Signaling to Cardiac Hypertrophy: Insights from Human and Mouse RASopathies

Cardiac hypertrophy is the heart’s response to a variety of extrinsic and intrinsic stimuli, some of which might finally lead up to a maladaptive state. An integral part of the pathogenesis of the hypertrophic cardiomyopathy disease (HCM) is the activation of the rat sarcoma (RAS)/RAF/MEK (mitogen-activated protein kinase kinase)/MAPK (mitogen-activated protein kinase) cascade. Therefore, the molecular signaling involving RAS has been the subject of intense research efforts, particularly after the identification of the RASopathies. These constitute a class of developmental disorders caused by germline mutations affecting proteins contributing to the RAS pathway. Among other phenotypic features, a subset of these syndromes is characterized by HCM, prompting researchers and clinicians to delve into the chief signaling constituents of cardiac hypertrophy. In this review, we summarize current advances in the knowledge of the molecular signaling events involved in the pathogenesis of cardiac hypertrophy through work completed on patients and on genetically manipulated animals with HCM and RASopathies. Important insights are drawn from the recognition of parallels between cardiac hypertrophy and cancer. Future research promises to further elucidate the complex molecular interactions responsible for cardiac hypertrophy, possibly pointing the way for the identification of new specific targets for the treatment of HCM.     

Signaling to Cardiac Hypertrophy: Insights from Human and Mouse RASopathies

Cardiac hypertrophy is the heart's response to a variety of extrinsic and intrinsic stimuli, some of which might finally lead up to a maladaptive state. An integral part of the pathogenesis of the hypertrophic cardiomyopathy disease (HCM) is the activation of the rat sarcoma (RAS)/RAF/MEK (mitogen-activated protein kinase kinase)/MAPK (mitogen-activated protein kinase) cascade. Therefore, the molecular signaling involving RAS has been the subject of intense research efforts, particularly after the identification of the RASopathies. These constitute a class of developmental disorders caused by germline mutations affecting proteins contributing to the RAS pathway. Among other phenotypic features, a subset of these syndromes is characterized by HCM, prompting researchers and clinicians to delve into the chief signaling constituents of cardiac hypertrophy. In this review, we summarize current advances in the knowledge of the molecular signaling events involved in the pathogenesis of cardiac hypertrophy through work completed on patients and on genetically manipulated animals with HCM and RASopathies. Important insights are drawn from the recognition of parallels between cardiac hypertrophy and cancer. Future research promises to further elucidate the complex molecular interactions responsible for cardiac hypertrophy, possibly pointing the way for the identification of new specific targets for the treatment of HCM.     

Therapeutic targeting of RAS: New hope for drugging the “undruggable”

RAS is the most frequently mutated oncogene in cancer and a critical driver of oncogenesis. Therapeutic targeting of RAS has been a goal of cancer research for more than 30 years due to its essential role in tumor formation and maintenance. Yet the quest to inhibit this challenging foe has been elusive. Although once considered “undruggable”, the struggle to directly inhibit RAS has seen recent success with the development of pharmacological agents that specifically target the KRAS(G12C) mutant protein, which include the first direct RAS inhibitor to gain entry to clinical trials. However, the limited applicability of these inhibitors to G12C-mutant tumors demands further efforts to identify more broadly efficacious RAS inhibitors. Understanding allosteric influences on RAS may open new avenues to inhibit RAS. Here, we provide a brief overview of RAS biology and biochemistry, discuss the allosteric regulation of RAS, and summarize the various approaches to develop RAS inhibitors.     

