A novel peroxisome-related gene signature predicts clinical prognosis and is associated with immune microenvironment in low-grade glioma.

Low-grade glioma (LGG), a common primary tumor, mainly originates from astrocytes and oligodendrocytes. Increasing evidence has shown that peroxisomes function in the regulation of tumorigenesis and development of cancer. However, the prognostic value of peroxisome-related genes (PRGs) in LGG has not been reported. Therefore, it is necessary to construct a prognostic risk model for LGG patients based on the expression profiles of peroxisome-related genes. Our study mainly concentrated on developing a peroxisome-related gene signature for overall survival (OS) prediction in LGG patients. First, according to these peroxisome-related genes, all LGG patients from The Cancer Genome Atlas (TCGA) database could be divided into two subtypes. Univariate Cox regression analysis was used to find prognostic peroxisome-related genes in TCGA_LGG dataset, and least absolute shrinkage and selection operator Cox regression analysis was employed to establish a 14-gene signature. The risk score based on the signature was positively associated with unfavorable prognosis. Then, multivariate Cox regression incorporating additional clinical characteristics showed that the 14-gene signature was an independent predictor of LGG. Time-dependent ROC curves revealed good performance of this prognostic signature in LGG patients. The performance about predicting OS of LGG was validated using the GSE107850 dataset derived from the Gene Expression Omnibus (GEO) database. Furethermore, we constructed a nomogram model based on the gene signature and age, which showed a better prognostic power. Gene ontology (GO) and Kyoto Encylopedia of Genes and Genomes (KEGG) analyses showed that neuroactive ligand-receptor interaction and phagosome were enriched and that the immune status was decreased in the high-risk group. Finally, cell counting kit-8 (CCK8) were used to detect cell proliferation of U251 and A172 cells. Inhibition of ATAD1 (ATPase family AAA domain-containing 1) and ACBD5 (Acyl-CoA binding-domain-containing-5) expression led to significant inhibition of U251 and A172 cell proliferation. Flow cytometry detection showed that ATAD1 and ACBD5 could induce apoptosis of U251 and A172 cells. Therefore, through bioinformatics methods and cell experiments, our study developed a new peroxisome-related gene signature that migh t help improve personalized OS prediction in LGG patients.

Pituitary adenoma with cavernous sinus compartment penetration and intracranial extension: surgical anatomy, approach, and outcomes.

Objectives: To understand the different characteristics and growth corridors of knosp grade 4 pituitary adenomas (Knosp4PA) with cavernous sinus (CS) compartments penetration and intracranial extension, aiming to improve the safety, effectiveness, and total resection rate of surgery. Methods: A case series of 120 Knosp4PA patients with 187 invaded compartments were retrospectively reviewed. A novel surgery-relevant grading system was proposed according to the CS penetrating features. The details of approach drafting, risk prediction, and complication avoidance were analyzed and integrated through illustrated cases. Results: All enrolled tumor was Knosp4PA which was derived from Knosp subgrades 3A(62.5%) and 3B(37.5%). Based on the tumor growth pathway and its relevant features, five subclassifications of intracranial extension(n=98,81.7%) were classified, which derived from the superior (Dolenc's and Oculomotor subtype, 5% and 24.2%), lateral (Parkinson's subtype,18.3%), and posterior (cerebral peduncle and Dorello's subtype, 5.8% and 1.7%) CS compartment penetration. The size of intracranial extension is assessed by Lou's scale proposed here based on preoperative MRI characteristics. Under Lou's scale, the gross total rate (GTR) decreased (82%, 53%, 22%, and 19%) with grades increased (grade 0,1,2,3, respectively), and presents significant difference between the four groups (p=0.000), as well as between single and multiple compartments involved (p=0.001). Preoperative cranial nerve deficits included the optic nerve (53%), oculomotor nerve (24.2%), and abducent nerve (4.2%), with an overall rate of visual function improvement in 68.1%. Postoperative complications of transient diabetes insipidus, cerebrospinal fluid (CSF) leakage, and cranial nerve deficits were 6.7%, 0.8%, and 0%. No new cranial nerve deficits occurred. The mortality rate was 0.8%. Conclusion: The concept of "penetration" refines the extracavernous growth pattern, and the five intracranial subclassifications help to understand the potential extension corridors, enhancing adequate exposure and targeted resection of Knosp4PA. This grading system may benefit from its predictive and prognostic value, from which a higher GTR rate can be achieved.

