CRTC1/MAML2 directs a PGC-1α-IGF-1 circuit that confers vulnerability to PPARγ inhibition.

Mucoepidermoid carcinoma (MEC) is a life-threatening salivary gland cancer that is driven primarily by a transcriptional coactivator fusion composed of cyclic AMP-regulated transcriptional coactivator 1 (CRTC1) and mastermind-like 2 (MAML2). The mechanisms by which the chimeric CRTC1/MAML2 (C1/M2) oncoprotein rewires gene expression programs that promote tumorigenesis remain poorly understood. Here, we show that C1/M2 induces transcriptional activation of the non-canonical peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) splice variant PGC-1α4, which regulates peroxisome proliferator-activated receptor gamma (PPARγ)-mediated insulin-like growth factor 1 (IGF-1) expression. This mitogenic transcriptional circuitry is consistent across cell lines and primary tumors. C1/M2-positive tumors exhibit IGF-1 pathway activation, and small-molecule drug screens reveal that tumor cells harboring the fusion gene are selectively sensitive to IGF-1 receptor (IGF-1R) inhibition. Furthermore, this dependence on autocrine regulation of IGF-1 transcription renders MEC cells susceptible to PPARγ inhibition with inverse agonists. These results yield insights into the aberrant coregulatory functions of C1/M2 and identify a specific vulnerability that can be exploited for precision therapy.

Visual Function Decline Resulting from Geographic Atrophy: Results from the Chroma and Spectri Phase 3 Trials.

PURPOSE: To assess visual function outcomes to 48 weeks in patients with bilateral geographic atrophy (GA) secondary to age-related macular degeneration included in 2 interventional clinical trials: relationship to baseline lesion size, outcomes by baseline lesion characteristic subgroups, and correlation of visual function outcomes with GA area. DESIGN: The Chroma and Spectri studies (ClinicalTrials.gov identifiers, NCT02247479 and NCT02247531, respectively) were identically designed phase 3, double-masked, multicenter, randomized, sham injection-controlled clinical trials that evaluated intravitreal lampalizumab in GA. PARTICIPANTS: Eligible patients were 50 years of age or older with well-demarcated bilateral GA (lesion size, 1-7 disc areas) without evidence of or previous treatment for choroidal neovascularization in either eye and best-corrected visual acuity (BCVA) letter score of 49 letters or more (≥1 GA lesion within 250 µm of foveal center if BCVA ≥79 letters). METHODS: Patients (pooled n = 1881) were randomized 2:1:2:1 to lampalizumab every 4 weeks, sham every 4 weeks, lampalizumab every 6 weeks, or sham every 6 weeks. Sham arms were pooled for analysis. MAIN OUTCOME MEASURES: Functional end points included change in BCVA from baseline to week 48, low-luminance visual acuity, mesopic microperimetry (number of absolute scotomatous points, mean macular sensitivity), binocular and monocular maximum reading speed, and 2 validated patient-reported outcome measures: Functional Reading Independence Index and 25-item National Eye Institute Visual Function Questionnaire. RESULTS: Enlargement of GA area, approximately 2 mm2/year on average across all treatment groups in each study, was accompanied by overall deterioration in all functional end points. No statistically significant differences were found between lampalizumab or sham arms for changes from baseline in functional assessment scores. Of visual function tests, only microperimetry outcomes were correlated moderately with GA lesion area when assessed cross-sectionally at baseline and week 48. CONCLUSIONS: Chroma and Spectri provide a unique data set of functional end points in GA that are relevant for future clinical trials. Patients with bilateral GA experienced a consistent decline in visual function over 48 weeks, but measures of visual function were not correlated strongly with GA lesion area. It is not possible to predict visual function outcomes from GA lesion size.

Natural History of Geographic Atrophy Secondary to Age-Related Macular Degeneration: Results from the Prospective Proxima A and B Clinical Trials.