HSPA5 negatively regulates lysosomal activity through ubiquitination of MUL1 in head and neck cancer

HSPA5/GRP78/BiP plays an important role in cell survival or tumor progression. For these reasons, HSPA5 is an emerging therapeutic target in cancer development. Here we report that HSPA5 contributes to head and neck cancer (HNC) survival via maintenance of lysosomal activity; however, a nonthermal plasma (NTP, considered as a next-generation cancer therapy)-treated solution (NTS) inhibits HNC progression through HSPA5-dependent alteration of lysosomal activity. HSPA5 prevents NTS-induced lysosome inhibition through lysosomal-related proteins or regulation of gene expression. However, NTS-induced MUL1/MULAN/GIDE/MAPL (mitochondrial ubiquitin ligase activator of NFKB 1) leads to downregulation of HSPA5 via K48-linked ubiquitination at the lysine 446 (K446) residue. MUL1 knockdown hinders NTS-induced lysosome inhibition or cytotoxicity through the reduction of HSPA5 ubiquitination in HNC cells. While MUL1 was suppressed, HSPA5 was overexpressed in tissues of HNC patients. NTS strongly inhibited HNC progression via alterations of expression of MUL1 and HSPA5, in vivo in a xenograft model. However, NTS did not induce inhibition of tumor progression or HSPA5 reduction in MUL1 knockout (KO) HNC cells which were generated by CRISPR/Cas9 system. The data provide compelling evidence to support the idea that the regulation of the MUL1-HSPA5 axis can be a novel strategy for the treatment of HNC.     

Comparative Oncogenomics Implicates the Neurofibromin 1 Gene (NF1) as a Breast Cancer Driver

Identifying genomic alterations driving breast cancer is complicated by tumor diversity and genetic heterogeneity. Relevant mouse models are powerful for untangling this problem because such heterogeneity can be controlled. Inbred Chaos3 mice exhibit high levels of genomic instability leading to mammary tumors that have tumor gene expression profiles closely resembling mature human mammary luminal cell signatures. We genomically characterized mammary adenocarcinomas from these mice to identify cancer-causing genomic events that overlap common alterations in human breast cancer. Chaos3 tumors underwent recurrent copy number alterations (CNAs), particularly deletion of the RAS inhibitor Neurofibromin 1 (Nf1) in nearly all cases. These overlap with human CNAs including NF1, which is deleted or mutated in 27.7% of all breast carcinomas. Chaos3 mammary tumor cells exhibit RAS hyperactivation and increased sensitivity to RAS pathway inhibitors. These results indicate that spontaneous NF1 loss can drive breast cancer. This should be informative for treatment of the significant fraction of patients whose tumors bear NF1 mutations.     

Advances in micro-CT imaging of small animals

PurposeMicron-scale computed tomography (micro-CT) imaging is a ubiquitous, cost-effective, and non-invasive three-dimensional imaging modality. We review recent developments and applications of micro-CT for preclinical research.MethodsBased on a comprehensive review of recent micro-CT literature, we summarize features of state-of-the-art hardware and ongoing challenges and promising research directions in the field.ResultsRepresentative features of commercially available micro-CT scanners and some new applications for both in vivo and ex vivo imaging are described. New advancements include spectral scanning using dual-energy micro-CT based on energy-integrating detectors or a new generation of photon-counting x-ray detectors (PCDs). Beyond two-material discrimination, PCDs enable quantitative differentiation of intrinsic tissues from one or more extrinsic contrast agents. When these extrinsic contrast agents are incorporated into a nanoparticle platform (e.g. liposomes), novel micro-CT imaging applications are possible such as combined therapy and diagnostic imaging in the field of cancer theranostics. Another major area of research in micro-CT is in x-ray phase contrast (XPC) imaging. XPC imaging opens CT to many new imaging applications because phase changes are more sensitive to density variations in soft tissues than standard absorption imaging. We further review the impact of deep learning on micro-CT. We feature several recent works which have successfully applied deep learning to micro-CT data, and we outline several challenges specific to micro-CT.ConclusionsAll of these advancements establish micro-CT imaging at the forefront of preclinical research, able to provide anatomical, functional, and even molecular information while serving as a testbench for translational research.     