Corrigendum: Endoscope-Assisted Retrosigmoid Approach for Vestibular Schwannomas with Intracanalicular Extensions: Facial Nerve Outcomes.

[This corrects the article DOI: 10.3389/fonc.2021.774462.].

Endoscope-Assisted Retrosigmoid Approach for Vestibular Schwannomas With Intracanalicular Extensions: Facial Nerve Outcomes.

OBJECTIVE: To explore the role of neuroendoscope assistance during surgical resection of the intracanalicular portion of vestibular schwannomas via the retrosigmoid approach and the subsequent early facial nerve outcomes. METHODS: Patients of vestibular schwannoma with intracanalicular extensions undergoing retrosigmoid dissection at a single institution were retrospectively analyzed in this study. Several surgical techniques were applied to ensure maximal and safe removal of tumors. Tumors extending less than 10 mm into the internal acoustic canal (IAC) were classified as Grade A, while those extending over 10 mm into IAC were taken as Grade B. Neuroendoscope was applied at the end of microscopic phase to search for potential remnants for Grade B tumors. Absolute tumor extension was defined and measured. House and Brackmann (HB) scale was used to evaluate immediate CN VII outcomes. RESULTS: Of the 61 patients, there were 38 females and 23 males. A total of 18 (29.51%) cases were Koos Grade II, 12 (19.67%) cases Koos Grade III, and 31 (50.82%) cases Koos Grade IV. There were 38 cases (62.30%) of Grade A and 23 cases (37.70%) of Grade B. Gross total resection was achieved in 60 cases (98.36%). Four cases of intracanalicular remnants were detected and completely removed under endoscopic visualizations. There was a significantly higher proportion (17%, p = 0.02) of intracanalicular remnants in Grade B than Grade A. CN VII and VIII were anatomically preserved in all cases. A total of 55 cases (90.16%) retained good (HB Grades 1 and 2) facial nerve outcomes. CONCLUSIONS: In Grade B vestibular schwannomas, after maximal microsurgical removal, endoscopic evaluation of the intracanalicular portion revealed residual tumors in 17% of the patients. Hence endoscopic evaluation of the potential intracanalicular remnants for tumor extending over 10 mm within IAC (Grade B) is recommended.

Endoscopic endonasal resection of sinonasal teratocarcinosarcoma with intracranial breakthrough: illustrative case.

BACKGROUND: Teratocarcinosarcoma traversing the anterior skull base is rarely reported in literature. The heterogenous and invasive features of the tumor pose challenges for surgical planning. With technological advancements, the endoscopic endonasal approach (EEA) has been emerging as a workhorse of anterior skull base lesions. To date, no case has been reported of EEA totally removing teratocarcinosarcomas with intracranial extensions. OBSERVATIONS: The authors provided an illustrative case of a 50-year-old otherwise healthy man who presented with left-sided epistaxis for a year. Imaging studies revealed a 31 × 60-mm communicating lesion of the anterior skull base. Gross total resection via EEA was achieved, and multilayered skull base reconstruction was performed. LESSONS: The endoscopic approach may be safe and effective for resection of extensive teratocarcinosarcoma of the anterior skull base. To minimize the risk of postoperative cerebrospinal fluid leaks, multilayered skull base reconstruction and placement of lumbar drainage are vitally important.

Deregulation of the Hippo Pathway Promotes Tumor Cell Proliferation Through YAP Activity in Human Sporadic Vestibular Schwannoma.