PURPOSE: To better characterize visual function decline and geographic atrophy (GA) progression secondary to age-related macular degeneration (AMD). DESIGN: Proxima A (NCT02479386)/Proxima B (NCT02399072) were global, prospective, noninterventional, observational clinical trials. PARTICIPANTS: Eligible patients were aged ≥50 years. Patients in Proxima A had bilateral GA without choroidal neovascularization (CNV) in either eye (N = 295). Patients in Proxima B had GA without CNV in the study eye and CNV±GA in the fellow eye (fellow eye CNV cohort, n = 168) or GA without CNV in the study eye, no CNV/GA in the fellow eye (fellow eye intermediate AMD cohort, n = 32). METHODS: Changes in visual function and imaging/anatomic parameters were evaluated over time using a mixed model for repeated measurement accounting for key baseline characteristics. MAIN OUTCOME MEASURES: Prespecified end points included change in GA area from baseline, best-corrected visual acuity (BCVA) score assessed by Early Treatment Diabetic Retinopathy Study (ETDRS), and visual acuity under low-luminance (LLVA). RESULTS: At 24 months, adjusted mean (standard error) change in GA lesion area from baseline was 3.87 (0.15) mm2 in participants with bilateral GA (Proxima A), 3.55 (0.16) mm2 in the fellow eye CNV cohort (Proxima B), and 2.96 (0.25) mm2 in the fellow eye intermediate AMD cohort (Proxima B). Progression of GA was greater in patients with baseline nonsubfoveal (vs. subfoveal) GA lesions and tended to increase as baseline low-luminance deficit increased (all patients). Conversion to GA or CNV in the fellow eye occurred in 30% and 6.7% of participants, respectively, in the Proxima B intermediate AMD cohort at month 12. Adjusted mean (standard error) changes in BCVA and LLVA (ETDRS letters) in the study eye from baseline to 24 months were -13.88 (1.40) and -7.64 (1.20) in Proxima A, -9.49 (1.29) and -7.57 (1.26) in Proxima B fellow eye CNV cohort, and -11.48 (3.39) and -8.37 (3.02) in Proxima B fellow eye intermediate AMD cohort, respectively. CONCLUSIONS: The prospective Proxima A and B studies highlight the severe functional impact of GA and the rapid rate of GA lesion progression over a 2-year period, including in patients with unilateral GA at baseline.

Engineered BRET-Based Biologic Light Sources Enable Spatiotemporal Control over Diverse Optogenetic Systems.

Light-inducible optogenetic systems offer precise spatiotemporal control over a myriad of biologic processes. Unfortunately, current systems are inherently limited by their dependence on external light sources for their activation. Further, the utility of laser/LED-based illumination strategies are often constrained by the need for invasive surgical procedures to deliver such devices and local heat production, photobleaching and phototoxicity that compromises cell and tissue viability. To overcome these limitations, we developed a novel BRET-activated optogenetics (BEACON) system that employs biologic light to control optogenetic tools. BEACON is driven by self-illuminating bioluminescent-fluorescent proteins that generate "spectrally tuned" biologic light via bioluminescence resonance energy transfer (BRET). Notably, BEACON robustly activates a variety of commonly used optogenetic systems in a spatially restricted fashion, and at physiologically relevant time scales, to levels that are achieved by conventional laser/LED light sources.

An Immunocompetent Mouse Model of HPV16(+) Head and Neck Squamous Cell Carcinoma.

The incidence of human papilloma virus (HPV)-associated head and neck squamous cell carcinoma (HNSCC) is increasing and implicated in more than 60% of all oropharyngeal carcinomas (OPSCCs). Although whole-genome, transcriptome, and proteome analyses have identified altered signaling pathways in HPV-induced HNSCCs, additional tools are needed to investigate the unique pathobiology of OPSCC. Herein, bioinformatics analyses of human HPV(+) HNSCCs revealed that all tumors express full-length E6 and identified molecular subtypes based on relative E6 and E7 expression levels. To recapitulate the levels, stoichiometric ratios, and anatomic location of E6/E7 expression, we generated a genetically engineered mouse model whereby balanced expression of E6/E7 is directed to the oropharyngeal epithelium. The addition of a mutant PIK3CAE545K allele leads to the rapid development of pre-malignant lesions marked by immune cell accumulation, and a subset of these lesions progress to OPSCC. This mouse provides a faithful immunocompetent model for testing treatments and investigating mechanisms of immunosuppression.