Investigation of urinary volatomic alterations in head and neck cancer: a non-invasive approach towards diagnosis and prognosis

Introduction

Head and neck cancer (HNC), like many other forms of cancer, is usually detected in advanced stages, causing poor survival outcomes. Lack of specific and sensitive screening markers for early detection of HNC has worsened the scenario for the patients as well as the clinicians. Therefore, identification of efficient, noninvasive and affordable screening marker/methodology with high specificity and sensitivity is imminent need of situation.

Objectives

This study aims to identify and characterize urinary volatomic alterations specific to HNC.

Methods

Volatomic analysis of urine samples collected from HNC patients (n?=?29) and healthy controls (n?=?31) was performed using headspace solid phase microextraction coupled to gas chromatography mass spectrometry (GC–MS). Both univariate and multivariate statistical approaches were used to investigate HNC specific volatomic alterations.

Results

Statistical analysis revealed a total of 28 metabolites with highest contribution towards discrimination of HNC patients from healthy controls (VIP >1, p?<?0.05, Log₂ FC ≥0.58/≤?0.57). The discrimination efficiency and accuracy of urinary VOCs was ascertained by ROC curve analysis that allowed the identification of four metabolites viz. 2,6-dimethyl-7-octen-2-ol, 1-butanol, p-xylene and 4-methyl-2-heptanone with highest sensitivity and specificity to discriminate HNC patients from healthy controls. Further, the metabolic pathway analysis identified several dysregulated pathways in HNC patients and their detailed investigations could unravel novel mechanistic insights into the disease pathophysiology.

Conclusion

Overall, this study provides valuable fingerprint of the volatile profile of HNC patients, which in turn, might help in improving the current understanding of this form of cancer and lead to the development of non-invasive approaches for HNC diagnosis.

     

Caspase-dependent apoptosis induced by two synthetic halogenated flavanones, 3′,7-dichloroflavanone and 3′,6-dichloroflavanone,on human breast and prostate cancer cells

The destruction of cancer cells with chemotherapeutic agents is normally achieved through apoptosis. We previously introduced two synthetic halogenated flavanone derivatives, 3^′,7-dichloroflavanone (3′-7 DCF) and 3^′,6-dichloroflavanone (3′-6 DCF), as potential apoptosis-inducing agents. In the current study, we investigated the ability of these compounds in triggering intrinsic or/and extrinsic pathway of apoptosis in breast and prostate cancer cells. Also, the synergistic effect of 3′-7 DCF with TLR3 (Toll-like receptor 3) agonist in apoptosis induction was evaluated on PC3 and LNCaP human prostate cancer cells. The involved pathway of apoptosis in the treated cells was delineated by caspase-3 activity assay, PARP-1 (poly(ADP-ribose)polymerase-1) cleavage, and procaspase-9 cleavage as markers of the intrinsic pathway and procaspase-8 cleavage as the marker of the extrinsic pathway. With the exception of the normal cells, treatment of all cell lines with both 3′-7 DCF and 3′-6 DCF triggered the cleavage of procaspase-8 and procaspase-9. These results indicate that the intrinsic and the extrinsic pathways of apoptosis are the mechanisms of the toxicity of flavanones in these cancer cell lines. However, the cytoxicity of the compound 3′-7 DCF was not synergistic with TLR3 agonist. Interestingly, the activation of caspases-9 preceeded that of caspase-8 suggesting that the intrinsic pathway is the primary reason for apoptosis induction by the flavanones.     