OBJECTIVE: Vestibular schwannomas (VSs) can cause serious neurological defects including hearing loss and facial paralysis. The aim of this study is to identify whether Hippo signaling could be a potential targetable pathway for clinical treatment in VSs. METHODS: Gene expression profiling was performed in 10 sporadic VSs and 4 normal nerves to identify aberrant genes expression of the Hippo pathway. Western blotting and immunohistochemical staining were used to examine the expression of Hippo core components in 20 VS samples. Neurofibromatosis type 2 (NF2) gene sequencing was also performed in all tumors using sanger sequencing. Verteporfin, inhibitor of yes-associated protein (YAP)-TEA domain family member, was used to assess the effect of proliferation inhibition in human primary VS cells and RT4-D6P2T cell line. RESULTS: We found 51 differentially expressed genes of the Hippo pathway between VSs and healthy controls. Unsupervised analysis identified the 2 molecular variants that significantly related with distinct NF2 mutation status. The phosphorylation levels of large tumor suppressor 1 and YAP were significantly decreased in NF2-mutated VSs compared with wild-type VSs and normal nerves. Immunohistochemical staining showed that increased nuclear YAP expression in VSs was positively correlated with high Ki-67 index and low Merlin expression. Verteporfin reduced viability of primary VS cells and RT4-D6P2T cells. CONCLUSIONS: Our findings implicate that deregulation of the Hippo pathway as a molecular mechanism of pathogenesis in human VSs, and suggest inhibition of this pathway as a potential treatment strategy.

Prognostic value of Ki-67 index in adult medulloblastoma after accounting for molecular subgroup: a retrospective clinical and molecular analysis.

INTRODUCTION: Medulloblastoma (MB) is a rare primary brain tumor in adults. We previously evaluated that combining both clinical and molecular classification could improve current risk stratification for adult MB. In this study, we aimed to identify the prognostic value of Ki-67 index in adult MB. METHOD: Ki-67 index of 51 primary adult MBs was reassessed using a computer-based image analysis (Image-Pro Plus). All patients were followed up ranging from 12 months up to 15 years. Gene expression profiling and immunochemistry were used to establish the molecular subgroups in adult MB. Combined risk stratification models were designed based on clinical characteristics, molecular classification and Ki-67 index, and identified by multivariable Cox proportional hazards analysis. RESULTS: In our cohort, the mean Ki-67 value was 30.0 ± 11.3% (range 6.56-63.55%). The average Ki-67 value was significantly higher in LC/AMB than in CMB and DNMB (P = .001). Among three molecular subgroups, Group 4-tumors had the highest average Ki-67 value compared with WNT- and SHH-tumors (P = .004). Patients with Ki-67 index large than 30% displayed poorer overall survival (OS) and progression free survival (PFS) than those with Ki-67 less than 30% (OS: P = .001; PFS: P = .006). Ki-67 index (i.e. > 30%, < 30%) was identified as an independent significant prognostic factor (OS: P = .017; PFS: P = .024) by using multivariate Cox proportional hazards model. CONCLUSIONS: In conclusion, Ki-67 index can be considered as a valuable independent prognostic biomarker for adult patients with MB.

Surgical treatment of large vestibular schwannomas in patients with neurofibromatosis type 2: outcomes on facial nerve function and hearing preservation.

Surgical treatment of vestibular schwannoma (VS) in patients with neurofibromatosis type 2 (NF2) along with functional preservation of cranial nerves is challenging. The aim of this study was to analyze the outcomes of hearing and facial nerve function in patients with NF2 who underwent large-size VS (> 2 cm) surgery. From 2006 to 2016, one hundred and forty NF2 patients were included with 149 large-size VS resections using retrosigmoid approach. Hearing function was classified according to the American Academy of Otolaryngology-Head and Neck Surgery (AAO-HNS) criteria. Preoperative and one-year postoperative facial nerve function were both assessed using the House-Brackmann (H-B) grading scale. A multivariate logistic regression was performed to identify preoperative predictors for facial function outcomes. No operative death we noted. Total tumor removal was achieved in 82.6% of the operated VSs. The anatomical integrity of the facial nerve was preserved in 67.8% of surgeries. Good facial nerve function (H-B Grades I-III) was maintained in 49.6% of patients at 12 months after surgery. Tumor size larger than 3 cm and preoperative facial weakness related with worse outcome of facial nerve function (P < 0.001; for both). Hearing preservation surgeries were attempted in 31 ears. Class B or C hearing according to the AAO-HNS criteria was maintained in 7 ears (22.5%), and measurable hearing was maintained 11 ears (35.5%). It is challenging to maintain hearing and facial nerve function in NF2 patients with large VSs. Early surgical intervention is an appropriate choice to decrease the risk of neurological functions deficit.

Awake craniotomy for assisting placement of auditory brainstem implant in NF2 patients.

