ABSTRACT
OBJECTIVES: Large (De Serres stage [IV-V]) head and neck lymphatic malformations (HNLMs) often have multiple, high-risk, invasive treatments (ITs) to address functional compromise. Logically reducing HNLM ITs should reduce treatment risk. We tested whether delaying HNLM ITs reduces total IT number. MATERIALS: Consecutive HNLM patients (n = 199) between 2010 and 2017, aged 0-18 years. METHODS: ITs (surgery or sclerotherapy) were offered for persistent or dysfunction causing HNLMs. Treatment effectiveness categorized by IT number: optimal (0-1), acceptable (2-5), or suboptimal (>5). Clinical data were summarized, and outcome associations tested (χ2 ). Relative risk (RR) with a Poisson working model tested whether HNLM observation or IT delay (>6 months post-diagnosis) predicts treatment success (i.e., ≤1 IT). RESULTS: Median age at HNLM diagnosis was 1.3 months (interquartile range [IQR] 0-45 m) with 107/199(54%) male. HNLM were stage I-III (174 [88%]), IV-V (25 [13%]). Initial treatment was observation (70 [35%]), invasive (129 [65%]). Treatment outcomes were optimal (137 [69%]), acceptable (36 [18%]), and suboptimal (26 [13%]). Suboptimal outcome associations: EXIT procedure, stage IV-V, oral location, and tracheotomy (p < 0.001). Stage I-III HNLMs were initially observed compared with stage I-III having ITs within 6 months of HNLM diagnosis, had a 82% lower relative treatment failure risk ([i.e., >1 IT], RR = 0.09, 95% CI 0.02-0.36, p < 0.001). Stage I-III HNLMs with non-delayed ITs had reduced treatment failure risk compared with IV-V (RR = 0.47, 95% CI 0.33-0.66, p < 0.001). CONCLUSION: Observation and delayed IT in stage I-III HNLM ("Grade 1") is safe and reduces IT (i.e., ≤1 IT). Stage IV-V HNLMs ("Grade 2") with early IT have a greater risk of multiple ITs. LEVEL OF EVIDENCE: 4 Laryngoscope, 133:956-962, 2023.
Subject(s)
Head , Lymphatic Abnormalities , Humans , Male , Infant , Female , Neck , Lymphatic Abnormalities/surgery , Treatment Outcome , Sclerotherapy/methodsABSTRACT
PURPOSE: PIK3CA-related overgrowth spectrum (PROS) conditions of the head and neck are treatment challenges. Traditionally, these conditions require multiple invasive interventions, with incomplete malformation removal, disfigurement, and possible dysfunction. Use of the PI3K inhibitor alpelisib, previously shown to be effective in PROS, has not been reported in PIK3CA-associated head and neck lymphatic malformations (HNLMs) or facial infiltrating lipomatosis (FIL). We describe prospective treatment of 5 children with PIK3CA-associated HNLMs or head and neck FIL with alpelisib monotherapy. METHODS: A total of 5 children with PIK3CA-associated HNLMs (n = 4) or FIL (n = 1) received alpelisib monotherapy (aged 2-12 years). Treatment response was determined by parental report, clinical evaluation, diary/questionnaire, and standardized clinical photography, measuring facial volume through 3-dimensional photos and magnetic resonance imaging. RESULTS: All participants had reduction in the size of lesion, and all had improvement or resolution of malformation inflammation/pain/bleeding. Common invasive therapy was avoided (ie, tracheotomy). After 6 or more months of alpelisib therapy, facial volume was reduced (range 1%-20%) and magnetic resonance imaging anomaly volume (range 0%-23%) were reduced, and there was improvement in swallowing, upper airway patency, and speech clarity. CONCLUSION: Individuals with head and neck PROS treated with alpelisib had decreased malformation size and locoregional overgrowth, improved function and symptoms, and fewer invasive procedures.
