- Email: [email protected]
Neurofibromatosis type 2 and multiple sclerosis
Journal Pre-proof
Neurofibromatosis type 2 and multiple sclerosis Dario-Lucas Helbing , Michael Brodhun MD , Oliver Tiedge MD , Helen Morrison PhD , Steffen K. Rosahl MD PII: DOI: Reference:
S2211-0348(19)30961-7 https://doi.org/10.1016/j.msard.2019.101890 MSARD 101890
To appear in:
Multiple Sclerosis and Related Disorders
Received date: Revised date: Accepted date:
15 January 2019 27 October 2019 8 December 2019
Please cite this article as: Dario-Lucas Helbing , Michael Brodhun MD , Oliver Tiedge MD , Helen Morrison PhD , Steffen K. Rosahl MD , Neurofibromatosis type 2 and multiple sclerosis, Multiple Sclerosis and Related Disorders (2019), doi: https://doi.org/10.1016/j.msard.2019.101890
This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier B.V.
Highlights
Multiple sclerosis can occur in patients with neurofibromatosis type 2. Peripheral schwannoma of a patient with both diseases showed a massive tcell infiltration. This t-cell infiltration correlated with a very rapid tumor growth.
Neurofibromatosis type 2 and multiple sclerosis
Dario-Lucas Helbing1,2, Michael Brodhun, MD3, Oliver Tiedge4, MD, Helen Morrison, PhD2, Steffen K. Rosahl, MD1
1
Department of Neurosurgery, Helios Klinikum, 99089 Erfurt, Germany
2
Leibniz Institute for Age Research, Fritz Lipmann Institute, 07745 Jena, Germany
3
Department of Pathology, Helios Klinikum, 99089 Erfurt, Germany
4
Department of Neurology, St.Georg Klinikum and MVZ, 99817 Eisenach, Germany
Corresponding author: Steffen K. Rosahl, MD Department of Neurosurgery HELIOS Klinikum, Nordhaeuser Str. 74, D-99089 Erfurt, Germany E-mail: [email protected] Number of figures: 3
tables:
0 Suppl. figures: 0
Number of characters in title: 62 Number of Characters in running head: 25 Number of words in abstract: 105 Total number of pages: 9 Short title: Nf2 and MS: A case report
Key words: macrophages; T-cells; neurofibromatosis type 2; schwannoma; multiple sclerosis Abbreviations MRI= Magnetic resonance imaging MS= Multiple sclerosis NF1= Neurofibromatosis type 1 NF2= Neurofibromatosis type 2 OMG= Oligodendrocyte myelin glycoprotein VS= Vestibular schwannoma
Abstract Comorbidity of neurofibromatosis type 2 (NF2) and multiple sclerosis (MS) has rarely been reported. Since immunological mechanisms have been implicated in Nf2, coexistence of the two entities may offer insights into schwannoma pathogenesis with respect to the impact of the immune system. We present the case of a woman with a de novo mutation in the NF2 gene who later developed MS. In addition, we found a significantly higher count of T cells in a laryngeal schwannoma of this patient as compared to a schwannoma removed from a NF2 patient without MS. This finding correlated with a higher growth rate in the case of NF+MS.
Introduction
Neurofibromatosis type 2 is an autosomal dominant inherited neurocutaneous disorder which is characterized by the development of multiple tumors in the nervous system. The disease hits approximately 1 in 25.000 livebirths with a wide phenotypic variability1. Comorbidity of NF2 and multiple sclerosis was only reported in one case in the literature until now2. An association between Neurofibromatosis type 1 and the development of MS has been previously described3. Patients with large deletions in the NF1 gene have a loss of a common set of 11 functional genes, including the loss of the OMG gene, encoding the oligodendrocyte myelin glycoprotein which lies in an intron of the NF1 gene4. However, this specific mutation is not a prerequisite to develop MS5, therefore a genetic association between these two diseases can be excluded. Further theories whether nerve sheath tumors or neurofibromas in NF1 or NF2 predispose for multiple sclerosis lack evidence6. We present a case of a woman with NF2 who developed relapsing remitting multiple sclerosis.