OBJECTIVES: Auditory brainstem implants (ABIs) may be the only opportunity for patients with NF2 to regain some sense of hearing sensation. However, only a very small number of individuals achieved open-set speech understanding and high sentence scores. Suboptimal placement of the ABI electrode array over the cochlear nucleus may be one of main factors for poor auditory performance. In the current study, we present a method of awake craniotomy to assist with ABI placement. METHODS: Awake surgery and hearing test via the retrosigmoid approach were performed for vestibular schwannoma resections and auditory brainstem implantations in four patients with NF2. Auditory outcomes and complications were assessed postoperatively. RESULTS: Three of 4 patients who underwent awake craniotomy during ABI surgery received reproducible auditory sensations intraoperatively. Satisfactory numbers of effective electrodes, threshold levels and distinct pitches were achieved in the wake-up hearing test. In addition, relatively few electrodes produced non-auditory percepts. There was no serious complication attributable to the ABI or awake craniotomy. CONCLUSIONS: It is safe and well tolerated for neurofibromatosis type 2 (NF2) patients using awake craniotomy during auditory brainstem implantation. This method can potentially improve the localization accuracy of the cochlear nucleus during surgery.

Distinctive localization and MRI features correlate of molecular subgroups in adult medulloblastoma.

Medulloblastoma (MB) is recognized as comprising four molecular subgroups with distinct transcriptional profiles, clinical features, and outcomes. Previous studies demonstrate that pediatric MBs present with subgroup-specific MRI manifestations. We hypothesized that combination of anatomical localization and conventional features based on MR imaging can predict these subgroups in adult MBs. MR Imaging manifestations of 125 adult patients with MB were analyzed retrospectively based on pre-operative MRI scans. MB molecular subgroups were evaluated by the expression profiling array and immunohistochemistry. A pediatric MB cohort of 60 patients were analyzed for comparison with data of adult patients. Multiple logistic regression analysis revealed that tumor location (P < 0.0001) and pattern of enhancement (P = 0.0048) were significantly correlated with molecular subgroups in adult MBs. Ninety-two percent of adult MBs were correctly predicted by using logistic regression model based on the anatomical localization patterns and pattern of enhancement. Exclusively intra-cerebellar growth, localization in the rostral cerebellum, and no brainstem contact were specific to adult SHH-MBs. Group 4-MBs in adult were characterized by minimal/no enhancement compared with other two subgroups. Infant SHH-MBs represented significant different localization patterns compared with SHH tumors in children and adults. We identified that molecular subgroups of adult MBs could be well predicted by tumor localization patterns and enhancement pattern. Our study also provided important evidence that MB subgroups in adult possibly derived from different cellular origins.

Risk Factors and Management of Intraoperative Cerebrospinal Fluid Leaks in Endoscopic Treatment of Pituitary Adenoma: Analysis of 492 Patients.

OBJECTIVES: To determine risk factors and management of intraoperative cerebrospinal fluid (CSF) leakage in endoscopic endonasal transsphenoidal pituitary adenoma surgery. METHODS: We conducted a retrospective review of 492 patients who, between April 2012 and August 2015, underwent endoscopic endonasal transsphenoidal surgeries for resection of pituitary adenoma. A multivariate statistical analysis was performed to investigate the association of some risk factors with intraoperative CSF leakage. Intraoperative CSF leaks were classified as grade 0, no leak observed; grade 1, small leak without obvious diaphragmatic defect; grade 2, moderate leak; or grade 3, large diaphragmatic defect. Repair methods were based on the CSF leak grade. RESULTS: Intraoperative CSF leakage occurred in 86 cases (17.5%). On univariate analysis, there were 3 factors associated with an increased intraoperative CSF leak rate: 1) repeat surgery (repeat 30.0% vs. primary 16.4%; P = 0.033), 2) consistency of the adenoma (tenacious, 27.3% vs. soft, 13.5%; P = 0.000), and 3) tumor size (22.0 ± 9.7mm vs. 25.4 ± 11.5 mm; P = 0.007). However, on multivariate analysis, only tumor consistency (P = 0.001; odds ratio, 2.379) and tumor size (P = 0.026; odds ratio, 1.032) were independently associated with intraoperative CSF leaks. In the 86 cases with intraoperative CSF leaks, the degree of intraoperative CSF leakage was categorized grade 1 in 30 cases, grade 2 in 25 cases, and grade 3 in 31 cases. Postoperative CSF leak repair failures occurred in 6 cases (1.2%). CONCLUSIONS: Intraoperative CSF leaks have a propensity to occur in cases with fibrous or large tumors. Once an intraoperative leak is identified, our graded cranial base repair method is safe and reliable.