Subject(s)
Phosphatidylinositol 3-Kinases , Thiazoles , Child , Humans , Phosphatidylinositol 3-Kinases/genetics , Mutation , Class I Phosphatidylinositol 3-Kinases/genetics , Thiazoles/therapeutic useABSTRACT
Somatic activating variants in PIK3CA, the gene that encodes the p110α catalytic subunit of phosphatidylinositol 3-kinase (PI3K), have been previously detected in â¼80% of lymphatic malformations (LMs).1 , 2 We report the presence of somatic activating variants in BRAF in individuals with LMs that do not possess pathogenic PIK3CA variants. The BRAF substitution p.Val600Glu (c.1799T>A), one of the most common driver mutations in cancer, was detected in multiple individuals with LMs. Histology revealed abnormal lymphatic channels with immunopositivity for BRAFV600E in endothelial cells that was otherwise indistinguishable from PIK3CA-positive LM. The finding that BRAF variants contribute to low-flow LMs increases the complexity of prior models associating low-flow vascular malformations (LM and venous malformations) with mutations in the PI3K-AKT-MTOR and high-flow vascular malformations (arteriovenous malformations) with mutations in the RAS-mitogen-activated protein kinase (MAPK) pathway.3 In addition, this work highlights the importance of genetic diagnosis prior to initiating medical therapy as more studies examine therapeutics for individuals with vascular malformations.
ABSTRACT
OBJECTIVE: Infant and toddler subdural haemorrhages (SDH) are often considered indicative of abuse or major trauma. However, accidental impact events, such as falls, cause contact extra-axial haemorrhages (EAHs). The current study sought to determine frequency and clinical behaviour of EAHs with infant and toddler accidental and abusive skull fractures. PATIENTS AND METHODS: Children aged <4 years with accidental skull fractures and abusive fractures identified by CT at two paediatric tertiary care centres. Clinical data were abstracted by child abuse paediatricians and images were reviewed by paediatric radiologists. Data were analysed using univariate and multivariate logistic regression as well as descriptive statistics. RESULTS: Among 227 subjects, 86 (37.9%) had EAHs. EAH was present in 73 (34.8%) accidental and 13 (76.5%) of the abusive injuries. Intracranial haemorrhage rates were not different for children with major or minor accidents but were fewer than abused. EAH was equally common with falls <4 and >4 ft. EAH depths did not differ by mechanism, but 69% of accidental EAHs were localised solely at fracture sites vs 38% abuse. Widespread and multifocal EAHs were more common with abuse. Children with abuse or major accidental injuries presented with lower initial Glasgow Coma Scales than those with minor accidents. Abused children had initial loss of consciousness more often than those with either minor or major accidents. CONCLUSIONS: Simple contact EAHs were common among children with minor and major accidental skull fractures. Accidental EAHs were more localised with less neurological dysfunction than abusive.
Subject(s)
Child Abuse , Skull Fractures , Accidents , Child , Child Abuse/diagnosis , Child, Preschool , Hematoma, Subdural , Humans , Infant , Retrospective Studies , Skull Fractures/complications , Skull Fractures/diagnostic imagingABSTRACT
OBJECTIVE: The aim of the study was to determine whether complex skull fractures are more indicative of child abuse or major trauma than simple skull fractures. DESIGN: This is a retrospective chart and imaging review of children diagnosed with a skull fracture. Subjects were from 2 pediatric tertiary care centers. Children younger than 4 years who underwent a head computed tomography with 3-dimensional rendering were included. We reviewed the medical records and imaging for type of skull fracture, abuse findings, and reported mechanism of injury. A complex skull fracture was defined as multiple fractures of a single skull bone, fractures of more than 1 skull bone, a nonlinear fracture, or diastasis of greater than 3 mm. Abuse versus accident was determined at the time of the initial evaluation with child abuse physician team confirmation. RESULTS: From 2011 to 2012, 287 subjects were identified by International Classification of Diseases, Ninth Revision, code. The 147 subjects with a cranial vault fracture and available 3-dimensional computed tomography composed this study's subjects. The average age was 12.3 months. Seventy four (50.3%) had complex and 73 (49.7%) had simple fractures. Abuse was determined in 6 subjects (4.1%), and a determination could not be made for 5 subjects. Adding abused children from 2013 to 2014 yielded 15 abused subjects. Twelve of the abused children (80%) had complex fractures; more than the 66 (48.5%) of 136 accidentally injured children (P = 0.001; relative risk = 1.65 [1.21-2.24]). However, among children with a complex fracture, the positive predictive value for abuse was only 7%. CONCLUSIONS: Complex skull fractures frequently occur from accidental injuries. This study suggests that the presence of complex skull fractures should not be used alone when making a determination of abuse.