The Progression of Geographic Atrophy Secondary to Age-Related Macular Degeneration.

Geographic atrophy (GA) is an advanced form of age-related macular degeneration (AMD) that leads to progressive and irreversible loss of visual function. Geographic atrophy is defined by the presence of sharply demarcated atrophic lesions of the outer retina, resulting from loss of photoreceptors, retinal pigment epithelium (RPE), and underlying choriocapillaris. These lesions typically appear first in the perifoveal macula, initially sparing the foveal center, and over time often expand and coalesce to include the fovea. Although the kinetics of GA progression are highly variable among individual patients, a growing body of evidence suggests that specific characteristics may be important in predicting disease progression and outcomes. This review synthesizes current understanding of GA progression in AMD and the factors known or postulated to be relevant to GA lesion enlargement, including both affected and fellow eye characteristics. In addition, the roles of genetic, environmental, and demographic factors in GA lesion enlargement are discussed. Overall, GA progression rates reported in the literature for total study populations range from 0.53 to 2.6 mm2/year (median, ∼1.78 mm2/year), assessed primarily by color fundus photography or fundus autofluorescence (FAF) imaging. Several factors that could inform an individual's disease prognosis have been replicated in multiple cohorts: baseline lesion size, lesion location, multifocality, FAF patterns, and fellow eye status. Because best-corrected visual acuity does not correspond directly to GA lesion enlargement due to possible foveal sparing, alternative assessments are being explored to capture the relationship between anatomic progression and visual function decline, including microperimetry, low-luminance visual acuity, reading speed assessments, and patient-reported outcomes. Understanding GA progression and its individual variability is critical in the design of clinical studies, in the interpretation and application of clinical trial results, and for counseling patients on how disease progression may affect their individual prognosis.

Targeting factor D of the alternative complement pathway reduces geographic atrophy progression secondary to age-related macular degeneration.

Geographic atrophy is an advanced form of age-related macular degeneration (AMD) and a leading cause of vision loss for which there are no approved treatments. Genetic studies in AMD patients have implicated dysregulation of the alternative complement pathway in the pathogenesis of geographic atrophy. Lampalizumab is a potential therapeutic that targets complement factor D, a pivotal activator of the alternative complement pathway. The MAHALO phase 2 clinical trial was a multicenter, randomized, controlled study that evaluated lampalizumab administered by intravitreal injection monthly (n = 42) and every other month (n = 41) versus sham control (n = 40) in patients with geographic atrophy secondary to AMD. The primary endpoint was the mean change in lesion area from baseline to month 18 as measured by fundus autofluorescence. Specific AMD-associated genetic polymorphisms were also analyzed. The MAHALO study met its primary efficacy endpoint with an acceptable safety profile; monthly lampalizumab treatment demonstrated a 20% reduction in lesion area progression versus sham control [80% confidence interval (CI), 4 to 37%]. A more substantial monthly treatment benefit of 44% reduction in geographic atrophy area progression versus sham control (95% CI, 15 to 73%) was observed in a subgroup of complement factor I (CFI) risk-allele carriers (57% of the patients analyzed were CFI risk-allele carriers). The MAHALO study shows a potential treatment effect in patients with geographic atrophy and supports therapeutic targeting of the alternative complement pathway for treating AMD pathogenesis.

THE PATHOPHYSIOLOGY OF GEOGRAPHIC ATROPHY SECONDARY TO AGE-RELATED MACULAR DEGENERATION AND THE COMPLEMENT PATHWAY AS A THERAPEUTIC TARGET.