A Retrospective Analysis of Vision-Impairing Tumors Among 467 Patients with Neurofibromatosis Type 2.

BACKGROUND: Vision is important for patients with hearing loss caused by neurofibromatosis type 2 (NF2). Tumors adjacent to the anterior visual pathway can potentially impair the vision. Only a few case reports and small-series studies have been reported. OBJECTIVE: To evaluate the clinical features of tumors adjacent to the anterior visual pathway in a large series of patients with NF2. METHODS: Seventy-three patients with potentially vision-impairing tumors were carefully screened from among 467 patients with NF2. RESULTS: Among the 73 patients, 31 had intraorbital tumors, 21 had suprasellar meningiomas, and 21 had medial sphenoid ridge meningiomas. Of the 31 patients with intraorbital tumors, 17 had optic nerve sheath meningiomas, 9 had intraorbital schwannomas, 3 had spheno-orbital meningiomas, 1 had an anterior cranial fossa-orbital meningioma, and 1 had a cranio-orbital schwannoma. To the date of the last follow-up, 43 patients (58.9%) experienced visual loss. In most cases, hearing loss tended to occur earlier than visual loss. Six patients underwent early operations, and they recovered well without any further vision damage. Six other patients underwent operations after having no functional visual ability in the affected eyes, and their visual ability was not saved. CONCLUSIONS: Tumors adjacent to the anterior visual pathway, although uncommon in patients with NF2, can cause progressive visual loss. Early surgical intervention seems to be the primary treatment strategy, except for in patients' optic nerve sheath meningiomas. If patients adopt a wait and see policy, regular visual examination seems to be mandatory.

Granular cell tumors in the central nervous system: a report on eight cases and a literature review.

OBJECTIVE: Granular cell tumors (GCTs) in the central nervous system (CNS) are extremely rare. We report here a series of eight GCTs at various locations in the CNS and provide a full review of the published literature. METHODS: Eight patients with pathologically confirmed GCT in the CNS were retrospectively reviewed. The patients were followed up via telephone interview or an outpatient department. RESULTS: Five patients were female, and three were male. Of the eight tumors, three were located at the sellar region, two were located in the spinal canal, one was located at the cerebral hemisphere, one was parasellar-nasal communicating, and one was combined with an enterogenous cyst at the ventral aspect of the medulla oblongata. Six patients were symptomatic, and two patients were found incidentally. Most GCTs tended to be homogeneous and well defined on radiological images. The parasellar-nasal communicating GCT showed destruction of the skull base and an evident cystic component in the sphenoid sinus. The various GCTs had similar histological features, and they tended to be histologically benign. Most tumors did not recur after the operation. After subtotal resection, one residual spinal GCT regrew during the follow-up period. The patient with the parasellar-nasal communicating GCT developed progressive visual loss in the right eye after the operation; and she received adjuvant radiation therapy. CONCLUSION: GCTs at different locations in the CNS can have significantly different clinical features and should be considered distinct entities. Except for the granular cell astrocytoma (GCA), most GCTs in the CNS tended to be benign. Malignant GCTs are rare and difficult to treat.

Clinical features of clear cell meningioma: a retrospective study of 36 cases among 10,529 patients in a single institution.

BACKGROUND: Clear cell meningioma (CCM) is a rare subtype of meningioma. We present the largest series of 36 CCMs and evaluate several prognostic factors of patient's clinical outcome. METHODS: Thirty-six patients with pathologically confirmed CCM among a total of 10,529 meningioma patients were retrospectively reviewed. RESULTS: CCM constituted 0.3 % of the intracranial meningiomas and 1.4 % of the intraspinal meningiomas. The male-to-female ratio (36 vs 64 %) for CCMs was similar to that for total meningiomas (28 vs 72 %) patients (chi-squared test, p = 0.3). The mean age at diagnosis of CCM patients (29.3 ± 18.4 years) was significantly younger than that of total meningiomas (49.8 ± 11.9 years) patients (t-test, p = 0). During the follow-up, 15 patients (42 %) suffered from tumor recurrence. The recurrence time ranged from 10 months to 12 years, with a median time of 29 months. Kaplan-Meier survival analysis showed that patients after total resection (Simpson grades I and II) had significantly longer progression-free survival (PFS) time than those after subtotal resection (Simpson grades III and IV) (log-rank test, p = 0.006). However, age (≤20 years or >20 years, p = 0.9), gender (p = 0.3), postoperative radiotherapy (p = 0.4), progesterone receptor staining (positivity or negativity, p = 0.2), and Ki-67 index (≤5 % or >5 %, p = 0.4) did not have significant effects on patients' PFS time. CONCLUSIONS: The proportion of CCM in spinal meningiomas is likely to be much larger than that in intracranial meningiomas. CCMs should be resected totally when possible to decrease the risk of recurrence or prolong patient's PFS time.