Subject(s)
Child Abuse , Craniocerebral Trauma , Skull Fractures , Child , Child Abuse/diagnosis , Humans , Infant , Retrospective Studies , Skull , Skull Fractures/diagnostic imaging , Skull Fractures/etiology , Tomography, X-Ray ComputedABSTRACT
OBJECTIVES: Head and neck lymphatic malformations (HNLM) are caused by gain-of-function somatic mutations in PIK3CA. Acetylsalicylic acid (ASA/aspirin) is thought to limit growth in PIK3CA-mutated neoplasms through PI3K pathway suppression. We sought to determine if ASA could be beneficial for HNLM. METHODS: Retrospective case series of patients (0-18 years) offered ASA (3-5 mg/kg/day) for HNLM treatment (2010-2018). Clinical and treatment characteristics, patient-reported symptom improvement, medication tolerance, compliance, and complications were recorded. Treatment response was determined by change in patient/caregiver-reported symptoms, or HNLM size [complete (resolved), partial (decreased), or stable]. RESULTS: Fifty-three patients were offered ASA, 23 (43%) accepted (median age 10 years, IQR 6-14). Compared to patients who declined, patients receiving ASA were more likely to have extensive malformations: ex-utero intrapartum treatment procedure, bilateral malformations, oral cavity location, ≥2 invasive treatments, or tracheotomy (p < 0.05). All patients with tissue available had PIK3CA mutations (13/23). Treatment indications included oral pain/blebs (12, 52%), recurrent pain/swelling (6, 26%), or sudden/persistent swelling (5, 22%). Treatment plan was commonly one 81 mg tablet daily (19, 83%) for 3-12 months (8, 42%). Therapeutic adherence was reported by 18 patients (78%). Symptoms improved in 18 patients [78%; decreased pain (9, 39%) and swelling (8, 35%)]. Treatment resulted in partial (14, 61%) or complete response (4, 17%). Three patients developed oral bleb bleeding, which resolved with medication discontinuation. CONCLUSION: ASA seems to be a well-tolerated, low-risk medication for HNLM treatment. This pilot study suggests that it often improves symptoms and reduces HNLM size. Further prospective, randomized studies are warranted to comprehensively assess indications, safety, and efficacy. LEVEL OF EVIDENCE: Level 4.
Subject(s)
Lymphatic Abnormalities , Phosphatidylinositol 3-Kinases , Aspirin/therapeutic use , Child , Humans , Lymphatic Abnormalities/drug therapy , Lymphatic Abnormalities/genetics , Pilot Projects , Retrospective StudiesABSTRACT
OBJECTIVES: The aim of this study was to determine how reliable scalp bruising and soft tissue swelling/cephalohematomas (STS) are for underlying young child skull fractures. METHODS: This was a retrospective clinical and imaging review from 2011 to 2012 of children younger than 4 years with skull fractures from 2 tertiary care hospitals. Imaging was reread by 3 pediatric radiologists. Descriptive statistics were utilized. The retrospective review had institutional review board approval. RESULTS: We identified 218 subjects for review: 210 unintentional and 8 abusive. One hundred forty-three had available 3-dimensional computed tomography reconstructions: 136 unintentional and 7 abused. Two-thirds were younger than 1 year. Twelve subjects had visible scalp bruising, but 73% had clinically and 93% radiologically apparent fracture-associated STS. There was no difference in clinical STS with simple (79%) versus complex (68%) fractures. Nor was there difference in subjects with fractures from minor (77%) versus major (70%) trauma. Unintentionally injured infants did not differ from abused for detectable STS (74% vs 50%). Parietal and frontal bones most frequently sustained fractures and most consistently had associated STS. CONCLUSIONS: Clinically apparent STS is present in approximately three-fourths of children with skull fractures. It may not be important to consistently identify skull fractures in unintentionally injured children. Point-of-care ultrasound may be adequate. For abuse concerns, it is important to identify skull fractures as evidence of cranial impacts and intracranial hemorrhages. The most sensitive, widely available imaging technique, cranial computed tomography scan with 3-dimensional skull reconstruction, should be utilized. Scalp bruising is present in a minority of young children with skull fractures. Its absence does not exclude cranial impact injury.