PURPOSE: Geographic atrophy (GA) is an advanced, vision-threatening form of age-related macular degeneration (AMD) affecting approximately five million individuals worldwide. To date, there are no approved therapeutics for GA treatment; however, several are in clinical trials. This review focuses on the pathophysiology of GA, particularly the role of complement cascade dysregulation and emerging therapies targeting the complement cascade. METHODS: Primary literature search on PubMed for GA, complement cascade in age-related macular degeneration. ClinicalTrials.gov was searched for natural history studies in GA and clinical trials of drugs targeting the complement cascade for GA. RESULTS: Cumulative damage to the retina by aging, environmental stress, and other factors triggers inflammation via multiple pathways, including the complement cascade. When regulatory components in these pathways are compromised, as with several GA-linked genetic risk factors in the complement cascade, chronic inflammation can ultimately lead to the retinal cell death characteristic of GA. Complement inhibition has been identified as a key candidate for therapeutic intervention, and drugs targeting the complement pathway are currently in clinical trials. CONCLUSION: The complement cascade is a strategic target for GA therapy. Further research, including on natural history and genetics, is crucial to expand the understanding of GA pathophysiology and identify effective therapeutic targets.

CLINICAL ENDPOINTS FOR THE STUDY OF GEOGRAPHIC ATROPHY SECONDARY TO AGE-RELATED MACULAR DEGENERATION.

PURPOSE: To summarize the recent literature describing the application of modern technologies in the study of patients with geographic atrophy (GA) secondary to age-related macular degeneration. METHODS: Review of the literature describing the terms and definitions used to describe GA, imaging modalities used to capture and measure GA, and the tests of visual function and functional deficits that occur in patients with GA. RESULTS: In this paper, we describe the evolution of the definitions used to describe GA. We compare imaging modalities used in the characterization of GA, report on the sensitivity and specificity of the techniques where data exist, and describe the correlations between these various modes of capturing the presence of GA. We review the functional tests that have been used in patients with GA, and critically examine their ability to detect and quantify visual deficits. CONCLUSION: Ophthalmologists and retina specialists now have a wide range of assessments available for the functional and anatomic characterization of GA in patients with age-related macular degeneration. To date, studies have been limited by their unimodal approach, and we recommend that future studies of GA use multimodal imaging. We also suggest strategies for the optimal functional testing of patients with GA.

An observational retrospective subgroup analysis of verteporfin photodynamic therapy-naive and previously treated patients in the focus trial.

PURPOSE: To perform a retrospective post hoc subgroup analysis of the FOCUS trial to assess the visual acuity outcomes and treatment benefits for patients receiving combination therapy who, at the time of enrollment, were naive to verteporfin photodynamic therapy (PDT) or had previously received PDT. METHODS: In this retrospective post hoc analysis of 24-month data from the FOCUS trial, PDT-naive and previously PDT-treated patients (n = 162) were included. Patients were randomized in a 2:1 ratio to receive 0.5 mg of ranibizumab monthly plus PDT or PDT alone. We retrospectively identified patients who had or had not received prior PDT for a post hoc subgroup analysis of 12- and 24-month outcomes. RESULTS: For the PDT-naive patients, mean change in the visual acuity at 24 months was +4.1 letters for the ranibizumab plus PDT group and -11.5 letters for the PDT monotherapy group, a treatment benefit over control group of 15.6 letters (95% confidence interval: 7.1-24.2). For the previously treated patients, mean change in the visual acuity at 24 months was +5.2 letters for the ranibizumab plus PDT group and -4.3 letters for the PDT monotherapy group, a treatment benefit over control group of 9.5 letters (95% confidence interval: 2.3-16.8). CONCLUSION: In the FOCUS subanalysis, the PDT-naive patients showed a trend toward greater treatment benefit over control subjects compared with patients previously treated with PDT. However, this study was not designed to address this question, and the confidence intervals were wide. Furthermore, the mean change in the visual acuity from baseline to 24 months was similar for both the PDT-naive and previously treated patients receiving combination therapy.

Central projections of trigeminal afferents innervating the face in naked mole-rats (Heterocephalus glaber).

