Hallmarks of response, resistance, and toxicity to immune checkpoint blockade.

Unprecedented advances have been made in cancer treatment with the use of immune checkpoint blockade (ICB). However, responses are limited to a subset of patients, and immune-related adverse events (irAEs) can be problematic, requiring treatment discontinuation. Iterative insights into factors intrinsic and extrinsic to the host that impact ICB response and toxicity are critically needed. Our understanding of the impact of host-intrinsic factors (such as the host genome, epigenome, and immunity) has evolved substantially over the past decade, with greater insights on these factors and on tumor and immune co-evolution. Additionally, we are beginning to understand the impact of acute and cumulative exposures-both internal and external to the host (i.e., the exposome)-on host physiology and response to treatment. Together these represent the current day hallmarks of response, resistance, and toxicity to ICB. Opportunities built on these hallmarks are duly warranted.

Targeting PD-L2-RGMb overcomes microbiome-related immunotherapy resistance.

The gut microbiota is a crucial regulator of anti-tumour immunity during immune checkpoint inhibitor therapy. Several bacteria that promote an anti-tumour response to immune checkpoint inhibitors have been identified in mice1-6. Moreover, transplantation of faecal specimens from responders can improve the efficacy of anti-PD-1 therapy in patients with melanoma7,8. However, the increased efficacy from faecal transplants is variable and how gut bacteria promote anti-tumour immunity remains unclear. Here we show that the gut microbiome downregulates PD-L2 expression and its binding partner repulsive guidance molecule b (RGMb) to promote anti-tumour immunity and identify bacterial species that mediate this effect. PD-L1 and PD-L2 share PD-1 as a binding partner, but PD-L2 can also bind RGMb. We demonstrate that blockade of PD-L2-RGMb interactions can overcome microbiome-dependent resistance to PD-1 pathway inhibitors. Antibody-mediated blockade of the PD-L2-RGMb pathway or conditional deletion of RGMb in T cells combined with an anti-PD-1 or anti-PD-L1 antibody promotes anti-tumour responses in multiple mouse tumour models that do not respond to anti-PD-1 or anti-PD-L1 alone (germ-free mice, antibiotic-treated mice and even mice colonized with stool samples from a patient who did not respond to treatment). These studies identify downregulation of the PD-L2-RGMb pathway as a specific mechanism by which the gut microbiota can promote responses to PD-1 checkpoint blockade. The results also define a potentially effective immunological strategy for treating patients who do not respond to PD-1 cancer immunotherapy.

Androgen receptor blockade promotes response to BRAF/MEK-targeted therapy.

Treatment with therapy targeting BRAF and MEK (BRAF/MEK) has revolutionized care in melanoma and other cancers; however, therapeutic resistance is common and innovative treatment strategies are needed1,2. Here we studied a group of patients with melanoma who were treated with neoadjuvant BRAF/MEK-targeted therapy ( NCT02231775 , n = 51) and observed significantly higher rates of major pathological response (MPR; ≤10% viable tumour at resection) and improved recurrence-free survival (RFS) in female versus male patients (MPR, 66% versus 14%, P = 0.001; RFS, 64% versus 32% at 2 years, P = 0.021). The findings were validated in several additional cohorts2-4 of patients with unresectable metastatic melanoma who were treated with BRAF- and/or MEK-targeted therapy (n = 664 patients in total), demonstrating improved progression-free survival and overall survival in female versus male patients in several of these studies. Studies in preclinical models demonstrated significantly impaired anti-tumour activity in male versus female mice after BRAF/MEK-targeted therapy (P = 0.006), with significantly higher expression of the androgen receptor in tumours of male and female BRAF/MEK-treated mice versus the control (P = 0.0006 and P = 0.0025). Pharmacological inhibition of androgen receptor signalling improved responses to BRAF/MEK-targeted therapy in male and female mice (P = 0.018 and P = 0.003), whereas induction of androgen receptor signalling (through testosterone administration) was associated with a significantly impaired response to BRAF/MEK-targeted therapy in male and female patients (P = 0.021 and P < 0.0001). Together, these results have important implications for therapy.