Significance of Lymph Node Metastasis in Cancer Dissemination of Head and Neck Cancer

Lymph node metastasis (LNM) in many solid cancers is a well-known prognostic factor; however, it has been debated whether regional LNM simply reflects tumor aggressiveness or is a source for further tumor dissemination. Similarly, the metastatic process in head and neck cancer (HNC) has not been fully evaluated. Thus, we aimed to investigate the relative significance of LNM in metastatic cascade of HNC using functional imaging of HNC patients and molecular imaging in in vivo models. First, we analyzed ¹⁸Fluorodeoxyglucose positron emission tomography (PET) parameters of 117 patients with oral cancer. The primary tumor and nodal PET parameters were measured separately, and survival analyses were conducted on the basis of clinical and PET variables to identify significant prognostic factors. In multivariate analyses, we found that only the metastatic node PET values were significant. Next, we compared the relative frequency of lung metastasis in primary ear tumors versus lymph node (LN) tumors, and we tested the rate of lung metastasis in another animal model, in which each animal had both primary and LN tumors that were expressing different colors. As a result, LN tumors showed higher frequencies of lung metastasis compared to orthotopic primary tumors. In color-matched comparisons, the relative contribution to lung metastasis was higher in LN tumors than in primary tumors, although both primary and LN tumors caused lung metastases. In summary, tumors growing in the LN microenvironment spread to systemic sites more commonly than primary tumors in HNC, suggesting that the adequate management of LNM can reduce further systemic metastasis.     

RAS and RHO family GTPase mutations in cancer: twin sons of different mothers?

Abstract

The RAS and RHO family comprise two major branches of the RAS superfamily of small GTPases. These proteins function as regulated molecular switches and control cytoplasmic signaling networks that regulate a diversity of cellular processes, including cell proliferation and cell migration. In the early 1980s, mutationally activated RAS genes encoding KRAS, HRAS and NRAS were discovered in human cancer and now comprise the most frequently mutated oncogene family in cancer. Only recently, exome sequencing studies identified cancer-associated alterations in two RHO family GTPases, RAC1 and RHOA. RAS and RHO proteins share significant identity in their amino acid sequences, protein structure and biochemistry. Cancer-associated RAS mutant proteins harbor missense mutations that are found primarily at one of three mutational hotspots (G12, G13 and Q61) and have been identified as gain-of-function oncogenic alterations. Although these residues are conserved in RHO family proteins, the gain-of-function mutations found in RAC1 are found primarily at a distinct hotspot. Unexpectedly, the cancer-associated mutations found with RHOA are located at different hotspots than those found with RAS. Furthermore, since the RHOA mutations suggested a loss-of-function phenotype, it has been unclear whether RHOA functions as an oncogene or tumor suppressor in cancer development. Finally, whereas RAS mutations are found in a broad spectrum of cancer types, RHOA and RAC1 mutations occur in a highly restricted range of cancer types. In this review, we focus on RHOA missense mutations found in cancer and their role in driving tumorigenesis, with comparisons to cancer-associated mutations in RAC1 and RAS GTPases.     

Stress-induced apoptosis: Toward a symmetry with receptor-mediated cell death

Apoptosis is a form of programmed cell death executed by caspases activated along signalling pathways initiated by ligation of cell-surface death receptors ( extrinsic pathway ) or by perturbation of the mithocondrial membrane promoted by physical or chemical stress agents ( intrinsic pathway ). In metazoans, this evolutionary conserved, genetically controlled process has a role in a variety of physiological settings, as development, homeostasis of tissues and maintenance of the organism integrity. When deranged by impaired regulation or inappropriate activation apoptosis contributes to the pathogenesis of diseases as autoimmunity, cancer, restenosis, ischaemia, heart failure and neurodegenerative disorders. In this review we will present a survey of the stress-induced intrinsic, mithochondrial, pathway and, based on recent experimental data, we will propose a view compatible with an emergent conceptual symmetry between the two apoptogenic extrinsic and intrinsic pathways. Elements of symmetry present in both the apoptogenic signalling pathways include: early activation of initiator caspases (feed-forwarded by a direct or post-mitocondrial effector caspase-mediated amplification loop in some cell types) and mitochondrial membrane permeabilization with required release of antagonists of active caspase inhibitors (IAPs) in high-level IAPs-expressing cells and apoptosome-mediated amplification of the caspase cascade more or less needed in different cell types.     

