Deep Domain Adversarial Learning for Species-Agnostic Classification of Histologic Subtypes of Osteosarcoma.

Osteosarcomas (OSs) are aggressive bone tumors with many divergent histologic patterns. During pathology review, OSs are subtyped based on the predominant histologic pattern; however, tumors often demonstrate multiple patterns. This high tumor heterogeneity coupled with scarcity of samples compared with other tumor types render histology-based prognosis of OSs challenging. To combat lower case numbers in humans, dogs with spontaneous OSs have been suggested as a model species. Herein, a convolutional neural network was adversarially trained to classify distinct histologic patterns of OS in humans using mostly canine OS data during training. Adversarial training improved domain adaption of a histologic subtype classifier from canines to humans, achieving an average multiclass F1 score of 0.77 (95% CI, 0.74-0.79) and 0.80 (95% CI, 0.78-0.81) when compared with the ground truth in canines and humans, respectively. Finally, this trained model, when used to characterize the histologic landscape of 306 canine OSs, uncovered distinct clusters with markedly different clinical responses to standard-of-care therapy.

Canine and murine models of osteosarcoma.

Osteosarcoma (OS) is the most common malignant bone tumor in children. Despite efforts to develop and implement new therapies, patient outcomes have not measurably improved since the 1980s. Metastasis continues to be the main source of patient mortality, with 30% of cases developing metastatic disease within 5 years of diagnosis. Research models are critical in the advancement of cancer research and include a variety of species. For example, xenograft and patient-derived xenograft (PDX) mouse models provide opportunities to study human tumor cells in vivo while transgenic models have offered significant insight into the molecular mechanisms underlying OS development. A growing recognition of naturally occurring cancers in companion species has led to new insights into how veterinary patients can contribute to studies of cancer biology and drug development. The study of canine cases, including the use of diagnostic tissue archives and clinical trials, offers a potential mechanism to further canine and human cancer research. Advancement in the field of OS research requires continued development and appropriate use of animal models. In this review, animal models of OS are described with a focus on the mouse and tumor-bearing pet dog as parallel and complementary models of human OS.

EML4-ALK induces cellular senescence in mortal normal human cells and promotes anchorage-independent growth in hTERT-transduced normal human cells.

BACKGROUND: Chromosomal inversions involving anaplastic lymphoma kinase (ALK) and echinoderm microtubule associated protein like 4 (EML4) generate a fusion protein EML4-ALK in non-small cell lung cancer (NSCLC). The understanding of EML4-ALK function can be improved by a functional study using normal human cells. METHODS: Here we for the first time conduct such study to examine the effects of EML4-ALK on cell proliferation, cellular senescence, DNA damage, gene expression profiles and transformed phenotypes. RESULTS: The lentiviral expression of EML4-ALK in mortal, normal human fibroblasts caused, through its constitutive ALK kinase activity, an early induction of cellular senescence with accumulated DNA damage, upregulation of p16INK4A and p21WAF1, and senescence-associated ß-galactosidase (SA-ß-gal) activity. In contrast, when EML4-ALK was expressed in normal human fibroblasts transduced with telomerase reverse transcriptase (hTERT), which is activated in the vast majority of NSCLC, the cells showed accelerated proliferation and acquired anchorage-independent growth ability in soft-agar medium, without accumulated DNA damage, chromosome aberration, nor p53 mutation. EML4-ALK induced the phosphorylation of STAT3 in both mortal and hTERT-transduced cells, but RNA sequencing analysis suggested that the different signaling pathways contributed to the different phenotypic outcomes in these cells. While EML4-ALK also induced anchorage-independent growth in hTERT-immortalized human bronchial epithelial cells in vitro, the expression of EML4-ALK alone did not cause detectable in vivo tumorigenicity in immunodeficient mice. CONCLUSIONS: Our data indicate that the expression of hTERT is critical for EML4-ALK to manifest its in vitro transforming activity in human cells. This study provides the isogenic pairs of human cells with and without EML4-ALK expression.

Targeting cellular senescence in cancer and aging: roles of p53 and its isoforms.