A facile magnetic extrusion method for preparing endosome-derived vesicles for cancer drug delivery.

To date, the scaled-up manufacturing and efficient drug loading of exosomes are two existing challenges limiting the clinical translation of exosome-based drug delivery. Herein, we developed a facile magnetic extrusion method for preparing endosome-derived vesicles, also known as exosome mimetics (EMs), which share the same biological origin and similar morphology, composition, and biofunctions with native exosomes. The high yield and consistency of this magnetic extrusion method help to overcome the manufacturing bottleneck in exosome research. Moreover, the proposed standardized multi-step method readily facilitates the ammonium sulfate gradient approach to actively load chemodrugs such as doxorubicin into EMs. The engineered EMs developed and tested here exhibit comparable drug delivery properties as do native exosomes and potently inhibit tumor growth by delivering doxorubicin in an orthotopic breast tumor model. These findings demonstrate that EMs can be prepared in a facile and scaled-up manner as a promising biological nanomedicine for cancer drug delivery.

Cdc42-Dependent Transfer of mir301 from Breast Cancer-Derived Extracellular Vesicles Regulates the Matrix Modulating Ability of Astrocytes at the Blood-Brain Barrier.

Breast cancer brain metastasis is a major clinical challenge and is associated with a dismal prognosis. Understanding the mechanisms underlying the early stages of brain metastasis can provide opportunities to develop efficient diagnostics and therapeutics for this significant clinical challenge. We have previously reported that breast cancer-derived extracellular vesicles (EVs) breach the blood-brain barrier (BBB) via transcytosis and can promote brain metastasis. Here, we elucidate the functional consequences of EV transport across the BBB. We demonstrate that brain metastasis-promoting EVs can be internalized by astrocytes and modulate the behavior of these cells to promote extracellular matrix remodeling in vivo. We have identified protein and miRNA signatures in these EVs that can lead to the interaction of EVs with astrocytes and, as such, have the potential to serve as targets for development of diagnostics and therapeutics for early detection and therapeutic intervention in breast cancer brain metastasis.

Craniofacial morphologic parameters in a Persian population: an anthropometric study.

Limited data are available regarding the reference ranges of facial proportions of the Persian population in Iran. This study aimed to establish the reference range of craniofacial anthropometric measurements in an adult Iranian population. On 100 individuals (men = women), aged 18 to 30 years with normal faces and occlusions, 34 linear and 7 angular measurements as well as 24 indices were calculated. The difference of measurements between men and women were evaluated by paired t-test. The data were compared with the norms of North American whites using 1-sample t-test. The subjects belonged to 5 ethnic groups (57% from Fars, 14% from Kord, 11% from Azari, 10% from Gilaki-Mazani, and 2% from Lor). All head measurements were greater in men except for the head index and the head height. The subjects had leptoprosopic faces. The intercanthal width was almost one third of the biocular width and greater than the eye fissure length. Although the nose width of women was significantly smaller, both sexes had leptorrhine noses. The chin height and lower chin height were greater in men. In comparison with North American whites, considerable differences were found regarding head height and width, biocular width, nose height, face height, mouth width, and upper chin height. In conclusion, the reference range of craniofacial anthropometric measurements established for the Iranian population might be efficiently used for esthetic treatments.

Stabilization of premaxilla repositioned during secondary bone grafting in complete bilateral cleft lip and palate patients.

Secondary bone grafting simultaneous to premaxillary repositioning is a well-recognized surgical procedure for the management of bilateral cleft lip and palate patients. Proper stabilization of the repositioned premaxilla is considered as a key factor for the success of secondary bone grafting because the mobility of the premaxillary segment jeopardizes graft integration. This case series reports a reliable method of premaxillary stabilization that incorporated the intrasurgical application of resin bone cement to cover and reinforce the arch bars or orthodontic brackets applied on the maxillary teeth. Occlusal loads were reduced by application of posterior bite blocks on the mandibular teeth. The stabilization method was performed on 7 patients (5 women and 2 men) with a mean age of 12.4 years. During postsurgery follow-ups, the repositioned premaxillary segments did not show mobility in any of the patients. The palatal fistulae were completely closed. Panoramic radiographies taken 2 months after surgery demonstrated acceptable graft integration. The patients have now been followed up to 5 years. No evidence of relapse has been observed. This technique seemed to be undemanding, included minimal laboratory procedure, and maintained the labial mucosa overlying the repositioned segment intact.