Propolis cinnamic acid derivatives induce apoptosis through both extrinsic and intrinsic apoptosis signaling pathways and modulate of miRNA expression

Propolis cinnamic acid derivatives have a number of biological activities including anti-oxidant and anti-cancer ones. In this study, we aimed to elucidate the mechanism of the anti-cancer activity of 3 representative propolis cinnamic acid derivatives, i.e., Artepilin C, Baccharin and Drupanin in human colon cancer cell lines. Our study demonstrated that these compounds had a potent apoptosis-inductive effect even on drug-resistant colon cancer cells. Combination treatment of human colon cancer DLD-1 cells with 2 of these compounds, each at its IC₂₀ concentration, induced apoptosis by stimulating both intrinsic and extrinsic apoptosis signaling pathways. Especially, Baccharin plus Drupanin exhibited a synergistic growth-inhibitory effect by strengthening both intrinsic and extrinsic apoptotic signaling transduction through TRAIL/DR4/5 and/or FasL/Fas death-signaling loops and by increasing the expression level of miR-143, resulting in decreased expression levels of the target gene MAPK/Erk5 and its downstream target c-Myc. These data suggest that the supplemental intake of these compounds found in propolis has enormous significance with respect to cancer prevention.     

The Mitochondrial Protein NLRX1 Controls the Balance between Extrinsic and Intrinsic Apoptosis

NLRX1 is a mitochondrial Nod-like receptor (NLR) protein whose function remains enigmatic. Here, we observed that NLRX1 expression was glucose-regulated and blunted by SV40 transformation. In transformed but not primary murine embryonic fibroblasts, NLRX1 expression mediated resistance to an extrinsic apoptotic signal, whereas conferring susceptibility to intrinsic apoptotic signals, such as glycolysis inhibition, increased cytosolic calcium and endoplasmic reticulum stress. In a murine model of colorectal cancer induced by azoxymethane, NLRX1−/− mice developed fewer tumors than wild type mice. In contrast, in a colitis-associated cancer model combining azoxymethane and dextran sulfate sodium, NLRX1−/− mice developed a more severe pathology likely due to the increased sensitivity to dextran sulfate sodium colitis. Together, these results identify NLRX1 as a critical mitochondrial protein implicated in the regulation of apoptosis in cancer cells. The unique capacity of NLRX1 to regulate the cellular sensitivity toward intrinsic versus extrinsic apoptotic signals suggests a critical role for this protein in numerous physiological processes and pathological conditions.     

Post-translational modification of RAS proteins

Mutations of RAS genes drive cancer more frequently than any other oncogene. RAS proteins integrate signals from a wide array of receptors and initiate downstream signaling through pathways that control cellular growth. RAS proteins are fundamentally binary molecular switches in which the off/on state is determined by the binding of GDP or GTP, respectively. As such, the intrinsic and regulated nucleotide-binding and hydrolytic properties of the RAS GTPase were historically believed to account for the entirety of the regulation of RAS signaling. However, it is increasingly clear that RAS proteins are also regulated by a vast array of post-translational modifications (PTMs). The current challenge is to understand what are the functional consequences of these modifications and which are physiologically relevant. Because PTMs are catalyzed by enzymes that may offer targets for drug discovery, the study of RAS PTMs has been a high priority for RAS biologists.     

Unraveling the regulatory role of endoplasmic-reticulum-associated degradation in tumor immunity

Abstract

During malignant transformation and cancer progression, tumor cells face both intrinsic and extrinsic stress, endoplasmic reticulum (ER) stress in particular. To survive and proliferate, tumor cells use multiple stress response pathways to mitigate ER stress, promoting disease aggression and treatment resistance. Among the stress response pathways is ER-associated degradation (ERAD), which consists of multiple components and steps working together to ensure protein quality and quantity. In addition to its established role in stress responses and tumor cell survival, ERAD has recently been shown to regulate tumor immunity. Here we summarize current knowledge on how ERAD promotes protein degradation, regulates immune cell development and function, participates in antigen presentation, exerts paradoxical roles on tumorigenesis and immunity, and thus impacts current cancer therapy. Collectively, ERAD is a critical protein homeostasis pathway intertwined with cancer development and tumor immunity. Of particular importance is the need to further unveil ERAD’s enigmatic roles in tumor immunity to develop effective targeted and combination therapy for successful treatment of cancer.     