Cellular senescence and the associated secretory phenotype (SASP) promote disease in the aged population. Targeting senescent cells by means of removal, modulation of SASP or through cellular reprogramming represents a novel therapeutic avenue for treating cancer- and age-related diseases such as neurodegeneration, pulmonary fibrosis and renal disease. Cellular senescence is partly regulated by the TP53 gene, a critical tumor suppressor gene which encodes 12 or more p53 protein isoforms. This review marks a significant milestone of 40 years of Carcinogenesis publication history and p53 research and 15 years of p53 isoform research. The p53 isoforms are produced through initiation at alternative transcriptional and translational start sites and alternative mRNA splicing. These truncated p53 isoform proteins are endogenously expressed in normal human cells and maintain important functional roles, including modulation of full-length p53-mediated cellular senescence, apoptosis and DNA repair. In this review, we discuss the mechanisms and functions of cellular senescence and SASP in health and disease, the regulation of cellular senescence by p53 isoforms, and the therapeutic potential of targeting cellular senescence to treat cancer- and age-associated diseases.

Cellular Senescence: Mechanisms, Morphology, and Mouse Models.

Cellular senescence is a cell cycle arrest in damaged or aged cells. Although this represents a critical mechanism of tumor suppression, persistence of senescent cells during aging induces chronic inflammation and tissue dysfunction through the adoption of the senescence-associated secretory phenotype (SASP). This has been shown to promote the progression of age-associated diseases such as Alzheimer's disease, pulmonary fibrosis, and atherosclerosis. As the global population ages, the role of cellular senescence in disease is becoming a more critical area of research. In this review, mechanisms, biomarkers, and pathology of cellular senescence and SASP are described with a brief discussion of literature supporting a role for cellular senescence in veterinary diseases. Cell culture and mouse models used in senescence studies are also reviewed including the senescence-accelerated mouse-prone (SAMP), senescence pathway knockout mice (p53, p21 [CDKN1A], and p16 [CDKN2A]), and the more recently developed senolysis mice, which allow for direct visualization and elimination (or lysis) of senescent cells in live mice (p16-3MR and INK-ATTAC). These and other mouse models have demonstrated the importance of cellular senescence in embryogenesis and wound healing but have also identified a therapeutic benefit for targeting persistent senescent cells in age-associated diseases including neurodegeneration, diabetes, and cardiac fibrosis.

Diagnosis of Collagen Type III Glomerulopathy Using Picrosirius Red and PASH/Masson's Trichrome Stain.

Canine collagen type III glomerulopathy (Col3GP) is a rare juvenile nephropathy in which irregular type III collagen fibrils and fibronectin accumulate in glomerular capillary walls and the mesangium. Necropsy findings were reviewed from 5 puppies diagnosed with Col3GP at 6 to 18 weeks of age. Histologically, with hematoxylin and eosin stain, the glomerular capillary walls and mesangium were diffusely and globally expanded by homogeneous pale eosinophilic material. Ultrastructurally, the subendothelial zone and mesangium were expanded by fibronectin and cross-banded collagen type III fibrils, diagnostic of Col3GP. Two additional stains were employed to identify the material within glomeruli as fibrillar collagen using light microscopy. In all 5 cases, the material was red with picrosirius red and birefringent under polarized light, and was blue with periodic acid-Schiff/hematoxylin/trichrome (PASH/TRI), thereby identifying it as fibrillar collagen. Based on these unique staining characteristics with picrosirius red and PASH/TRI, Col3GP may be reliably diagnosed with light microscopy alone.

Surfactant Protein A and Napsin A in the Immunohistochemical Characterization of Canine Pulmonary Carcinomas: Comparison With Thyroid Transcription Factor-1.

Thyroid transcription factor-1 (TTF-1) is a specific and sensitive marker for canine pulmonary tumors but is also expressed in thyroid carcinomas, which commonly metastasize to lung. Napsin A and surfactant protein A (SP-A) are used in the histologic diagnosis of non-small-cell lung cancer in humans but have not been thoroughly evaluated in neoplasms of dogs. The objective of this study was to compare the efficacy of immunohistochemistry for SP-A, napsin A, and TTF-1 in the diagnosis of canine pulmonary carcinomas. TTF-1, napsin A, and SP-A antibodies were applied to 67 formalin-fixed, paraffin-embedded canine pulmonary tumors. Although each marker had good sensitivity, only 3% (2/67) of lung tumors were negative for SP-A compared with 7% (5/67) and 9% (6/67) for napsin A and TTF-1, respectively. Each antigen was detected in a greater percentage of cells of tumors with acinar or papillary patterns compared with those with squamous differentiation. SP-A immunoreactivity was absent in all 113 nonpulmonary tumors tested. Of 108 normal tissues, SP-A was detected only in lung and in 1 of 6 adrenal, 1 of 3 endometrial, and 1 of 4 hepatic sections. Based on these findings, SP-A and napsin A are useful markers of canine lung epithelial neoplasia. Of these, SP-A is the most sensitive and specific (a possible pitfall is the need to distinguish entrapped normal pulmonary epithelial cells or alveolar macrophages from neoplastic cells) and can be used in combination with TTF-1 or napsin A to improve detection and differentiation of pulmonary carcinomas from metastatic tumors in the canine lung.