Case Report and Results
The patient (NF+MS) is a 45-year-old woman with diagnosed mosaic NF2 who first presented in our clinic with bilateral tinnitus in 2000. Due to the presence of bilateral vestibular schwannoma (Fig.1C) and a frontal falx meningioma she was diagnosed with neurofibromatosis type 2 (Fig.1A). The woman (Subject 6, Fig.1A) has no obvious family history of NF2 or MS despite an aunt with an unclear nerve disease (Subject 4, Fig.1A). Sequencing of lymphocyte extracted DNA revealed a mutation in exon 15 leading to a frameshift in protein translation, resulting in a non-functional protein (Fig.1B). This specific mutation of the bases 1614-1615 has not been reported previously.
Figure 1 (A) Family History Tree: Black arrow marks NF+MS. (B) Schematic alignment of DNA sequence of wildtype merlin and mutated merlin of NF+MS with translation into amino acid sequence. synthesis due to the mutation.
*
indicates termination of protein
(C) T1 weighted MRI image shows typical NF2-related bilateral schwannoma at two different time points and of two different transversal slices (upper and lower row). Note the decrease in size of the non-operated schwannoma on the left side (for quantification see Fig.3).
The patient presented with right hypoesthesia in V1-V3, disequilibrium and a tendency to fall with her eyes closed. In another institution, she underwent bilateral stereotactic radiotherapy of both vestibular schwannoma (VS) and of the frontal meningioma (total dose of 40 Gy in 10 fractions) from 01/2004 to 03/2004. Due to the growth of the right VS between 2004 and 11/2006 with subsequent abducens paresis and brain stem compression, the tumor was removed microsurgically. Pathological assessment confirmed the presence of a schwannoma (WHO grade 1). In 11/2007 the patient presented with nightly paresthesia in the supplying area of both ulnar nerves. A positive Tinel-Hoffmann-Sign suggested local nerve irritation. In 05/2008 a substantially growing intramedullary schwannoma at L2 was removed. In September 2008 hypoesthesia in the right L4 and L5 dermatomes and paresthesia in the right foot developed. An intradural tumor at L2/3 with a diameter of 1.2 cm and a small tumor at L4/5 remained stable on several follow-up scans. In 03/2010 progressive paresthesia in both legs occurred. A year later, the patient reported difficulties in swallowing. The MRI showed a tumor near the trachea. This schwannoma with esophageal infiltration was removed in April 2011. Later, the patient also presented with hypoesthesia below the armpits, appearing only after she exercised. In retrospect, this was classified as Uhthoff’s phenomenon, a classical sign of MS. Over the course of time she developed numbness and hypoesthesia in the right digits III-V in October/November of 2011. This condition became steadily worse until July 2013 when she reported that the numbness had now spread involving her whole right hand. Due to the presence of a new periventricular T2-weighted MRI hyperintensity (Fig.2), we decided to include further diagnostic procedures to test for MS in her regular check-up in 2014. The suspicion was confirmed by the presence of specific oligoclonal bands in the cerebrospinal fluid and by proven intrathecal IgG-synthesis. Alongside multiple hyperintense lesions in T2-weighted MRIs and at least two clinical relapses, the diagnosis of relapsingremitting multiple sclerosis was established according to the McDonald criteria7, 8. Accordingly, the patient was treated with 125 µg intramuscular peginterferon beta1a injections every two weeks.
Figure 2 T2 weighted MRI image showing multiple MS lesions during course of the disease and in different transversal sections (vertical panels) as well as a frontal falx meningioma.