Thickness of labial alveolar bone overlying healthy maxillary and mandibular anterior teeth.

AIM: This study aimed to measure the thickness of labial bone overlying maxillary and mandibular anterior teeth and the distance between cementoenamel junction and bone crest in a Persian population. MATERIALS & METHODS: Two calibrated examiners evaluated tomographic data of 152 maxillary and 200 mandibular anterior teeth. Labial bone width was assessed at levels 1.0 to 5.0 mm apical to bone crest. Moreover, the distance between cementoenamel junction and bone crest was measured for both maxillary and mandibular teeth and its potential effect on the amount of labial bone thickness was assessed. RESULTS: One hundred-twenty nine maxillary central incisors, 77 lateral incisors, 70 canines, 105 mandibular central incisors, 103 lateral incisors and 81 canines were included for measurements. In maxilla, width of bone averaged 1.08mm, 1.11mm, and 1.3mm for central incisors, lateral incisors, and canines, respectively. Corresponding numbers for mandibular central incisors, lateral incisors, and canines were 0.74mm, 0.66mm and 0.40mm. High variation of cementoenamel junction to bone crest distance (range 0.5 to 5.15 mm) was detected. The mean amount of labial bone width was not statistically different in patients with different distances between cementoenamel junction and bone crest; except for mandibular lateral incisors. CONCLUSION: The mean thickness of the labial alveolar bone overlying maxillary anterior teeth was found to be between 1 to 1.2 mm and between 0.5 to 0.8 mm for mandibular anterior teeth at the first 5 mm from bone crest in a Persian population.

Multidisciplinary reconstruction of a palatomaxillary defect with nonvascularized fibula bone graft and distraction osteogenesis.

Reconstruction of extensive palatomaxillary defects with oronasal/antral communication represents a challenge to surgeons. Bone-containing microvascular flaps have been suggested as a promising option for rehabilitation of function and esthetics. These types of flaps, however, might be associated with high morbidity. A combination of less complicated treatments might also provide acceptable results while diminishing potential donor site complications. This clinical report presents a bilateral maxillary defect with oronasal communication due to resection of malignant melanoma of the palate. The lost alveolar bone was initially reconstructed with a nonvascularized fibula bone graft. After 6 months, the alveolar segment was subjected to vertical distraction osteogenesis to increase bone height. After a 3-month consolidation period, the patient received 10 dental implants and an implant-supported fixed prosthesis. To preclude graft harvesting morbidity for reconstructing the oronasal fistula, the frame of the prosthesis was designed to include 3 ball attachments on which a palatal obturator, merely covering the palate, could be stabilized. The removable implant-retained obturator restored function perfectly. During the 5-year follow-up, no complication regarding bone graft, the dental implants, and the obturator has been observed.

Healing of extraction sockets and augmented alveolar defects following 1-year treatment with bisphosphonate.