Preparation, structural characterization, assessment of potential antiaromaticity and metalation of 21-oxyazuliporphyrins.

Oxidation of tetraarylazuliporphyrins with silver(I) acetate in refluxing chloroform-acetonitrile afforded good yields of 21-oxyazuliporphyrins. Although hydroxyazuliporphyrin tautomers can be considered for this system, spectroscopic results and density functional theory calculations indicate that the keto form is favored, and this was confirmed by single-crystal X-ray diffraction. Oxyazuliporphyrins formally possess a 24π electron delocalization pathway, but the proton NMR spectra are consistent with macrocycles that have diatropic ring currents. Nucleus independent chemical shift and anisotropy of induced current density calculations also confirmed the diatropic nature of these macrocycles, although these results indicated that the seven-membered ring is antiaromatic. However, while the NMR spectra showed the azulene protons at atypically high field values, the results are consistent with a nonaromatic cycloheptatrienyl unit. Protonation gave dicationic products that exhibited enhanced diatropic character. Oxyazuliporphyrins readily form metalated derivatives with Ni(II), Pd(II), and Pt(II), and these complexes exhibited significant diatropic character even though the macrocycle is highly distorted. X-ray diffraction characterization of palladium(II) and platinum(II) complexes demonstrated that these derivatives are structurally virtually identical to a previously reported copper(II) oxyazuliporphyrin.

Synthesis, structural characterization and reactivity of heteroazuliporphyrins.

A series of hetero-azuliporphyrins have been prepared by the "3 + 1" variant on the MacDonald condensation. Azulitripyrranes with tert-butyl and phenyl substituents reacted with thiophene or selenophene dialdehydes in the presence of TFA to give, following an oxidation step, thia- and selena-azuliporphyrins in 45-55% yield. Two of these compounds gave crystals suitable for X-ray crystallographic analysis and the data were consistent with the presence of a 17-atom delocalization pathway. The hetero-azuliporphyrins have significant diatropic character that is enhanced by the presence of an electron-donating tert-butyl substituent. The aromatic character is further increased in polar solvents such as DMSO, which are believed to stabilize dipolar resonance contributors with 18π electron delocalization pathways. Protonation also greatly increases the diatropic characteristics of these macrocycles. The porphyrinoids underwent an oxidative ring contraction with t-BuOOH-KOH to give moderate yields of benzoheterocarbaporphyrins. Reaction of azulitripyrranes with 2,5-furandicarbaldehyde afforded oxa-azuliporphyrins, a class of carbaporphyrinoids that had previously been inaccessible. These "missing links" in the study of heteroazuliporphyrins were isolated as the dihydrochloride salts. Protonated oxa-azuliporphyrins are stable aromatic compounds, but the free base forms underwent rapid decomposition in solution.

Measuring psychotherapeutic processes in the context of psychedelic experiences: Validation of the General Change Mechanisms Questionnaire (GCMQ).

BACKGROUND: Therapeutic and salutogenic effects of psychedelic drugs have been attributed to psychotherapeutic or psychotherapy-like processes that can unfold during the acute psychedelic experience and beyond. Currently, there are no psychometric instruments available to comprehensively assess psychotherapeutic processes (as conceptualized by empirical psychotherapy research) in the context of psychedelic experiences. AIMS: We report the initial validation of the General Change Mechanisms Questionnaire (GCMQ), a self-report instrument designed to measure five empirically established general change mechanisms (GCMs) of psychotherapy-(1) resource activation, (2) therapeutic relationship, (3) problem actuation, (4) clarification, and (5) mastery-in the context of psychedelic experiences. METHODS: An online survey in a sample of 1153 English-speaking and 714 German-speaking psychedelic users was conducted to evaluate simultaneously developed English- and German-language versions of the GCMQ. RESULTS: The theory-based factor structure was confirmed. The five GCMQ scales showed good internal consistency. Evidence for convergent validity with external measures was obtained. Significant associations with different settings and with therapeutic, hedonic, and escapist use motives confirmed the hypothesized context dependence of GCM-related psychedelic experiences. Indicating potential therapeutic effects, the association between cumulative stressful life events and well-being was significantly moderated by resource activation, clarification, and mastery. Factor mixture modeling revealed five distinct profiles of GCM-related psychedelic experiences. CONCLUSION: Initial testing indicates that the GCMQ is a valid and reliable instrument that can be used in future clinical and nonclinical psychedelic research. The five identified profiles of GCM-related experiences may be relevant to clinical uses of psychedelics and psychedelic harm reduction.