Subject(s)
Child Abuse , Contusions , Craniocerebral Trauma , Skull Fractures , Child , Child Abuse/diagnosis , Child, Preschool , Contusions/diagnostic imaging , Contusions/etiology , Craniocerebral Trauma/complications , Craniocerebral Trauma/diagnostic imaging , Humans , Infant , Retrospective Studies , Skull Fractures/complications , Skull Fractures/diagnostic imagingABSTRACT
OBJECTIVES: An increasing number of treatment modalities for lymphatic malformations are being described, complicating therapeutic decisions. Understanding lymphatic malformation natural history is essential. We describe management of head and neck lymphatic malformations where decisions primarily addressed lesion-induced functional compromise (ie, breathing, swallowing) to identify factors associated with invasive treatment and active observation. We hypothesize that non-function threatening malformations can be observed. STUDY DESIGN: Retrospective case series. METHODS: Retrospective case series of consecutive head and neck lymphatic malformation patients (2000-2017) with over 2 years of follow-up. Patient characteristics were summarized and associations with invasive treatment (surgery or sclerotherapy) tested using Fisher's exact. In observed patients, factors associated with spontaneous regression were assessed with Fisher's exact test. RESULTS: Of 191 patients, 101 (53%) were male, 97 (51%) Caucasian, and 98 (51.3%) younger than 3 months. Malformations were de Serres I-III 167 (87%), or IV-V 24 (12%), and commonly located in the neck (101, 53%), or oral cavity (36, 19%). Initial treatments included observation (65, 34%) or invasive treatments such as primary surgery (80, 42%), staged surgery (25, 13%), or primary sclerotherapy (9, 5%). Of 65 initially observed malformations, 8 (12%) subsequently had invasive treatment, 36 (58%) had spontaneous regression, and 21 (32%) elected for no invasive therapy. Spontaneous regression was associated with location in the lateral neck (P = .003) and macrocystic malformations (P = .017). CONCLUSION: Head and neck lymphatic malformation treatment selection can be individualized after stratifying by stage, presence of functional compromise, and consideration of natural history. Recognizing the spectrum of severity is essential in evaluating efficacy of emerging treatments, as selected malformations may respond to observation. LEVEL OF EVIDENCE: 4 Laryngoscope, 131:1392-1397, 2021.
Subject(s)
Head/abnormalities , Lymphatic Abnormalities/therapy , Neck/abnormalities , Watchful Waiting , Child, Preschool , Clinical Decision-Making , Databases, Factual , Female , Humans , Infant , Lymphatic Abnormalities/pathology , Male , Otorhinolaryngologic Surgical Procedures/statistics & numerical data , Prospective Studies , Retrospective Studies , Sclerotherapy/statistics & numerical data , Treatment OutcomeABSTRACT
PURPOSE: Vascular malformations (VM) are primarily caused by somatic activating pathogenic variants in oncogenes. Targeted pharmacotherapies are emerging but require molecular diagnosis. Since variants are currently only detected in malformation tissue, patients may be ineligible for clinical trials prior to surgery. We hypothesized that cell-free DNA (cfDNA) could provide molecular diagnoses for patients with isolated VM. METHODS: cfDNA was isolated from plasma or cyst fluid from patients with arteriovenous malformations (AVM), venous malformations (VeM), or lymphatic malformations (LM), and assayed for known pathogenic variants using droplet digital polymerase chain reaction (ddPCR). Cyst fluid cfDNA from an independent cohort of LM patients was prospectively screened for variants using a multiplex ddPCR assay. RESULTS: Variants were detected in plasma cfDNA in patients with AVM (2/8) and VeM (1/3). Variants were detected in cyst fluid cfDNA (7/7) but not plasma (0/26) in LM patients. Prospective testing of cyst fluid cfDNA with multiplex ddPCR identified variants in LM patients who had never undergone surgery (4/5). CONCLUSION: Variants were detected in plasma from AVM and VeM patients, and in cyst fluid from patients with LM. These data support investigation of cfDNA-based molecular diagnostics for VM patients, which may provide opportunities to initiate targeted pharmacotherapies without prior surgery.