In this study, we examined the topography of projections from facial afferents to the trigeminal brainstem nuclear complex (TBNC) in naked mole-rats using the neuronal tracer CTB-HRP. Tracer injections were made in a ventral to dorsal sequence that included the tooth pulp and dental ligament, ventral buccal pad, vibrissae, and the forehead. Labeled terminals were identified throughout the rostrocaudal extent of the TBNC, including the principal nucleus (Pr5), pars oralis (Sp5O), pars interpolaris (Sp5I), and pars caudalis (Sp5C) of the spinal trigeminal nucleus. Injections that labeled afferents from the tooth pulp and dental ligament resulted in heavy transport to dorsomedial portions of the TBNC, whereas injections made into progressively more dorsal regions of the face resulted in labeled terminals progressively more ventral and lateral in the nuclei. Injections that included dental afferents also labeled the mesencephalic nucleus of V, whereas injections into the skin of the face labeled cell bodies in the facial nucleus, and in most cases the motor nucleus of 5. Dental afferents in more rostral portions of the TBNC were coextensive with a cytochrome oxidase-dense region visible in alternate sections processed for chemoarchitecture.

Growing out of a caste--reproduction and the making of the queen mole-rat.

Naked mole-rats have a eusocial colony structure consisting of non-reproductive workers and a reproductively active caste where a single, dominant queen and 1-3 males produce all of the offspring. Well-established queens have elongated bodies that characterize their caste. Worker females retain the ability to transform into queens, however the trigger and time course for this physical transformation remain a mystery. Here, we show a direct link between periods of pregnancy and vertebral lengthening in nascent queens. Adult female mole-rats were paired with a male and radiographed weekly for two and a half years to track the growth of the lumbar vertebrae as the mole-rats became sexually mature and experienced pregnancies. The lumbar vertebrae of breeding females grew at an increased rate during each pregnancy but growth rates returned to normal between pregnancies and during extended periods without reproduction. The rate of lumbar lengthening was reduced to normal rates in older, established queens experiencing pregnancies. Our results imply that the length of a new queen mole-rat is proportional to the number of pregnancies experienced and suggest that hormones related to pregnancy may play the critical role in bone growth associated with caste transformation.

Touching on somatosensory specializations in mammals.

Specialized species often reveal general principles of brain organization and provide systems for analysis of sensory function. Subterranean species dependent on touch have particularly large somatosensory areas with modular cortical representations of sensory surfaces. Some species have added cortical areas to processing networks, have developed tactile foveas and have superior colliculi primarily devoted to somatosensation rather than vision. Recent studies reveal surprisingly large cortical representations of oral structures in primates and mole-rats. Cortical modules represent a range of different sensory surfaces in rodents, star-nosed moles and primates, indicating that similar developmental mechanisms operate in diverse species. Finally, manipulation of patterning genes in mice suggests evolutionary mechanisms for producing the specialized corticies of subterranean species.

Cortical, callosal, and thalamic connections from primary somatosensory cortex in the naked mole-rat (Heterocephalus glaber), with special emphasis on the connectivity of the incisor representation.

We investigated the distribution of cortical, callosal, and thalamic connections from the primary somatosensory area (S1) in naked mole-rats, concentrating on lower incisor and forelimb representations. A neuronal tracer (WGA-HRP) was injected into the center of each respective representation under guidance from microelectrode recordings of neuronal activity. The locations of cells and terminals were determined by aligning plots of labeled cells with flattened cortical sections reacted for cytochrome oxidase. The S1 lower incisor area was found to have locally confined intrahemispheric connections and longer connections to a small cluster of cells in the presumptive secondary somatosensory (S2) and parietal ventral (PV) incisor fields. The S1 incisor area also had sparse connections with anterior cortex, in presumptive primary motor cortex. Homotopic callosal projections were identified between the S1 lower incisor areas in each hemisphere. Thalamocortical connections related to the incisor were confined to ventromedial portions of the ventral posterior medial subnucleus (VPM) and posterior medial nucleus (Po). Injections into the S1 forelimb area revealed reciprocal intrahemispheric connections to S2 and PV, to two areas in frontal cortex, and to two areas posterior to S1 that appear homologous to posterior lateral area and posterior medial area in rats. The S1 forelimb representation also had callosal projections to the contralateral S1 limb area and to contralateral S2 and PV. Thalamic distribution of label from forelimb injections included ventral portions of the ventral posterior lateral subnucleus (VPL), dorsolateral Po, the ventral lateral nucleus, and the ventral medial nucleus and neighboring intralaminar nuclei.

Organization of somatosensory cortical areas in the naked mole-rat (Heterocephalus glaber).