Comparative responses to demethylating therapy in animal models of osteosarcoma.

Background: The demethylating agent decitabine (DAC) effectively inhibits tumor growth and metastasis by targeting ESR1 methylation to restore estrogen receptor alpha (ERα) signaling and promoting cellular differentiation in models of human osteosarcoma (OSA). Whether this pathway can be targeted in canine OSA patients is unknown. Methods: Canine OSA tumor samples were tested for ERα expression and ESR1 promoter methylation. Human (MG63.3) and canine (MC-KOS) OSA cell lines and murine xenografts were treated with DAC in vitro and in vivo, respectively. Samples were assessed using mRNA sequencing and tissue immunohistochemistry. Results: ESR1 is methylated in a subset of canine OSA patient samples and the MC-KOS cell line. DAC treatment led to enhanced differentiation as demonstrated by increased ALPL expression, and suppressed tumor growth in vitro and in vivo. Metastatic progression was inhibited, particularly in the MG63.3 model, which expresses higher levels of DNA methyltransferases DNMT1 and 3B. DAC treatment induced significant alterations in immune response and cell cycle pathways. Conclusion: DAC treatment activates ERα signaling, promotes bone differentiation, and inhibits tumor growth and metastasis in human and canine OSA. Additional DAC-altered pathways and species- or individual-specific differences in DNMT expression may also play a role in DAC treatment of OSA.

Clinical, pathologic and molecular findings in 2 Rottweiler littermates with appendicular osteosarcoma.

Appendicular osteosarcoma was diagnosed and treated in a pair of littermate Rottweiler dogs, resulting in distinctly different clinical outcomes despite similar therapy within the context of a prospective, randomized clinical trial (NCI-COTC021/022). Histopathology, immunohistochemistry, mRNA sequencing, and targeted DNA hotspot sequencing techniques were applied to both dogs' tumors to define factors that could underpin their differential response to treatment. We describe the comparison of their clinical, histologic and molecular features, as well as those from a companion cohort of Rottweiler dogs, providing new insight into potential prognostic biomarkers for canine osteosarcoma.

Large Scale Comparative Deconvolution Analysis of the Canine and Human Osteosarcoma Tumor Microenvironment Uncovers Conserved Clinically Relevant Subtypes.

Osteosarcoma is a relatively rare but aggressive cancer of the bones with a shortage of effective biomarkers. Although less common in humans, Osteosarcomas are fairly common in adult pet dogs and have been shown to share many similarities with their human analogs. In this work, we analyze bulk transcriptomic data of 213 primary and 100 metastatic Osteosarcoma samples from 210 pet dogs enrolled in nation-wide clinical trials to uncover three Tumor Microenvironment (TME)-based subtypes: Immune Enriched (IE), Immune Enriched Dense Extra-Cellular Matrix-like (IE-ECM) and Immune Desert (ID) with distinct cell type compositions, oncogenic pathway activity and chromosomal instability. Furthermore, leveraging bulk transcriptomic data of canine primary tumors and their matched metastases from different sites, we characterize how the Osteosarcoma TME evolves from primary to metastatic disease in a standard of care clinical setting and assess its overall impact on clinical outcomes of canines. Most importantly, we find that TME-based subtypes of canine Osteosarcomas are conserved in humans and predictive of progression free survival outcomes of human patients, independently of known prognostic biomarkers such as presence of metastatic disease at diagnosis and percent necrosis following chemotherapy. In summary, these results demonstrate the power of using canines to model the human Osteosarcoma TME and discover novel biomarkers for clinical translation.

Transcriptional profiling of canine osteosarcoma identifies prognostic gene expression signatures with translational value for humans.