Subject(s)
Cell-Free Nucleic Acids , Vascular Malformations , Cell-Free Nucleic Acids/genetics , Humans , Multiplex Polymerase Chain Reaction , Mutation , Prospective Studies , Vascular Malformations/diagnosis , Vascular Malformations/geneticsABSTRACT
OBJECTIVE: To describe the prevalence and clinical characteristics of airway findings in a multi-institutional cohort of PHACE patients. STUDY DESIGN: Multicenter retrospective case series. SETTING: Multidisciplinary vascular anomalies clinics at 2 institutions. METHODS: Data were collected from the electronic medical record, including clinical presentation, airway findings, treatment, and outcomes. RESULTS: Of 55 PHACE patients, 22 (40%) had airway hemangiomas. Patients with airway involvement were more commonly female (P = .034, odds ratio [OR] 23, 95% confidence interval [CI] 1.3-410) and of Caucasian ethnicity (P = .020, OR 5.3, 95% CI 1.3-21). Anatomically, patients with bilateral S3 involvement had higher rates of airway disease (P = .0012, OR 15, 95% CI 2.9-77). Most patients with airway hemangiomas had stridor (68%). Of the patients managed in the propranolol era (2008 or later, n = 35), 14 had airway involvement. All 14 were treated with propranolol, whereas 13 (62%) of 21 nonairway patients were treated with propranolol. The average treatment duration was longer in the airway patients (22.1 vs 16.7 months). All patients who underwent tracheostomy (n = 4) did so before 2008. CONCLUSION: Risk factors for airway involvement in PHACE include female gender, Caucasian ethnicity, and stridor. Since the widespread use of propranolol, fewer patients have required surgical management of their airway disease. Given the high prevalence of airway involvement even in patients without stridor, assessment of the airway is a crucial component of a comprehensive PHACE workup.
Subject(s)
Aortic Coarctation/complications , Eye Abnormalities/complications , Hemangioma/epidemiology , Hemangioma/therapy , Neurocutaneous Syndromes/complications , Respiratory Tract Neoplasms/epidemiology , Respiratory Tract Neoplasms/therapy , Aortic Coarctation/diagnosis , Aortic Coarctation/therapy , Eye Abnormalities/diagnosis , Eye Abnormalities/therapy , Female , Hemangioma/diagnosis , Humans , Infant , Infant, Newborn , Male , Neurocutaneous Syndromes/diagnosis , Neurocutaneous Syndromes/therapy , Prevalence , Propranolol/therapeutic use , Respiratory Tract Neoplasms/diagnosis , Retrospective Studies , Tracheostomy , Vasodilator Agents/therapeutic useABSTRACT
AIM: We sought to determine the frequency and patterns of delayed medical care seeking for young children with skull fractures. METHODS: We identified accidental skull fractures <4 years old, 2011-2012. Child abuse paediatricians abstracted retrospective data and paediatric radiologists re-reviewed images. 'Delays' were defined as presentation at ≥6 h. 'Minor accidents' included falls <4 feet and low force trauma, while 'major accidents' included higher height falls and major force events. We studied the frequency and duration of care delays, the signs or symptoms leading to care, and the duration of delays after signs or symptoms developed. RESULTS: Two hundred and ten children had accidental skull fractures. Delays were less likely with major accidents (4.9%), than with minor accidents (25.8%) (RR = 0.32 [0.15-0.70]). Children came to care for scalp swelling (STS) (39%), the injury event (36.2%), altered consciousness (15.2%) and vomiting (10.5%). Delayed onset of STS (78.6%) caused most delayed care. Early STS was firm, (17.6%) versus delayed (5.0%), as opposed to soft or fluctuant. CONCLUSION: Delayed care seeking is common for minor, but not major accidental infant and toddler skull fractures. Most followed delayed onset of signs and symptoms. Delayed care seeking alone should not imply child abuse.
Subject(s)
Child Abuse , Skull Fractures , Accidental Falls , Child , Child, Preschool , Humans , Infant , Retrospective Studies , Skull Fractures/diagnostic imaging , Skull Fractures/epidemiology , Skull Fractures/etiologyABSTRACT
PURPOSE: To investigate retinal function and visual outcomes in infants with retinal hemorrhages due to non-accidental trauma (NAT). METHODS: This is a retrospective review of full-field or multifocal electroretinogram (ERG) recordings, visual acuity in log minimum angle of resolution (logMAR), clinical status, and neuroimaging. Multifocal ERGs from the central 40° were compared to corresponding fundus imaging. Visual acuity was measured by Teller cards at follow-up. ERGs were compared to controls recorded under anesthesia. RESULTS: Sixteen children met inclusion criteria (14 recorded during the acute phase and 2 during long-term follow-up). During the acute phase, ERGs (n = 4 full field; n = 10 multifocal ERG) showed abnormal amplitude, latency, or both in at least one eye. Ten subjects had significantly reduced responses in both eyes, 3 of which had an ERG dominated by a negative waveform (absent b-wave or P1). The remaining six subjects had responses in one eye that were near normal (≥ 50% of controls). ERGs were sometimes abnormal in local areas without hemorrhage. ERGs could be preserved in local areas adjacent to traumatic retinoschisis. Two subjects with reduced visual acuity had belated ERGs: One had an abnormal macular ERG and the other had a normal macular ERG implying cortical visual impairment. At follow-up, 10 of 14 subjects had significant visual acuity loss (≥ 0.7 age-corrected logMAR); four subjects had mild vision loss (≤ 0.5 age-corrected logMAR). Visual acuity outcome was not reliably associated with the fundus appearance in the acute phase. All subjects with a negative ERG waveform had severe vision loss on follow-up. CONCLUSIONS: Retinal dysfunction was common during the acute phase of NAT. A near normal appearing fundus did not imply normal retinal function, and ERG abnormality did not always predict a poor visual acuity outcome. However, a negative ERG waveform was associated severe visual acuity loss. Potential artifacts of retinal hemorrhages and anesthesia could not fully account for multifocal ERG abnormalities. Retinal function can be preserved in areas adjacent to traumatic retinoschisis.