Resistance mechanisms to immune checkpoints blockade by monoclonal antibody drugs in cancer immunotherapy: Focus on myeloma

Multiple myeloma (MM) is a clonal B‐cell malignancy characterized by the accumulation of neoplastic proliferation of a plasma cell in the bone marrow that produces a monoclonal immunoglobulin. The immune checkpoint inhibitors against programmed death‐1/programmed death‐1 ligand and cytotoxic T‐lymphocyte antigen 4 axis have demonstrated appropriate anticancer activity in several solid tumors and liquid cancers, and are rapidly transforming the practice of medical oncology. However, in a high percentage of patients, the efficacy of immune checkpoints blockade remains limited due to innate or primary resistance. Moreover, the malignancies progress in many patients due to acquired or secondary resistance, even after the clinical response to immune checkpoints' blockade. The evidence shows that multiple tumor‐intrinsic and tumor‐extrinsic factors and alterations in signaling pathways are involved in primary and secondary resistance to immune checkpoints blockade. Improved identification of intrinsic and extrinsic factors and mechanisms of resistance or response to immune checkpoints blockade may not only provide novel prognostic or predictive biomarkers but also guide the optimal combination/sequencing of immune checkpoint blockade therapy in the clinic. Here, we review the underlying biology and role of immune checkpoints blockade in patients with MM. Furthermore, we review the host and tumor‐related factor effects on immune checkpoints blockade in MM immunotherapy.     

Role of DNA methylation in head and neck cancer

Head and neck cancer (HNC) is a heterogenous and complex entity including diverse anatomical sites and a variety of tumor types displaying unique characteristics and different etilogies. Both environmental and genetic factors play a role in the development of the disease, but the underlying mechanism is still far from clear. Previous studies suggest that alterations in the genes acting in cellular signal pathways may contribute to head and neck carcinogenesis. In cancer, DNA methylation patterns display specific aberrations even in the early and precancerous stages and may confer susceptibility to further genetic or epigenetic changes. Silencing of the genes by hypermethylation or induction of oncogenes by promoter hypomethylation are frequent mechanisms in different types of cancer and achieve increasing diagnostic and therapeutic importance since the changes are reversible. Therefore, methylation analysis may provide promising clinical applications, including the development of new biomarkers and prediction of the therapeutic response or prognosis. In this review, we aimed to analyze the available information indicating a role for the epigenetic changes in HNC.     

Induction of intrinsic and extrinsic apoptosis through oxidative stress in drug-resistant cancer by a newly synthesized Schiff base copper chelate

Multidrug resistance (MDR) in cancer represents a variety of strategies employed by tumor cells to evade the beneficial cytotoxic effects of structurally different anticancer drugs and thus confers impediments to the successful treatment of cancers. Efflux of drugs by MDR protein-1, functional P-glycoprotein and elevated level of reduced glutathione confer resistance to cell death or apoptosis and thus provide a possible therapeutic target for overcoming MDR in cancer. Previously, we reported that a Schiff base ligand, potassium-N-(2-hydroxy 3-methoxy-benzaldehyde)-alaninate (PHMBA) overcomes MDR in both in vivo and in vitro by targeting intrinsic apoptotic/necrotic pathway through induction of reactive oxygen species (ROS). The present study describes the synthesis and spectroscopic characterization of a copper chelate of Schiff base, viz., copper (II)-N-(2-hydroxy-3-methoxy-benzaldehyde)-alaninate (CuPHMBA) and the underlying mechanism of cell death induced by CuPHMBA in vitro. CuPHMBA kills both the drug-resistant and sensitive cell types irrespective of their drug resistance phenotype. The cell death induced by CuPHMBA follows apoptotic pathway and moreover, the cell death is associated with intrinsic mitochondrial and extrinsic receptor-mediated pathways. Oxidative stress plays a pivotal role in the process as proved by the fact that antioxidant enzyme; polyethylene glycol conjugated-catalase completely blocked CuPHMBA-induced ROS generation and abrogated cell death. To summarize, the present work provides a compelling rationale for the future clinical use of CuPHMBA, a redox active copper chelate in the treatment of cancer patients, irrespective of their drug-resistance status.     

