The Role of the p16 and p53 Tumor Suppressor Proteins and Viral HPV16 E6 and E7 Oncoproteins in the Assessment of Survival in Patients with Head and Neck Cancers Associated with Human Papillomavirus Infections

Funding Information: This research was supported by the European Social Fund and the Latvian state budget within the frame of the project No. 8.2.2.0/20/l/004 “Support for involving doctoral students in scientific research and studies” at Riga Stradiņš University. The article processing charge was covered by Riga Stradinš University Department of Otorhinolaryngology. Publisher Copyright: © 2023 by the authors.The role of HPV in the survival prognosis of patients with head and neck squamous cell carcinoma, especially patients with laryngeal squamous cell carcinoma (LSCC) and hypopharyngeal squamous cell carcinoma (HPSCC), is still somewhat ambiguous. The present study aimed to explore the significance of tumor suppressor proteins and HPV16 E6 and E7 oncoproteins in the assessment of survival in patients with oropharyngeal squamous cell carcinoma (OPSCC), LSCC, and HPSCC associated with high-risk (HR-) and low-risk (LR-) HPV infections. By utilizing molecular and immunohistochemical investigations of HNSCC samples and patient data, univariate and multivariate survival analyses were conducted. The presence of HPV DNA (LR- and HR-HPV) was associated with a better 5-year OS and DSS for OPSCC and LSCC. The IHC overexpression of HPV16 E6 protein and p16 protein was associated with better survival in the univariate (for OPSCC) and multivariate (OPSCC and HPSCC) survival analyses. The overexpression of p53 was associated with better survival in OPSCC. HPV infection plays a significant role in the tumorigenesis of HNSCC, and the immunohistochemical assessment of HPV16 E6 protein expression should be interpreted as a useful prognostic marker for OPSCC and HPSCC.publishersversionPeer reviewe

Survival analysis of oropharyngeal squamous cell carcinoma patients linked to histopathology, disease stage, tumor stage, risk factors, and received therapy

Publisher Copyright: Copyright © Experimental Oncology, 2020. Copyright: Copyright 2020 Elsevier B.V., All rights reserved.Background: Survival of oropharyngeal squamous cell carcinoma (OSCC) patients depends on the risk and environmental factors, tumor biology, achievements in diagnostics and treatment approaches. Aim: To perform a survival analysis of the patients with OSCC treated over a 10-year period in a single hospital in Latvia linking these data to histopathological findings, risk factors and received therapy. Materials and Methods: The main outcome measures were overall and disease-specific survival (OS and DS) along with histopathology analysis. Results: Kaplan - Meier survival analysis showed better survival for females, younger patients lacking bad habits, operated and received radiotherapy, with lower T grade and disease stage. Cox regression showed diminished early death risk in patients with lower T grade, no regional metastases (N0) and bad habits, operated and received radiotherapy. A vast majority of tumors were localized in palatine tonsils and the base of the tongue. The localization did not correlate with mean survival time/survival. Lower OS (p = 0.03) and DS (p = 0.026) were estimated for patients with pharyngeal wall and tonsillar involvement compared to tumors localized in the soft palate. A histological variant of tumor seemed irrelevant estimating OS and DS, whereas therapeutic modalities significantly affected survival. Conclusions: OSCC patients with lower T grade, N0 status, lacking bad habits, and surgically treated had better survival.publishersversionPeer reviewe

Identification of High-Risk Human Papillomavirus DNA, p16, and E6/E7 Oncoproteins in Laryngeal and Hypopharyngeal Squamous Cell Carcinomas