Canine osteosarcoma is increasingly recognized as an informative model for human osteosarcoma. Here we show in one of the largest clinically annotated canine osteosarcoma transcriptional datasets that two previously reported, as well as de novo gene signatures devised through single sample Gene Set Enrichment Analysis (ssGSEA), have prognostic utility in both human and canine patients. Shared molecular pathway alterations are seen in immune cell signaling and activation including TH1 and TH2 signaling, interferon signaling, and inflammatory responses. Virtual cell sorting to estimate immune cell populations within canine and human tumors showed similar trends, predominantly for macrophages and CD8+ T cells. Immunohistochemical staining verified the increased presence of immune cells in tumors exhibiting immune gene enrichment. Collectively these findings further validate naturally occurring osteosarcoma of the pet dog as a translationally relevant patient model for humans and improve our understanding of the immunologic and genomic landscape of the disease in both species.

Exploration of Imaging Biomarkers for Metabolically-Targeted Osteosarcoma Therapy in a Murine Xenograft Model.

Background: Osteosarcoma (OS) is an aggressive pediatric cancer with unmet therapeutic needs. Glutaminase 1 (GLS1) inhibition, alone and in combination with metformin, disrupts the bioenergetic demands of tumor progression and metastasis, showing promise for clinical translation. Materials and Methods: Three positron emission tomography (PET) clinical imaging agents, [18F]fluoro-2-deoxy-2-D-glucose ([18F]FDG), 3'-[18F]fluoro-3'-deoxythymidine ([18F]FLT), and (2S, 4R)-4-[18F]fluoroglutamine ([18F]GLN), were evaluated in the MG63.3 human OS xenograft mouse model, as companion imaging biomarkers after treatment for 7 d with a selective GLS1 inhibitor (CB-839, telaglenastat) and metformin, alone and in combination. Imaging and biodistribution data were collected from tumors and reference tissues before and after treatment. Results: Drug treatment altered tumor uptake of all three PET agents. Relative [18F]FDG uptake decreased significantly after telaglenastat treatment, but not within control and metformin-only groups. [18F]FLT tumor uptake appears to be negatively affected by tumor size. Evidence of a flare effect was seen with [18F]FLT imaging after treatment. Telaglenastat had a broad influence on [18F]GLN uptake in tumor and normal tissues. Conclusions: Image-based tumor volume quantification is recommended for this paratibial tumor model. The performance of [18F]FLT and [18F]GLN was affected by tumor size. [18F]FDG may be useful in detecting telaglenastat's impact on glycolysis. Exploration of kinetic tracer uptake protocols is needed to define clinically relevant patterns of [18F]GLN uptake in patients receiving telaglenastat.

Δ133p53α Protects Human Astrocytes from Amyloid-beta Induced Senescence and Neurotoxicity.

Cellular senescence is an important contributor to aging and age-related diseases such as Alzheimer's disease (AD). Senescent cells are characterized by a durable cell proliferation arrest and the acquisition of a proinflammatory senescence-associated secretory phenotype (SASP), which participates in the progression of neurodegenerative disorders. Clearance of senescent glial cells in an AD mouse model prevented cognitive decline suggesting pharmacological agents targeting cellular senescence might provide novel therapeutic approaches for AD. Δ133p53α, a natural protein isoform of p53, was previously shown to be a negative regulator of cellular senescence in primary human astrocytes, with clinical implications from its diminished expression in brain tissues from AD patients. Here we show that treatment of proliferating human astrocytes in culture with amyloid-beta oligomers (Aß), an endogenous pathogenic agent of AD, results in reduced expression of Δ133p53α, as well as induces the cells to become senescent and express proinflammatory SASP cytokines such as IL-6, IL-1ß and TNFα. Our data suggest that Aß-induced astrocyte cellular senescence is associated with accelerated DNA damage, and upregulation of full-length p53 and its senescence-inducing target gene p21WAF1. We also show that exogenously enhanced expression of Δ133p53α rescues human astrocytes from Aß-induced cellular senescence and SASP through both protection from DNA damage and dominant-negative inhibition of full-length p53, leading to inhibition of Aß-induced, astrocyte-mediated neurotoxicity. The results presented here demonstrate that Δ133p53α manipulation could modulate cellular senescence in the context of AD, possibly opening new therapeutic avenues.

Creatine riboside is a cancer cell-derived metabolite associated with arginine auxotrophy.