Subject(s)
Battered Child Syndrome/complications , Hematoma, Subdural/physiopathology , Retina/physiopathology , Retinal Hemorrhage/physiopathology , Visual Acuity/physiology , Child, Preschool , Electroretinography/methods , Female , Fundus Oculi , Hematoma, Subdural/diagnostic imaging , Hematoma, Subdural/etiology , Humans , Infant , Infant, Newborn , Male , Retrospective Studies , Tomography, Optical Coherence , Vision Disorders/physiopathologyABSTRACT
Lymphatic malformations (LMs) are congenital, nonneoplastic vascular malformations associated with postzygotic activating PIK3CA mutations. The mutation spectrum within LMs is narrow, with the majority having 1 of 3 hotspot mutations. Despite this relative genetic homogeneity, clinical presentations differ dramatically. We used molecular inversion probes and droplet digital polymerase chain reaction to perform deep, targeted sequencing of PIK3CA in 271 affected and unaffected tissue samples from 81 individuals with isolated LMs and retrospectively collected clinical data. Pathogenic PIK3CA mutations were identified in affected LM tissue in 64 individuals (79%) with isolated LMs, with variant allele fractions (VAFs) ranging from 0.1% to 13%. Initial analyses revealed no correlation between VAF and phenotype variables. Recognizing that different mutations activate PI3K to varying degrees, we developed a metric, the genotype-adjusted VAF (GVAF), to account for differences in mutation strength, and found significantly higher GVAFs in LMs with more severe clinical characteristics including orofacial location or microcystic structure. In addition to providing insight into LM pathogenesis, we believe GVAF may have broad applicability for genotype-phenotype analyses in mosaic disorders.
Subject(s)
Class I Phosphatidylinositol 3-Kinases/genetics , Genotype , Lymphatic Vessels/abnormalities , Mutation , Adolescent , Child , Child, Preschool , Female , Humans , Lymphatic Vessels/diagnostic imaging , Magnetic Resonance Imaging , Male , Retrospective Studies , Severity of Illness IndexABSTRACT
Our previous studies found that B16-F10 melanoma growth in the rear footpad of immunocompetent mice induces marked B cell accumulation within tumor-draining popliteal lymph nodes (TDLN). This B cell accumulation drives TDLN remodeling that precedes and promotes metastasis, indicating a tumor-promoting role for TDLN B cells. Here we show that phenotypic characterization of lymphocytes in mice bearing B16-F10 melanomas identifies preferential accumulation of T2-MZP B cells in the TDLN. Comparison of non-draining LNs and spleens of tumor-bearing mice with LNs and spleens from naïve mice determined that this pattern of B cell accumulation was restricted to the TDLN. B cell-deficient and immunocompetent mice reconstituted with T2-MZP B cells but not with other B cell subsets displayed accelerated tumor growth, demonstrating that T2-MZP B cells possess regulatory activity in tumor-bearing mice. Unlike splenic regulatory B cells, however, these TDLN B cells did not exhibit increased IL-10 production, nor did they promote Treg generation in the TDLN. These findings demonstrate that tumors initially signal via the lymphatic drainage to stimulate the preferential accumulation of T2-MZP regulatory B cells. This local response may be an early and critical step in generating an immunosuppressive environment to permit tumor growth and metastasis.