In 11/2016, there was still a peripheral facial paralysis on the right side, right hypoesthesia in V1-V3, complete sensory hearing loss on the right ear and alternating but preserved hearing on the left side. Intermittent paresthesia in both legs and disequilibrium with a tendency to fall also prevailed, and she reported paresthesia in the left first three digits. A meningioma colliding with her vestibular schwannoma on the right side had grown significantly and threatened to affect hearing in her only remaining ear. The meningioma was removed, and hearing remained stable. At present, her therapy still consists of 125 µg peginterferon beta-1a i.m. every two weeks. Histopathologic workup and tumor volumetry To evaluate the possible influence of the MS on tumor growth and tumor-related immune system tissue inflammation, we performed neuropathological assessment of the laryngeal schwannoma and compared macrophage inflammation and T cell infiltration to a hypoglossal schwannoma from a control patient (NF) who suffered from NF2 without MS. Both were neuropathologically diagnosed as WHO grade I schwannoma. To rule out staining bias due to tumor hemorrhage Berlin blue staining was performed which showed no hemorrhage and siderophages. The tumor of the NF+MS patient showed a higher cellularity in all sections (Fig.3A). It is reported for schwannoma that the tumor cells express the classical macrophage and lysosomal marker CD689, which seemed to be expressed on a higher level in our NF+MS patient. CD68 staining was diffuse and extended to tumor cells. We decided to perform further staining with another macrophage marker, MAC387, in order to quantify the number of infiltrating macrophages10. We could confirm the previously reported finding of a massive macrophage infiltration into the schwannomas11 in both patients, although the tumor from our Pat.NF+MS displayed less infiltrating macrophages compared to the tissue of the control patient. Interestingly, we observed T cell infiltration in both schwannoma and also a massively higher number of T cells in the tumor of NF+MS (Fig.3A). This correlated with a higher growth rate of the hypopharyngeal schwannoma of NF+MS (97% volume increase in one year) compared to the hypoglossal schwannoma of NF (29% in 3 years and 46% in 2 years) (Fig.3B). The volume increase of 97% of the hypopharyngeal schwannoma of NF+MS during one year only by far exceeds the median increase of non-vestibular schwannomas in NF2 patients that has recently been reported to be 25,6% ± 12,98% per year12.
Figure 3 (A) Neuropathological evaluation of tumor tissue with hematoxylin-eosin staining and CD3, MAC387 and CD68 markers. Scale bars: HE Stainings: 100 µm; Pat.NF: CD68, MAC387: 100 µm, CD3: 200 µm; Pat. NF+MS: CD68: 100 µm, MAC387, CD3: 200 µm. Representative areas have been selected from images taken by automatic virtual microscopy with an Axio Scan.Z1 device. (B) Magnetic resonance tumor volumetry. Data set shows tumor growth curves of both neuropathologically assessed peripheral tumors and the vestibular schwannomas of both patients. Volume change in % compared to previous tumor volume is depicted for the peripheral schwannomas.
Discussion Consistent with recent findings about the influence of the microenvironment on the pathogenesis of NF2-associated schwannoma, these tumors often arise at confined spaces in the body e.g. the internal acoustic meatus9. It is hypothesized that this mechanical factor induces neural microlesions . In consequence of a lesion, the NF2deficient tissue has a regenerative deficit, which in turn results in the development of tumors usually originating at these exact locations10. The regeneration of microlesions of the nerve is controlled by a variety of different cells, especially cells of the immune system which give proliferation and differentiation signals like cytokines and growth factors to Schwann cells and axons10. In multiple sclerosis this interplay and crosstalk between immune cells, neurons and glial cells is dysregulated, which results in the infiltration of immune cells and the subsequent demyelination and death of axons 13. Whether MS leads to a higher risk of developing tumors, possibly due to a dysregulated immune system, is still under debate14. T-cell infiltration into the tumor tissue was elevated in our NF2+MS patient as compared to a non-MS NF2 case. However, whether the observed higher T-cell number can explain the observed faster tumor growth in comparison to the median tumor growth rate of vestibular schwannoma in NF2 patients in general, remains speculative. To explore a possible causal or mediative relationship of reactive T-cells in vestibular schwannoma growth in NF2 patients a larger number of patients with both, MS and NF2 need to be identified and analyzed.