To assess the effect of bisphosphonates on healing of extraction sockets and augmented alveolar defects, 12 adult female mongrel dogs were assigned to 2 experimental groups and a control group. The experimental groups received oral alendronate (ALN, 3.5 mg/kg/wk) or IV pamidronate (PAM, 1 mg/kg/wk) for 12 months. Animals were randomly tested for serum C-terminal telopeptide of collagen I (CTx). The right first and second premolars were extracted. After 8 weeks, extraction sites were evaluated for healing. Subsequently, 3-wall defects were created in ridges and filled with human mineralized cortical particulate bone. Two months post-augmentation, animals were sacrificed and mandibles were collected for cone-beam computed tomography (CBCT) and histomorphometric appraisal. The obtained data were compared using 1-way ANOVA test. CTx test results in both experimental groups were comparable (<10 pg/mL) but lower than that of the control group (minimum 159.2 pg/mL). Two months post-extraction, bone sequestra were noticed in extraction sites in BP-treated groups, involving the entire alveolar bone in the PAM group and the upper rim of the alveoli in the ALN group. Histologically, bone sequestra from the PAM group demonstrated empty osteocyte lacunae, while in the ALN group areas of necrotic bone along with evidence of active bone remodeling was distinguished. Eight weeks post-augmentation, the experimental groups showed no evidence of bone formation in the augmented area, while bone formation ratio was measured to be 18.32% in the control group. The mean amount of pixel intensity calculated from the CBCT images of the ALN, PAM, and control group was 113.69 ± 11.04, 124.94 ± 4.72, and 113.69 ± 6.63, respectively. Pixel intensity in PAM-treated group was significantly higher than both other groups. This study demonstrated that 1-year treatment with ALN/PAM was associated with impairment of post-extraction and post-augmentation bone healing in dogs.

Cortical tenting technique versus onlay layered technique for vertical augmentation of atrophic posterior mandibles: a split-mouth pilot study.

PURPOSE: This study intended to compare the efficacy of onlay layered and cortical tenting grafting techniques for vertical alveolar augmentation. MATERIALS AND METHODS: Six patients with bilaterally edentulous atrophic posterior mandibles received 2 thin ramus blocks, covered with bovine bone mineral on 1 side (onlay layered technique), whereas in the opposite side (cortical tenting technique), a thin ramus block was placed over the ridge keeping a distance that was filled with particulate autogenous bone and bovine bone mineral (50:50). Bone height over the mandibular canal was measured on panoramic radiographies taken preoperatively, and 2 weeks and 4 months postoperatively. RESULTS: The amount of augmentation in the tenting group (5.2 ± 0.76 mm) was higher than the onlay group (4.48 ± 0.51 mm). Graft resorption was 1.75 ± 1.08 mm in the onlay group and 1.17 ± 0.41 mm in the tenting group. Neither of the differences regarding the amount of augmentation and graft resorption was statistically significant (P = 0.345 and 0831, respectively). CONCLUSION: Both of the examined techniques might be favorable methods for vertical augmentation.

Clinical importance of recipient site characteristics for vertical ridge augmentation: a systematic review of literature and proposal of a classification.

This review evaluated the characteristics of vertical alveolar defects that were augmented via onlay bone grafting or guided bone regeneration. Information regarding the anatomic site, type of edentulism, and defects' dimensions were extracted. The experiments differed vastly in the description of the defects' features. Aiming to mitigate the confounding effect of recipient site's morphology in future experiments, a classification of vertically deficient recipient sites is proposed.

Vertical bone augmentation with simultaneous implant placement using particulate mineralized bone and mesenchymal stem cells: a preliminary study in rabbit.

This study aimed to assess vertical bone augmentation with simultaneous implant placement in rabbit tibiae using particulate mineralized bone/fibrin glue/mesenchymal stem cell. Bone marrow was aspirated from tibiae of five 10-week-old New Zealand White male rabbits. Right and left tibiae of each rabbit were prepared, and a 3-mm protruding implant from tibial bone was placed in each side. Particulate allogenic bone/fibrin glue/mesenchymal stem cell combination was placed around test implants and particulate bone graft/fibrin glue around controls. Two months postoperatively, the animals were euthanized, and sections were prepared for histological analysis. The mean amount of vertical bone length was higher in the experimental group than the control group (2.09 mm vs 1.03 mm; P < .05). New supracrestal trabecular bone formation was also significantly higher in the test group (28.5 ± 4.5% vs 4.3 ± 1.8%; P < .05). Mesenchymal stem cell/particulate allograft/fibrin glue appears to be a promising combination for vertical bone augmentation around simultaneously inserted implants in rabbit tibia.

Dissecting the Impact of the Gut Microbiome on Cancer Immunotherapy.