Funding Information: The authors would like to thank the Riga Stradi?? University Department of Doctoral studies for reagents and publishing support. We would also like to thank Elza Rate, MD at the Department of Head and Neck Surgery, Oncology Centre of Latvia, for additional support with the collection of some tumor specimens. We are grateful to Simons Svirskis for his valuable help with statistical analysis. Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland.Human papillomavirus (HPV) was proven to play a significant role in cancer development in the oropharynx. However, its role in the development of laryngeal (LSCC) and hypopharyngeal squamous cell carcinoma (HPSCC) remains to be clarified. High-risk HPV (HR-HPV) viral proteins E6 and E7 are considered to be pertinent to HPV-related carcinogenesis. Hence, our aim was to estimate LSCC and HPSCC for HR-HPV DNA, p16, and E6/E7 oncoprotein status by using molecular virology and immunohistochemistry methods. The prevalence of HPV16 infection was 22/41 (53.7%) and 20/31 (64.5%) for LSCC and HPSCC, accordingly. The majority of HPV16+ tumor samples were stage III or IV. In most samples, the presence of either HPV16 E6 or HPV16 E7 viral protein in dysplastic or tumor cells was confirmed using immunohistochemistry. Our results suggest a high prevalence of HPV16 as a primary HR-HPV type in LSCC and HPSCC. The lack of HPV E6/E7 oncoproteins in some tumor samples may suggest either the absence of viral integration or the presence of other mechanisms of tumorigenesis. The utilization of p16 IHC as a surrogate marker of HR-HPV infection is impractical in LSCC and HPSCC.publishersversionPeer reviewe

Argon laser microirradiation of mitochondria in rat myocardial cells. VI. Correlation of contractility and ultrastructure.

A correlation is made between individual laser microbeam lesions (0.25-1 μm in diameter) in single mitochondria of myocardial cells and alterations in cellular contractility. A moderate lesion is described ultrastructurally that has three concentric zones of damage. In addition, the outer mitochondrial membrane is broken, and the intercristae matrix outside the lesion may be affected. Two kinds of weak lesions are described. No concentric zones of damage are observed, and the cristae and mitochondrial membranes outside the lesion area are not grossly affected. All cells that exhibited an alteration in contractile patterns had moderate lesions. A correlation between laser energy density and type of lesion was observed. The nature of the lesions and explanations of the contractile responses are discussed. © 1976

DNA replication defect in Salmonella typhimurium mutants lacking the editing (epsilon) subunit of DNA polymerase III.

In Salmonella typhimurium, dnaQ null mutants (encoding the epsilon editing subunit of DNA polymerase III [Pol III]) exhibit a severe growth defect when the genetic background is otherwise wild type. Suppression of the growth defect requires both a mutation affecting the alpha (polymerase) subunit of DNA polymerase III and adequate levels of DNA polymerase I. In the present paper, we report on studies that clarify the nature of the physiological defect imposed by the loss of epsilon and the mechanism of its suppression. Unsuppressed dnaQ mutants exhibited chronic SOS induction, indicating exposure of single-stranded DNA in vivo, most likely as gaps in double-stranded DNA. Suppression of the growth defect was associated with suppression of SOS induction. Thus, Pol I and the mutant Pol III combined to reduce the formation of single-stranded DNA or accelerate its maturation to double-stranded DNA. Studies with mutants in major DNA repair pathways supported the view that the defect in DNA metabolism in dnaQ mutants was at the level of DNA replication rather than of repair. The requirement for Pol I was satisfied by alleles of the gene for Pol I encoding polymerase activity or by rat DNA polymerase beta (which exhibits polymerase activity only). Consequently, normal growth is restored to dnaQ mutants when sufficient polymerase activity is provided and this compensatory polymerase activity can function independently of Pol III. The high level of Pol I polymerase activity may be required to satisfy the increased demand for residual DNA synthesis at regions of single-stranded DNA generated by epsilon-minus pol III. The emphasis on adequate polymerase activity in dnaQ mutants is also observed in the purified alpha subunit containing the suppressor mutation, which exhibits a modestly elevated intrinsic polymerase activity relative to that of wild-type alpha

Argon laser microirradiation of mitochondria in rat myocardial cells. VI. Correlation of contractility and ultrastructure.