Conclusion This case features a very rare comorbidity of NF2 and MS. We found a considerably higher number of tumor-infiltrating T-cells and a by far higher growth rate in a peripheral schwannoma of this female patient compared to a peripheral schwannoma of another woman suffering from NF2 only. Even though the growth rate of the NF2 + MS schwannoma was abnormally high compared to the growth rate, found in most other NF2 patients it would be premature to draw conclusions about a possible causal or modulating role of T-cells in NF2-associated schwannoma growth. More patients with the comorbidity of MS and NF2 will need to be identified and their tumors will have to be compared to other NF2 patients.
Authors Contributions SR examined and operated patients. DLH wrote the manuscript, initiated experiments, performed tumor volumetry and participated in neuropathological analysis. MB performed neuropathological assessment. OT examined and treated the presented patient and performed immunotherapy. DLH, SR and HM analyzed and discussed results, edited and corrected the manuscript. Conflict of Interest The authors have no conflict of interest to declare. Informed Consent Written informed consent was obtained from all patients. Acknowledgements The authors would like to thank Birgit Perner for help in setting up an imaging protocol for the Axio Scan.Z1 device and Leonie Karoline Stabenow for critical reading and editing of the manuscript.
References 1. 2.
3. 4. 5.
6. 7. 8. 9.
10.
11. 12. 13. 14.
Asthagiri AR, Parry DM, Butman JA, et al. Neurofibromatosis type 2. Lancet (London, England). Jun 6 2009;373(9679):1974-1986. Naghshineh H, Shahin D, Sahraian MA, Minagar A. Co-existence of neurofibromatosis type 2 and multiple sclerosis: A case report. Multiple sclerosis and related disorders. May 2014;3(3):384-386. Etemadifar M, Fatehi F, Sahraian M, et al. Multiple sclerosis and neurofibromatosis type 1: report of seven patients from Iran. Multiple Sclerosis Journal. 2009;15(9):1126-1130. Jenne DE, Tinschert S, Stegmann E, et al. A common set of at least 11 functional genes is lost in the majority of NF1 patients with gross deletions. Genomics. May 15 2000;66(1):93-97. Johnson MR, Ferner RE, Bobrow M, Hughes RAC. Detailed analysis of the oligodendrocyte myelin glycoprotein gene in four patients with neurofibromatosis 1 and primary progressive multiple sclerosis. Journal of Neurology, Neurosurgery & Psychiatry. 2000;68(5):643646. Perini P, Gallo P. The range of multiple sclerosis associated with neurofibromatosis type 1. Journal of Neurology, Neurosurgery & Psychiatry. 2001;71(5):679-681. Polman CH, Reingold SC, Banwell B, et al. Diagnostic criteria for multiple sclerosis: 2010 revisions to the McDonald criteria. Ann Neurol. Feb 2011;69(2):292-302. Thompson AJ, Banwell BL, Barkhof F, et al. Diagnosis of multiple sclerosis: 2017 revisions of the McDonald criteria. The Lancet Neurology. 2018;17(2):162-173. Kurtin PJ, Bonin DM. Immunohistochemical demonstration of the lysosome-associated glycoprotein CD68 (KP-1) in granular cell tumors and schwannomas. Human pathology. Nov 1994;25(11):1172-1178. Goebeler M, Roth J, Teigelkamp S, Sorg C. The monoclonal antibody MAC387 detects an epitope on the calcium-binding protein MRP14. Journal of leukocyte biology. Feb 1994;55(2):259-261. Schulz A, Buttner R, Hagel C, et al. The importance of nerve microenvironment for schwannoma development. Acta neuropathologica. Aug 2016;132(2):289-307. Lawson McLean AC, Rosahl SK. Growth Dynamics of Intracranial Tumors in Patients with Neurofibromatosis Type 2. World Neurosurg. Feb 2017;98:152-161. Dendrou CA, Fugger L, Friese MA. Immunopathology of multiple sclerosis. Nature reviews. Immunology. Sep 15 2015;15(9):545-558. Sun LM, Lin CL, Chung CJ, Liang JA, Sung FC, Kao CH. Increased breast cancer risk for patients with multiple sclerosis: a nationwide population-based cohort study. European journal of neurology. Feb 2014;21(2):238-244.