The gut microbiome has emerged as a key regulator of response to cancer immunotherapy. However, there is a gap in our understanding of the underlying mechanisms by which the microbiome influences immunotherapy. To this end, we developed a mathematical model based on i) gut microbiome data derived from preclinical studies on melanomas after fecal microbiota transplant, ii) mechanistic modeling of antitumor immune response, and iii) robust association analysis of murine and human microbiome profiles with model-predicted immune profiles. Using our model, we could distill the complexity of these murine and human studies on microbiome modulation in terms of just two model parameters: the activation and killing rate constants of immune cells. We further investigated associations between specific bacterial taxonomies and antitumor immunity and immunotherapy efficacy. This model can guide the design of studies to refine and validate mechanistic links between the microbiome and immune system.

The role of extracellular vesicles in the physiological and pathological regulation of the blood-brain barrier.

Extracellular vesicles (EVs) are a subclass of biological nanoparticles secreted by most cell types. Once secreted, EVs can travel long distances to deliver their content to target cells thereby playing a key role in cell-to-cell communication and supporting both physiological and pathological processes. In recent years, the functional versatility of EVs has come to be more widely appreciated. Their heterogeneous structure encloses solubilized bioactive cargoes including proteins and nucleic acids. EVs mirror the secreting cell in composition therefore representing a novel source of diagnostic and prognostic biomarkers. Moreover, due to their unique structure, EVs constitute a promising class of biocompatible nanovehicles for drug delivery as well. Importantly, and of burgeoning interest, is the fact that EVs have the intrinsic ability to breach biological barriers including the complex blood-brain barrier (BBB), whose restrictive nature represents a significant therapeutic challenge. EVs have been shown to contribute to the progression of a variety of brain diseases including metastatic brain cancer, neurodegenerative diseases, and acute pathologies including infections and ischemia. In this review, the role of EVs in the maintenance and regulation of the BBB under normal physiological and pathologic conditions are discussed. Applications of EVs as therapeutic and diagnostic tools in the treatment of diseases that affect the central nervous system are presented as are limitations hindering their broad translation and potential solutions to resolve them.

Gut microbiota signatures are associated with toxicity to combined CTLA-4 and PD-1 blockade.

Treatment with combined immune checkpoint blockade (CICB) targeting CTLA-4 and PD-1 is associated with clinical benefit across tumor types, but also a high rate of immune-related adverse events. Insights into biomarkers and mechanisms of response and toxicity to CICB are needed. To address this, we profiled the blood, tumor and gut microbiome of 77 patients with advanced melanoma treated with CICB, with a high rate of any ≥grade 3 immune-related adverse events (49%) with parallel studies in pre-clinical models. Tumor-associated immune and genomic biomarkers of response to CICB were similar to those identified for ICB monotherapy, and toxicity from CICB was associated with a more diverse peripheral T-cell repertoire. Profiling of gut microbiota demonstrated a significantly higher abundance of Bacteroides intestinalis in patients with toxicity, with upregulation of mucosal IL-1ß in patient samples of colitis and in pre-clinical models. Together, these data offer potential new therapeutic angles for targeting toxicity to CICB.

Dietary fiber and probiotics influence the gut microbiome and melanoma immunotherapy response.

Gut bacteria modulate the response to immune checkpoint blockade (ICB) treatment in cancer, but the effect of diet and supplements on this interaction is not well studied. We assessed fecal microbiota profiles, dietary habits, and commercially available probiotic supplement use in melanoma patients and performed parallel preclinical studies. Higher dietary fiber was associated with significantly improved progression-free survival in 128 patients on ICB, with the most pronounced benefit observed in patients with sufficient dietary fiber intake and no probiotic use. Findings were recapitulated in preclinical models, which demonstrated impaired treatment response to anti­programmed cell death 1 (anti­PD-1)­based therapy in mice receiving a low-fiber diet or probiotics, with a lower frequency of interferon-γ­positive cytotoxic T cells in the tumor microenvironment. Together, these data have clinical implications for patients receiving ICB for cancer.