Regulation of Epidermal Growth Factor Receptor Signaling and Erlotinib Sensitivity in Head and Neck Cancer Cells by miR-7

Elevated expression and activity of the epidermal growth factor receptor (EGFR)/protein kinase B (Akt) signaling pathway is associated with development, progression and treatment resistance of head and neck cancer (HNC). Several studies have demonstrated that microRNA-7 (miR-7) regulates EGFR expression and Akt activity in a range of cancer cell types via its specific interaction with the EGFR mRNA 3′-untranslated region (3′-UTR). In the present study, we found that miR-7 regulated EGFR expression and Akt activity in HNC cell lines, and that this was associated with reduced growth in vitro and in vivo of cells (HN5) that were sensitive to the EGFR tyrosine kinase inhibitor (TKI) erlotinib (Tarceva). miR-7 acted synergistically with erlotinib to inhibit growth of erlotinib-resistant FaDu cells, an effect associated with increased inhibition of Akt activity. Microarray analysis of HN5 and FaDu cell lines transfected with miR-7 identified a common set of downregulated miR-7 target genes, providing insight into the tumor suppressor function of miR-7. Furthermore, we identified several target miR-7 mRNAs with a putative role in the sensitization of FaDu cells to erlotinib. Together, these data support the coordinate regulation of Akt signaling by miR-7 in HNC cells and suggest the therapeutic potential of miR-7 alone or in combination with EGFR TKIs in this disease.     

Association of CYP1A1 and CYP2D6 gene polymorphisms with head and neck cancer in Tunisian patients

The purpose of this study was to investigate the relationship between head and neck cancer (HNC) and environmental agents and polymorphisms in CYP1A1, CYP2D6, NAT1 and NAT2 metabolic enzymes genes. To the best of our knowledge, this is the first report on polymorphisms in CYP1A1 6310C>T, CYP2D6 Arg365His, NAT1 52936A>T and NAT2 Arg268Lys (NAT2*12A) genes and susceptibility to HNC in Tunisian population. We study the prevalence of these polymorphisms in 169 patients with HNC and 261 control subjects using polymerase chain reaction based methods in a Tunisian population. We detected an association between HNC and CYP1A1 6310C>T (TT) and CYP2D6 Arg365His (His/His) variant carriers (OR 1.75, P = 0.008 and OR 1.66, P = 0.016, respectively). No association was found between the polymorphisms genotypes of NAT1 52936T>A and NAT2 Arg268Lys and risk of HNC. An association between HNC and CYP1A1 (TT) genotype was found among patients with smoking (P = 0.011) and drinking habit (P = 0.009). The combinations of NAT1 (AT or AA) and NAT2 (AA) at-risk genotypes increased HNC risk (OR 4.23, P = 0.005 and OR 3.60, P = 0.048, respectively). However, the combinations of CYP1A1 (AA) and CYP2D6 (CC) genotypes decreased risk of HNC (OR 0.20; P = 0.006). Genetic polymorphisms in CYP1A1 and CYP2D6 may significantly associate with HNC in the Tunisian population. The results of this study suggest a possible gene–environment interaction for certain carcinogen metabolizing enzymes, but larger studies that fully evaluate the interaction are needed.