Neurofibromatosis type 2 and multiple sclerosis Dario-Lucas Helbing , Michael Brodhun MD , Oliver Tiedge MD , Helen Morrison PhD , Steffen K. Rosahl MD PII: DOI: Reference:
S2211-0348(19)30961-7 https://doi.org/10.1016/j.msard.2019.101890 MSARD 101890
To appear in:
Multiple Sclerosis and Related Disorders
Received date: Revised date: Accepted date:
15 January 2019 27 October 2019 8 December 2019
Please cite this article as: Dario-Lucas Helbing , Michael Brodhun MD , Oliver Tiedge MD , Helen Morrison PhD , Steffen K. Rosahl MD , Neurofibromatosis type 2 and multiple sclerosis, Multiple Sclerosis and Related Disorders (2019), doi: https://doi.org/10.1016/j.msard.2019.101890
This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier B.V.
Highlights
Multiple sclerosis can occur in patients with neurofibromatosis type 2. Peripheral schwannoma of a patient with both diseases showed a massive tcell infiltration. This t-cell infiltration correlated with a very rapid tumor growth.
Neurofibromatosis type 2 and multiple sclerosis
Dario-Lucas Helbing1,2, Michael Brodhun, MD3, Oliver Tiedge4, MD, Helen Morrison, PhD2, Steffen K. Rosahl, MD1
1
Department of Neurosurgery, Helios Klinikum, 99089 Erfurt, Germany
2
Leibniz Institute for Age Research, Fritz Lipmann Institute, 07745 Jena, Germany
3
Department of Pathology, Helios Klinikum, 99089 Erfurt, Germany
4
Department of Neurology, St.Georg Klinikum and MVZ, 99817 Eisenach, Germany
Corresponding author: Steffen K. Rosahl, MD Department of Neurosurgery HELIOS Klinikum, Nordhaeuser Str. 74, D-99089 Erfurt, Germany E-mail: [email protected] Number of figures: 3
tables:
0 Suppl. figures: 0
Number of characters in title: 62 Number of Characters in running head: 25 Number of words in abstract: 105 Total number of pages: 9 Short title: Nf2 and MS: A case report
Key words: macrophages; T-cells; neurofibromatosis type 2; schwannoma; multiple sclerosis Abbreviations MRI= Magnetic resonance imaging MS= Multiple sclerosis NF1= Neurofibromatosis type 1 NF2= Neurofibromatosis type 2 OMG= Oligodendrocyte myelin glycoprotein VS= Vestibular schwannoma
Abstract Comorbidity of neurofibromatosis type 2 (NF2) and multiple sclerosis (MS) has rarely been reported. Since immunological mechanisms have been implicated in Nf2, coexistence of the two entities may offer insights into schwannoma pathogenesis with respect to the impact of the immune system. We present the case of a woman with a de novo mutation in the NF2 gene who later developed MS. In addition, we found a significantly higher count of T cells in a laryngeal schwannoma of this patient as compared to a schwannoma removed from a NF2 patient without MS. This finding correlated with a higher growth rate in the case of NF+MS.
Introduction
Neurofibromatosis type 2 is an autosomal dominant inherited neurocutaneous disorder which is characterized by the development of multiple tumors in the nervous system. The disease hits approximately 1 in 25.000 livebirths with a wide phenotypic variability1. Comorbidity of NF2 and multiple sclerosis was only reported in one case in the literature until now2. An association between Neurofibromatosis type 1 and the development of MS has been previously described3. Patients with large deletions in the NF1 gene have a loss of a common set of 11 functional genes, including the loss of the OMG gene, encoding the oligodendrocyte myelin glycoprotein which lies in an intron of the NF1 gene4. However, this specific mutation is not a prerequisite to develop MS5, therefore a genetic association between these two diseases can be excluded. Further theories whether nerve sheath tumors or neurofibromas in NF1 or NF2 predispose for multiple sclerosis lack evidence6. We present a case of a woman with NF2 who developed relapsing remitting multiple sclerosis.