Multiunit electrophysiology was combined with histological analysis of cortical sections to investigate the organization of somatosensory areas in the naked mole-rat. We provide new details for the organization of primary somatosensory cortex (S1) and identify cortical modules and barrels that correspond to the representations of different body parts. In addition, details of the location and organization of secondary somatosensory cortex (S2) are reported, and evidence for a third somatosensory representation, likely the parietal ventral area (PV), is provided and discussed. S1 contained a complete and systematic representation of the contralateral body surface and oral structures. The orientation of S1 was inverted, with the lower body represented medially and the face and oral structures located rostrolaterally. The S2 representation was found in caudolateral cortex forming a mirror image of S1. The two areas were joined at the representation of the vibrissae and snout, so that the orientation of S2 formed an upright representation of the body in cortex. Receptive fields for S2 were consistently larger than those in S1. Evidence for the presumptive parietal ventral area, lateral to S2, suggests that this area may be an inverted mirror image of S2. By aligning the electrophysiological maps of body representations with cytochrome oxidase-reacted cortical sections we were able to identify modules related to the buccal pad, chin, vibrissae, forelimb, hindlimb, trunk, tongue, lower incisor, and upper incisors. The orofacial modules in lateral cortex resemble similar modules reported to relate to oral structures previously described in the laboratory rat, owl monkey, and squirrel monkey.

Plasticity of the cortical dentition representation after tooth extraction in naked mole-rats.

Naked mole-rats (Heterocephalus glaber) have a large cortical representation of their behaviorally important front teeth, accounting for 30% of primary somatosensory cortex (SI). Here we investigated the plasticity of this dental representation after the extraction of a single lower tooth. The representation of the contralateral lower incisor normally accounts for approximately 15% of somatosensory cortex in mole-rats. In five mole-rats the lower right incisor was extracted on either postnatal day 7 or 21. After 5-8 months the deprived tooth zone in S1 was investigated with multiunit microelectrode recordings. The results revealed a dramatic reorganization of the orofacial representation in SI. Neurons in the cortical lower tooth representation were responsive to tactile inputs from surrounding orofacial structures, including the contralateral upper incisor, ipsilateral lower incisor, tongue, chin, gums, and buccal pad. Neurons in the former lower tooth zone had complex receptive fields that often encompassed multiple sensory surfaces surrounding the extracted tooth in the periphery. These results suggest that the representation of the dentition in mammals is capable of significant reorganization after the loss of sensory inputs from the teeth. These data parallel findings in the somatosensory hand area of primates after deafferentation where cortex can become activated by a mixture of widely spaced surrounding sensory surfaces (e.g., chin and upper arm).

Organization of somatosensory cortex in the laboratory rat (Rattus norvegicus): Evidence for two lateral areas joined at the representation of the teeth.

Lateral somatosensory areas have not been explored in detail in rats, and theories on the organization of this region are based largely on anatomical tracing experiments. We investigated the topography of this region by using microelectrode recordings, which were related to flattened cortical sections processed for cytochrome oxidase (CO). Two lateral somatosensory areas were identified, each containing a complete representation of the body. A larger, more medial representation formed a mirror image of S1 along the rostrocaudal axis of the head region corresponding to the previously identified secondary somatosensory area (S2). A smaller, more lateral representation formed a mirror image of S2 along the rostrocaudal axis of the forelimb and hindlimb regions and likely corresponds to the parietal ventral area (PV) identified in other mammals. We also investigated the representation of the dentition and identified regions of cortex responsive to tooth stimulation. The lower incisor representation was rostral to the lower lip region of S1, and the upper incisor representation was lateral to the buccal pad region of S1. The upper and lower incisors flanked the tongue representation. An additional large region of far lateral cortex responded to both incisors. Finally, five CO-dense modules were consistently identified rostral and lateral to the S1 face representation, which we refer to as OM1, OM2, OM3, FM, and HM. These modules closely correspond to the physiologically identified areas representing the lower incisor (OM1) and tongue (OM2) regions of S1 and the mixed tooth (OM3), forelimb (FM1), and hindlimb (HM) representations of S2 and PV.