Subject(s)
B-Lymphocytes, Regulatory/immunology , Lymph Nodes/immunology , Neoplasms/immunology , Neoplasms/pathology , Adoptive Transfer , Animals , Cell Proliferation , Female , Lymphocyte Subsets/immunology , Male , Mice, Inbred C57BL , T-Lymphocytes/immunologyABSTRACT
BACKGROUND: To use high resolution MRI lymphography to characterize altered tumor-draining lymph node (TDLN) lymph drainage in response to growth of aggressive tumors. METHODS: Six mice bearing B16-F10 melanomas in one rear footpad were imaged by 3.0 Tesla (T) MRI before and after subcutaneous injection of Gadofosveset trisodium (Gd-FVT) contrast agent into both rear feet. Gd-FVT uptake into the left and right draining popliteal LNs was quantified and compared using Wilcoxon signed-rank test. Fluorescent dextran lymphography compared patterns of LN lymph drainage with the pattern of immunostained lymphatic sinuses by fluorescence microscopy. RESULTS: TDLNs exhibited greater Gd-FVT uptake than contralateral uninvolved LNs, although this difference did not reach significance (P < 0.06). Foci of contrast agent consistently surrounded the medulla and cortex of TDLNs, while Gd-FVT preferentially accumulated in the cortex of contralateral LNs at 5 and 15 min after injection. Fluorescent dextran lymphography confirmed these distinct contrast agent uptake patterns, which correlated with lymphatic sinus growth in TDLNs. CONCLUSION: 3.0T MRI lymphography using Gd-FVT identified several distinctive alterations in the uptake of contrast agent into TDLNs, which could be useful to identify the correct TDLN, and to characterize TDLN lymphatic sinus growth that may predict metastatic potential.
Subject(s)
Gadolinium , Image Interpretation, Computer-Assisted/methods , Lymph Nodes/pathology , Magnetic Resonance Imaging/methods , Melanoma/pathology , Melanoma/secondary , Organometallic Compounds , Algorithms , Animals , Cell Line, Tumor , Contrast Media , Female , Image Enhancement/methods , Lymphatic Metastasis , Male , Mice , Mice, Inbred C57BL , Reproducibility of Results , Sensitivity and SpecificityABSTRACT
Metabolomics is one of the newcomers among the "omics" techniques, perhaps also constituting the most relevant for the study of pathophysiological conditions. Metabolomics may indeed yield not only disease-specific biomarkers but also profound insights into the etiology and progression of a variety of human disorders. Various metabolomic approaches are currently available to study oncogenesis and tumor progression in vivo, in murine tumor models. Many of these models rely on the xenograft of human cancer cells into immunocompromised mice. Understanding how the metabolism of these cells evolves in vivo is critical to evaluate the actual pertinence of xenograft models to human pathology. Here, we discuss various tumor xenograft models and methods for their metabolomic profiling to provide a short guide to investigators interested in this field of research.
Subject(s)
Metabolomics , Neoplasms, Experimental/metabolism , Animals , Biomarkers, Tumor/metabolism , Heterografts , Mice , Neoplasms, Experimental/pathology , Nuclear Magnetic Resonance, Biomolecular , Species Specificity , Tandem Mass SpectrometryABSTRACT
Metabolomics is increasingly being used in cancer biology for biomarker discovery and identification of potential novel therapeutic targets. However, a systematic metabolomics study of multiple biofluids to determine their interrelationships and to describe their use as tumor proxies is lacking. Using a mouse xenograft model of kidney cancer, characterized by subcapsular implantation of Caki-1 clear cell human kidney cancer cells, we examined tissue, serum, and urine all obtained simultaneously at baseline (urine) and at, or close to, animal sacrifice (urine, tissue, and plasma). Uniform metabolomics analysis of all three "matrices" was accomplished using gas chromatography- and liquid chromatography-mass spectrometry. Of all the metabolites identified (267 in tissue, 246 in serum, and 267 in urine), 89 were detected in all 3 matrices, and the majority was altered in the same direction. Heat maps of individual metabolites showed that alterations in serum were more closely related to tissue than was urine. Two metabolites, cinnamoylglycine and nicotinamide, were concordantly and significantly (when corrected for multiple testing) altered in tissue and serum, and cysteine-glutathione disulfide showed the highest change (232.4-fold in tissue) of any metabolite. On the basis of these and other considerations, three pathways were chosen for biologic validation of the metabolomic data, resulting in potential therapeutic target identification. These data show that serum metabolomics analysis is a more accurate proxy for tissue changes than urine and that tryptophan degradation (yielding anti-inflammatory metabolites) is highly represented in renal cell carcinoma, and support the concept that PPAR-α antagonism may be a potential therapeutic approach for this disease.