Transplantation of Human Neural Stem Cells in a Parkinsonian Model Exerts Neuroprotection via Regulation of the Host Microenvironment.

Parkinson's disease (PD) is characterized by a progressive loss of dopaminergic neurons and consequent dopamine (DA) deficit, and current treatment still remains a challenge. Although neural stem cells (NSCs) have been evaluated as appealing graft sources, mechanisms underlying the beneficial phenomena are not well understood. Here, we investigate whether human NSCs (hNSCs) transplantation could provide neuroprotection against DA depletion by recruiting endogenous cells to establish a favorable niche. Adult mice subjected to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) were transplanted with hNSCs or vehicle into the striatum. Behavioral and histological analyses demonstrated significant neurorescue response observed in hNSCs-treated animals compared with the control mice. In transplanted animals, grafted cells survived, proliferated, and migrated within the astrocytic scaffold. Notably, more local astrocytes underwent de-differentiation, acquiring the properties of NSCs or neural precursor cells (NPCs) in mice given hNSCs. Additionally, we also detected significantly higher expression of host-derived growth factors in hNSCs-transplanted mice compared with the control animals, together with inhibition of local microglia and proinflammatory cytokines. Overall, our results indicate that hNSCs transplantation exerts neuroprotection in MPTP-insulted mice via regulating the host niche. Harnessing synergistic interaction between the grafts and host cells may help optimize cell-based therapies for PD.

The progress of immunotherapy for glioblastoma.

Glioblastoma is the most common primary brain tumor in adults, accounting for about half of all primary brain tumors. Despite multiple therapeutic interventions such as surgical resection, radiotherapy, and systemic chemotherapy, the prognosis for glioblastoma remains poor. Due to the scientific community's enhanced understanding of the CNS immune system and significant achievements in tumor immunotherapy in recent years, immunotherapy has become a promising GBM treatment. In vaccine therapy, a number of clinical trials have achieved encouraging results. In antibody therapy, antibodies are used to target immune checkpoints such as ipilimumab and nivolumab. Bioengineering technology has also lead to a new field of tumor immunotherapy, whereby genetically modified tumor-specific T cells are reintroduced into a patient's body.

Brain Stem and Entire Spinal Leptomeningeal Dissemination of Supratentorial Glioblastoma Multiforme in a Patient during Postoperative Radiochemotherapy: Case Report and Review of the Literatures.

Glioblastoma multiforme (GBM) is the most common primary malignancy of the central nervous system in adults. Macroscopically evident and symptomatic spinal metastases occur rarely. Autopsy series suggest that approximately 25% of patients with intracranial GBM have evidence of spinal subarachnoid seeding, although the exact incidence is not known as postmortem examination of the spine is not routinely performed. Herein, we present a rare case of symptomatic brain stem and entire spinal dissemination of GBM in a 36-year-old patient during postoperative adjuvant radiochemotherapy with temozolomide and cisplatin. Visual deterioration, intractable stomachache, and limb paralysis were the main clinical features. The results of cytological and immunohistochemical tests on the cerebrospinal fluid cells were highly suggestive of spinal leptomeningeal dissemination. After 1 month, the patient's overall condition deteriorated and succumbed to his disease. To the best of our knowledge, this is the first reported case of GBM dissemination presenting in this manner. Because GBM extracranial dissemination is rare, we also reviewed pertinent literature regarding this uncommon entity. Although metastases to spinal cord from GBM are uncommon, it is always important to have in mind when patients with a history of GBM present with symptoms that do not correlate with the primary disease pattern.