: Joyce N Mbekeani, Maaly Abdel Fattah, Wahibo Chebbo, Randa Al Nounou, and Mohammed Asif Dogar. ACKNOWLEDGMENTS Naeem A. Chaudhri, MD and Ahmad Saleh Al Otaibi, MD: Dept of Hematologic Oncology, King Faisal Specialist Hospital &Research Center, Riyadh, KSA. Hassan Omairah, AAS, COT, OCT-C, CRA, Senior Ophthalmic Photographer: Ophthalmic Photography, Dept of Ophthalmology, King Faisal Specialist Hospital &Research Center, Riyadh, KSA. REFERENCES 1. Kincaid MC, Green WR. Ocular and orbital involvement in leukemia. Surv Ophthalmol. 1983;27:211-232. 2. Schocket LS, Massaro-Giordano M, Volpe NJ, Galetta SL. Bilateral optic nerve infiltration in central nervous system leukemia. Am J Ophthalmol. 2003;135:94-99. 3. Mandic BD, Potocnjak V, Bencic G, Mandic Z, Pantz A. Visual loss as initial presentation of chronic myelogenous leukemia. Coll Antropol. 2005;29(suppl 1):141-143. 4. Jabbour E, Kantarjan H. Chronic myeloid leukemia: 2014 update on diagnosis, monitoring and management. Am J Hematol. 2014;89:548-556. Mbekeani et al: J Neuro-Ophthalmol 2016; 36: 73-77 5. Takayama N, Sato N, O'Brien SG, Ikeda Y, Okamoto S. Imatinib mesylate has limited activity against the central nervous system involvement of Philadelphia chromosome-positive acute lymphoblastic leukaemia due to poor penetration into cerebrospinal fluid. Br J Haematol. 2002;119:106-108. 6. Wolff NC, Richardson JA, Egorin M, Illaria RL. The CNS is a sanctuary for leukemic cells in mice receiving imatinib mesylate for Bcr/Abl-induced leukemia. Blood. 2003;101:5010-5013. 7. Deininger MJ. Central nervous system blast crisis in chronic myeloid leukemia on imatinib mesylate therapy: report of two cases. J Natl Compr Canc Netw. 2008;6(suppl 2):S11-S21. 8. Radhika N, Minakshi M, Rajesh M, Manas BR, Kumar MD. Resistance and relapse with imatinib in CML: causes and consequences. Indian J Hematol Blood Transfus. 2011;27:51-54. 9. Rajappa S, Uppin SG, Raghunadharao D, Rao IS, Surath A. Isolated central nervous system blast crisis in chronic myeloid leukemia. Hematol Oncol. 2004;22:179-181. 10. Altintas A, Cil T, Kilinc I, Kaplan MA, Ayyildiz O. Central nervous system blastic crisis in chronic myeloid leukemia on imatinib mesylate therapy: a case report. J Neurooncol. 2007;84:103-105. 11. Rytting ME, Wierda WG. Central nervous system relapse in two patients with chronic myelogenous leukemia in myeloid blastic phase on imatinib mesylate therapy. Leuk Lymphoma. 2004;45:1623-1626. 12. Puvanachandra N, Goddard K, Lyons CJ. Dramatic visual recovery after prompt radiotherapy and chemotherapy for leukaemic infiltration of the optic nerve in a child. Eye (Lond). 2010;24:927-928. 13. Lin YC, Wang AG, Yen MY, Hsu WM. Leukemic infiltration of the optic nerve. Eye (Lond). 2004;18:546-550. 14. Ellis W, Little HL. Leukemic infiltration of the optic nerve head. Am J Ophthalmol. 1973;75:867-871. 15. Hendrick AM, Rogers JS II, Guliner RJ. Ocular complications following blast transformation in chronic myelogenous leukemia. Am J Hematol. 1979;7:389-394. 16. Costagliola C, Rinaldi M, Cotticelli L, Sbordone S, Nastri G. Isolated optic nerve involvement in chronic myeloid leukemia. Leuk Res. 1992;16:411-413. 17. Yamamoto H, Yamamoto M, Omoto E, Kato T, Tajima K. An isolated myeloid blast crisis presenting as optic nerve sarcoma in a patient with chronic myeloid leukemia treated with imatinib. Br J Haematol. 2015;170:290. 18. Gomez J, Duenas V. Isolated central nervous system relapse in chronic myeloid leukemia. Case Rep Med. 2015;2015:232915. 19. Shah NP, Guilhot F, Cortes JE, Schiffer CA, le Coutre P, Brümmendorf TH, Kantarjian HM, Hochhaus A, Rousselot P, Mohamed H, Healey D, Cunnningham M, Saglio G. Long-term outcome with dasatinib after imatinib failure in chronic-phase chronic myeloid leukemia: follow-up of phase 3 study. Blood. 2014;123:2317-2324. 20. Kantarjian H, Shah NP, Hochhaus A, Cortes J, Shah S, Ayala M, Moiraghi B, Shen Z, Mayer J, Pasquini R, Nakamae H, Huguet F, Boqué C, Chuah C, Bleickardt E, Bradley-Garelik MB, Zhu C, Szatrowski T, Shapiro D, Baccarani M. Dasatinib versus imatinib in newly diagnosed chronic-phase chronic myeloid leukemia. N Engl J Med. 2010;362:2260-2270. 21. Jabbour E, Kantarjian HM, Saglio G, Steegmann JL, Shah NP, Boquë C, Chuah C, Pavlovsky C, Mayer J, Cortes J, Baccarani M, Kim DW, Bradley-Garelik MB, Mohamed H, Wildgust M, Hochhaus A. Early response with dasatinib or imatinib in chronic myeloid leukemia: 3-year follow-up from a randomized phase 3 trial (DASISION). Blood. 2014;123:494-500. 22. Kwon SI, Lee DH, Kim YJ. Optic disc edema as a possible complication of imatinib mesylate (Gleevec). Jpn J Ophthalmol. 2008;52:331-333. 23. DeLuca C, Shenouda-Awad N, Haskes C, Wrzesinski S. Imatinib mesylate (Gleevec) induced unilateral optic disc edema. Optom Vis Sci. 2012;89:e16-e22. 77 Copyright © North American Neuro-Ophthalmology Society. Unauthorized reproduction of this article is prohibited.