Case Report and Results
The patient (NF+MS) is a 45-year-old woman with diagnosed mosaic NF2 who first presented in our clinic with bilateral tinnitus in 2000. Due to the presence of bilateral vestibular schwannoma (Fig.1C) and a frontal falx meningioma she was diagnosed with neurofibromatosis type 2 (Fig.1A). The woman (Subject 6, Fig.1A) has no obvious family history of NF2 or MS despite an aunt with an unclear nerve disease (Subject 4, Fig.1A). Sequencing of lymphocyte extracted DNA revealed a mutation in exon 15 leading to a frameshift in protein translation, resulting in a non-functional protein (Fig.1B). This specific mutation of the bases 1614-1615 has not been reported previously.
Figure 1 (A) Family History Tree: Black arrow marks NF+MS. (B) Schematic alignment of DNA sequence of wildtype merlin and mutated merlin of NF+MS with translation into amino acid sequence. synthesis due to the mutation.
*
indicates termination of protein
(C) T1 weighted MRI image shows typical NF2-related bilateral schwannoma at two different time points and of two different transversal slices (upper and lower row). Note the decrease in size of the non-operated schwannoma on the left side (for quantification see Fig.3).
The patient presented with right hypoesthesia in V1-V3, disequilibrium and a tendency to fall with her eyes closed. In another institution, she underwent bilateral stereotactic radiotherapy of both vestibular schwannoma (VS) and of the frontal meningioma (total dose of 40 Gy in 10 fractions) from 01/2004 to 03/2004. Due to the growth of the right VS between 2004 and 11/2006 with subsequent abducens paresis and brain stem compression, the tumor was removed microsurgically. Pathological assessment confirmed the presence of a schwannoma (WHO grade 1). In 11/2007 the patient presented with nightly paresthesia in the supplying area of both ulnar nerves. A positive Tinel-Hoffmann-Sign suggested local nerve irritation. In 05/2008 a substantially growing intramedullary schwannoma at L2 was removed. In September 2008 hypoesthesia in the right L4 and L5 dermatomes and paresthesia in the right foot developed. An intradural tumor at L2/3 with a diameter of 1.2 cm and a small tumor at L4/5 remained stable on several follow-up scans. In 03/2010 progressive paresthesia in both legs occurred. A year later, the patient reported difficulties in swallowing. The MRI showed a tumor near the trachea. This schwannoma with esophageal infiltration was removed in April 2011. Later, the patient also presented with hypoesthesia below the armpits, appearing only after she exercised. In retrospect, this was classified as Uhthoff’s phenomenon, a classical sign of MS. Over the course of time she developed numbness and hypoesthesia in the right digits III-V in October/November of 2011. This condition became steadily worse until July 2013 when she reported that the numbness had now spread involving her whole right hand. Due to the presence of a new periventricular T2-weighted MRI hyperintensity (Fig.2), we decided to include further diagnostic procedures to test for MS in her regular check-up in 2014. The suspicion was confirmed by the presence of specific oligoclonal bands in the cerebrospinal fluid and by proven intrathecal IgG-synthesis. Alongside multiple hyperintense lesions in T2-weighted MRIs and at least two clinical relapses, the diagnosis of relapsingremitting multiple sclerosis was established according to the McDonald criteria7, 8. Accordingly, the patient was treated with 125 µg intramuscular peginterferon beta1a injections every two weeks.
Figure 2 T2 weighted MRI image showing multiple MS lesions during course of the disease and in different transversal sections (vertical panels) as well as a frontal falx meningioma.