The metabolic dependencies of cancer cells have substantial potential to be exploited to improve the diagnosis and treatment of cancer. Creatine riboside (CR) is identified as a urinary metabolite associated with risk and prognosis in lung and liver cancer. However, the source of high CR levels in patients with cancer as well as their implications for the treatment of these aggressive cancers remain unclear. By integrating multiomics data on lung and liver cancer, we have shown that CR is a cancer cell-derived metabolite. Global metabolomics and gene expression analysis of human tumors and matched liquid biopsies, together with functional studies, revealed that dysregulation of the mitochondrial urea cycle and a nucleotide imbalance were associated with high CR levels and indicators of a poor prognosis. This metabolic phenotype was associated with reduced immune infiltration and supported rapid cancer cell proliferation that drove aggressive tumor growth. CRhi cancer cells were auxotrophic for arginine, revealing a metabolic vulnerability that may be exploited therapeutically. This highlights the potential of CR not only as a poor-prognosis biomarker but also as a companion biomarker to inform the administration of arginine-targeted therapies in precision medicine strategies to improve survival for patients with cancer.

Astrocyte senescence and SASP in neurodegeneration: tau joins the loop.

Tau accumulation is a core component of Alzheimer's disease and other neurodegenerative tauopathies. While tau's impact on neurons is a major area of research, the effect of extracellular tau on astrocytes is largely unknown. This article summarizes our recent studies showing that astrocyte senescence plays a critical role in neurodegenerative diseases and integrates extracellular tau into the regulatory loop of senescent astrocyte-mediated neurotoxicity. Human astrocytes in vitro undergoing senescence were shown to acquire the inflammatory senescence-associated secretory phenotype (SASP) and toxicity to neurons, which may recapitulate aging- and disease-associated neurodegeneration. Here, we show that human astrocytes exposed to extracellular tau in vitro also undergo cellular senescence and acquire a neurotoxic SASP (e.g. IL-6 secretion), with oxidative stress response (indicated by upregulated NRF2 target genes) and a possible activation of inflammasome (indicated by upregulated ASC and IL-1ß). These findings suggest that senescent astrocytes induced by various conditions and insults, including tau exposure, may represent a therapeutic target to inhibit or delay the progression of neurodegenerative diseases. We also discuss the pathological activity of extracellular tau in microglia and astrocytes, the disease relevance and diversity of tau forms, therapeutics targeting senescence in neurodegeneration, and the roles of p53 and its isoforms in astrocyte-mediated neurotoxicity and neuroprotection.

Consideration of Both Discriminated and Generalized Responding When Teaching Children with Autism Abduction Prevention Skills.

We taught three children with autism how to respond to abduction lures presented by strangers. We then tested undesirable generalization of the safety response to matched instructions to leave by a familiar adult. Following training, all three participants engaged in the safety response across both strangers and familiar adults. Thus, we evaluated a set of procedures for establishing discriminated responding. Appropriate responding to instructions to leave by strangers versus familiar adults was achieved only after discrimination training. Discriminated responding occurred across a novel setting and maintained across 3 months; however, performance during stimulus generalization probes within community settings was variable. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s40617-020-00541-9.

The Δ133p53 Isoforms, Tuners of the p53 Pathway.

The TP53 gene is a critical tumor suppressor and key determinant of cell fate which regulates numerous cellular functions including DNA repair, cell cycle arrest, cellular senescence, apoptosis, autophagy and metabolism. In the last 15 years, the p53 pathway has grown in complexity through the discovery that TP53 differentially expresses twelve p53 protein isoforms in human cells with both overlapping and unique biologic activities. Here, we summarize the current knowledge on the Δ133p53 isoforms (Δ133p53α, Δ133p53ß and Δ133p53γ), which are evolutionary derived and found only in human and higher order primates. All three isoforms lack both of the transactivation domains and the beginning of the DNA-binding domain. Despite the absence of these canonical domains, the Δ133p53 isoforms maintain critical functions in cancer, physiological and premature aging, neurodegenerative diseases, immunity and inflammation, and tissue repair. The ability of the Δ133p53 isoforms to modulate the p53 pathway functions underscores the need to include these p53 isoforms in our understanding of how the p53 pathway contributes to multiple physiological and pathological mechanisms. Critically, further characterization of p53 isoforms may identify novel regulatory modes of p53 pathway functions that contribute to disease progression and facilitate the development of new therapeutic strategies.