A correlation is made between individual laser microbeam lesions (0.25-1 μm in diameter) in single mitochondria of myocardial cells and alterations in cellular contractility. A moderate lesion is described ultrastructurally that has three concentric zones of damage. In addition, the outer mitochondrial membrane is broken, and the intercristae matrix outside the lesion may be affected. Two kinds of weak lesions are described. No concentric zones of damage are observed, and the cristae and mitochondrial membranes outside the lesion area are not grossly affected. All cells that exhibited an alteration in contractile patterns had moderate lesions. A correlation between laser energy density and type of lesion was observed. The nature of the lesions and explanations of the contractile responses are discussed. © 1976

Smoking and alcohol abuse – predictive factors in oropharyngeal squamous cell carcinoma: A retrospective study

Oropharynx is one of the most common localizations for malignant neoplasms in the head and neck region. HPV-positive OPSCCs are the most common subtypes in Western countries with tobacco and alcohol use, remaining the major risk factors for cancer development in this region. The study aims to address the significance of smoking and alcohol abuse in OPSCC cases and perform a survival analysis of the patients with OPSCC treated over a 10-year period in a single hospital in Latvia, correlating these data with patients' hazardous habits (smoking and alcohol abuse). Out of 247 patients, the majority were smokers (75.95%, n = 180), approximately one third were habitual drinkers (35.19%, n = 82) or were exposed to both aforementioned major risk factors (31.47%, n = 73). Kaplan-Meier survival analysis showed significantly better survival for the patients without the aforementioned hazardous habits. Cox regression analysis proved a higher early death rate when both factors (smoking and alcohol abuse) were present. It seems that smoking has a higher predictive value than alcohol abuse

Nucleotide sequences of dnaE, the gene for the polymerase subunit of DNA polymerase III in Salmonella typhimurium, and a variant that facilitates growth in the absence of another polymerase subunit.

The dnaE gene of Salmonella typhimurium, like that of Escherichia coli, encodes the alpha subunit containing the polymerase activity of the principal replicative enzyme, DNA polymerase III. This gene, or one nearby, has been identified as the locus of suppressor mutations that promote growth by cells deleted for dnaQ, the gene for the editing subunit of this enzyme complex. Using a combination of nucleotide sequencing and marker rescue experiments, the alteration in one such suppressor was identified as a valine-to-glycine substitution at amino acid 832 of the 1,160-amino-acid alpha polypeptide. The alpha polypeptides of E. coli and S. typhimurium are identical in size and in 97% of their amino acid residues. Their identity includes the valine residue that was changed in the suppressor allele of S. typhimurium. We also localized a temperature-sensitive dnaE mutation to the 3' half of dnaE

Isolation and characterization of mutants with deletions in dnaQ, the gene for the editing subunit of DNA polymerase III in Salmonella typhimurium.

dnaQ (mutD) encodes the editing exonuclease subunit (epsilon) of DNA polymerase III. Previously described mutations in dnaQ include dominant and recessive mutator alleles as well as leaky temperature-sensitive alleles. We describe the properties of strains bearing null mutations (deletion-substitution alleles) of this gene. Null mutants exhibited a growth defect as well as elevated spontaneous mutation. As a consequence of the poor growth of dnaQ mutants and their high mutation rate, these strains were replaced within single colonies by derivatives carrying an extragenic suppressor mutation that compensated the growth defect but apparently not the mutator effect. Sixteen independently derived suppressors mapped in the vicinity of dnaE, the gene for the polymerization subunit (alpha) of DNA polymerase III, and one suppressor that was sequenced encoded an altered alpha polypeptide. Partially purified DNA polymerase III containing this altered alpha subunit was active in polymerization assays. In addition to their dependence on a suppressor mutation affecting alpha, dnaQ mutants strictly required DNA polymerase I for viability. We argue from these data that in the absence of epsilon, DNA replication falters unless secondary mechanisms, including genetically coded alteration in the intrinsic replication capacity of alpha and increased use of DNA polymerase I, come into play. Thus, epsilon plays a role in DNA replication distinct from its known role in controlling spontaneous mutation frequency