In 11/2016, there was still a peripheral facial paralysis on the right side, right hypoesthesia in V1-V3, complete sensory hearing loss on the right ear and alternating but preserved hearing on the left side. Intermittent paresthesia in both legs and disequilibrium with a tendency to fall also prevailed, and she reported paresthesia in the left first three digits. A meningioma colliding with her vestibular schwannoma on the right side had grown significantly and threatened to affect hearing in her only remaining ear. The meningioma was removed, and hearing remained stable. At present, her therapy still consists of 125 µg peginterferon beta-1a i.m. every two weeks. Histopathologic workup and tumor volumetry To evaluate the possible influence of the MS on tumor growth and tumor-related immune system tissue inflammation, we performed neuropathological assessment of the laryngeal schwannoma and compared macrophage inflammation and T cell infiltration to a hypoglossal schwannoma from a control patient (NF) who suffered from NF2 without MS. Both were neuropathologically diagnosed as WHO grade I schwannoma. To rule out staining bias due to tumor hemorrhage Berlin blue staining was performed which showed no hemorrhage and siderophages. The tumor of the NF+MS patient showed a higher cellularity in all sections (Fig.3A). It is reported for schwannoma that the tumor cells express the classical macrophage and lysosomal marker CD689, which seemed to be expressed on a higher level in our NF+MS patient. CD68 staining was diffuse and extended to tumor cells. We decided to perform further staining with another macrophage marker, MAC387, in order to quantify the number of infiltrating macrophages10. We could confirm the previously reported finding of a massive macrophage infiltration into the schwannomas11 in both patients, although the tumor from our Pat.NF+MS displayed less infiltrating macrophages compared to the tissue of the control patient. Interestingly, we observed T cell infiltration in both schwannoma and also a massively higher number of T cells in the tumor of NF+MS (Fig.3A). This correlated with a higher growth rate of the hypopharyngeal schwannoma of NF+MS (97% volume increase in one year) compared to the hypoglossal schwannoma of NF (29% in 3 years and 46% in 2 years) (Fig.3B). The volume increase of 97% of the hypopharyngeal schwannoma of NF+MS during one year only by far exceeds the median increase of non-vestibular schwannomas in NF2 patients that has recently been reported to be 25,6% ± 12,98% per year12.
Figure 3 (A) Neuropathological evaluation of tumor tissue with hematoxylin-eosin staining and CD3, MAC387 and CD68 markers. Scale bars: HE Stainings: 100 µm; Pat.NF: CD68, MAC387: 100 µm, CD3: 200 µm; Pat. NF+MS: CD68: 100 µm, MAC387, CD3: 200 µm. Representative areas have been selected from images taken by automatic virtual microscopy with an Axio Scan.Z1 device. (B) Magnetic resonance tumor volumetry. Data set shows tumor growth curves of both neuropathologically assessed peripheral tumors and the vestibular schwannomas of both patients. Volume change in % compared to previous tumor volume is depicted for the peripheral schwannomas.
Discussion Consistent with recent findings about the influence of the microenvironment on the pathogenesis of NF2-associated schwannoma, these tumors often arise at confined spaces in the body e.g. the internal acoustic meatus9. It is hypothesized that this mechanical factor induces neural microlesions . In consequence of a lesion, the NF2deficient tissue has a regenerative deficit, which in turn results in the development of tumors usually originating at these exact locations10. The regeneration of microlesions of the nerve is controlled by a variety of different cells, especially cells of the immune system which give proliferation and differentiation signals like cytokines and growth factors to Schwann cells and axons10. In multiple sclerosis this interplay and crosstalk between immune cells, neurons and glial cells is dysregulated, which results in the infiltration of immune cells and the subsequent demyelination and death of axons 13. Whether MS leads to a higher risk of developing tumors, possibly due to a dysregulated immune system, is still under debate14. T-cell infiltration into the tumor tissue was elevated in our NF2+MS patient as compared to a non-MS NF2 case. However, whether the observed higher T-cell number can explain the observed faster tumor growth in comparison to the median tumor growth rate of vestibular schwannoma in NF2 patients in general, remains speculative. To explore a possible causal or mediative relationship of reactive T-cells in vestibular schwannoma growth in NF2 patients a larger number of patients with both, MS and NF2 need to be identified and analyzed.