Subject(s)
Biomarkers, Tumor/analysis , Carcinoma, Renal Cell/metabolism , Kidney Neoplasms/metabolism , Metabolomics/methods , Animals , Cell Line, Tumor , Cell Proliferation , Chromatography, Liquid , Humans , Mass Spectrometry , Mice , Mice, Nude , PPAR alpha/pharmacology , Transplantation, Heterologous , Tryptophan/metabolism , Tumor Escape , Validation Studies as TopicABSTRACT
Kidney cancer often diagnosed at late stages when treatment options are severely limited. Thus, greater understanding of tumor metabolism leading ultimately to novel approaches to diagnosis is needed. Our laboratory has been utilizing metabolomics to evaluate compounds appearing in kidney cancer patients' biofluids at concentrations different from control patients. Here, we collected urine samples from kidney cancer patients and analyzed them by chromatography coupled to mass spectrometry. Once normalized to control for urinary concentration, samples were analyzed by two independent laboratories. After technical validation, we now show differential urinary concentrations of several acylcarnitines as a function of both cancer status and kidney cancer grade, with most acylcarnitines being increased in the urine of cancer patients and in those patients with high cancer grades. This finding was validated in a mouse xenograft model of human kidney cancer. Biological validation shows carbon chain length-dependent effects of the acylcarnitines on cytotoxicity in vitro, and higher chain length acylcarnitines demonstrated inhibitory effects on NF-κB activation, suggesting an immune modulatory effect of these compounds. Thus, acylcarnitines in the kidney cancer urine may reflect alterations in metabolism, cell component synthesis and/or immune surveillance, and may help explain the profound chemotherapy resistance seen with this cancer. This study shows for the first time the value of a novel class of metabolites which may lead to new therapeutic approaches for cancer and may prove useful in cancer biomarker studies. Furthermore, these findings open up a new area of investigation into the metabolic basis of kidney cancer.
Subject(s)
Carnitine/analogs & derivatives , Kidney Neoplasms/urine , NF-kappa B/biosynthesis , Animals , Biomarkers, Tumor/urine , Carnitine/biosynthesis , Carnitine/metabolism , Carnitine/urine , Cell Line, Tumor , Humans , Kidney , Kidney Neoplasms/pathology , Metabolomics , Mice , NF-kappa B/metabolism , Neoplasm Grading , Neoplasm Transplantation , Transplantation, HeterologousABSTRACT
Renal cell carcinoma (RCC) is one of the few human cancers whose incidence is increasing. The disease regularly progresses asymptomatically and is frequently metastatic upon presentation, thereby necessitating the development of an early method of detection. A metabolomic approach for biomarker detection using urine as a biofluid is appropriate since the tumor is located in close proximity to the urinary space. By comparing the composition of urine from individuals with RCC to control individuals, differences in metabolite composition of this biofluid can be identified, and these data can be utilized to create a clinically applicable and, possibly, bedside assay. Recent studies have shown that sample handling and processing greatly influences the variability seen in the urinary metabolome of both cancer and control patients. Once a standard method of collection is developed, identifying metabolic derangements associated with RCC will also lead to the investigation of novel targets for therapeutic intervention. The objective of this review is to discuss existing methods for sample collection, processing, data analysis, and recent findings in this emerging field.
Subject(s)
Biomarkers, Tumor/urine , Kidney Neoplasms/diagnosis , Kidney Neoplasms/urine , Metabolomics , HumansABSTRACT
Kidney cancer is the seventh most common cancer in the Western world, its incidence is increasing, and it is frequently metastatic at presentation, at which stage patient survival statistics are grim. In addition, there are no useful biofluid markers for this disease, such that diagnosis is dependent on imaging techniques that are not generally used for screening. In the present study, we use metabolomics techniques to identify metabolites in kidney cancer patients' urine, which appear at different levels (when normalized to account for urine volume and concentration) from the same metabolites in nonkidney cancer patients. We found that quinolinate, 4-hydroxybenzoate, and gentisate are differentially expressed at a false discovery rate of 0.26, and these metabolites are involved in common pathways of specific amino acid and energetic metabolism, consistent with high tumor protein breakdown and utilization, and the Warburg effect. When added to four different (three kidney cancer-derived and one "normal") cell lines, several of the significantly altered metabolites, quinolinate, α-ketoglutarate, and gentisate, showed increased or unchanged cell proliferation that was cell line-dependent. Further evaluation of the global metabolomics analysis, as well as confirmation of the specific potential biomarkers using a larger sample size, will lead to new avenues of kidney cancer diagnosis and therapy.