Conclusion This case features a very rare comorbidity of NF2 and MS. We found a considerably higher number of tumor-infiltrating T-cells and a by far higher growth rate in a peripheral schwannoma of this female patient compared to a peripheral schwannoma of another woman suffering from NF2 only. Even though the growth rate of the NF2 + MS schwannoma was abnormally high compared to the growth rate, found in most other NF2 patients it would be premature to draw conclusions about a possible causal or modulating role of T-cells in NF2-associated schwannoma growth. More patients with the comorbidity of MS and NF2 will need to be identified and their tumors will have to be compared to other NF2 patients.
Authors Contributions SR examined and operated patients. DLH wrote the manuscript, initiated experiments, performed tumor volumetry and participated in neuropathological analysis. MB performed neuropathological assessment. OT examined and treated the presented patient and performed immunotherapy. DLH, SR and HM analyzed and discussed results, edited and corrected the manuscript. Conflict of Interest The authors have no conflict of interest to declare. Informed Consent Written informed consent was obtained from all patients. Acknowledgements The authors would like to thank Birgit Perner for help in setting up an imaging protocol for the Axio Scan.Z1 device and Leonie Karoline Stabenow for critical reading and editing of the manuscript.
References 1. 2.
3. 4. 5.
6. 7. 8. 9.
10.
11. 12. 13. 14.
Asthagiri AR, Parry DM, Butman JA, et al. Neurofibromatosis type 2. Lancet (London, England). Jun 6 2009;373(9679):1974-1986. Naghshineh H, Shahin D, Sahraian MA, Minagar A. Co-existence of neurofibromatosis type 2 and multiple sclerosis: A case report. Multiple sclerosis and related disorders. May 2014;3(3):384-386. Etemadifar M, Fatehi F, Sahraian M, et al. Multiple sclerosis and neurofibromatosis type 1: report of seven patients from Iran. Multiple Sclerosis Journal. 2009;15(9):1126-1130. Jenne DE, Tinschert S, Stegmann E, et al. A common set of at least 11 functional genes is lost in the majority of NF1 patients with gross deletions. Genomics. May 15 2000;66(1):93-97. Johnson MR, Ferner RE, Bobrow M, Hughes RAC. Detailed analysis of the oligodendrocyte myelin glycoprotein gene in four patients with neurofibromatosis 1 and primary progressive multiple sclerosis. Journal of Neurology, Neurosurgery & Psychiatry. 2000;68(5):643646. Perini P, Gallo P. The range of multiple sclerosis associated with neurofibromatosis type 1. Journal of Neurology, Neurosurgery & Psychiatry. 2001;71(5):679-681. Polman CH, Reingold SC, Banwell B, et al. Diagnostic criteria for multiple sclerosis: 2010 revisions to the McDonald criteria. Ann Neurol. Feb 2011;69(2):292-302. Thompson AJ, Banwell BL, Barkhof F, et al. Diagnosis of multiple sclerosis: 2017 revisions of the McDonald criteria. The Lancet Neurology. 2018;17(2):162-173. Kurtin PJ, Bonin DM. Immunohistochemical demonstration of the lysosome-associated glycoprotein CD68 (KP-1) in granular cell tumors and schwannomas. Human pathology. Nov 1994;25(11):1172-1178. Goebeler M, Roth J, Teigelkamp S, Sorg C. The monoclonal antibody MAC387 detects an epitope on the calcium-binding protein MRP14. Journal of leukocyte biology. Feb 1994;55(2):259-261. Schulz A, Buttner R, Hagel C, et al. The importance of nerve microenvironment for schwannoma development. Acta neuropathologica. Aug 2016;132(2):289-307. Lawson McLean AC, Rosahl SK. Growth Dynamics of Intracranial Tumors in Patients with Neurofibromatosis Type 2. World Neurosurg. Feb 2017;98:152-161. Dendrou CA, Fugger L, Friese MA. Immunopathology of multiple sclerosis. Nature reviews. Immunology. Sep 15 2015;15(9):545-558. Sun LM, Lin CL, Chung CJ, Liang JA, Sung FC, Kao CH. Increased breast cancer risk for patients with multiple sclerosis: a nationwide population-based cohort study. European journal of neurology. Feb 2014;21(2):238-244.