Uncovering How HPV Alters the Genome in Ugandan Cervical Cancer Patients
, by Nick Griner, Ph.D. and Peggy I. Wang, Ph.D.
While human papillomavirus (HPV) infection is very common, persistent infection with certain types of the virus can cause many types of cancer, including cervical cancer. However, the exact molecular mechanisms by which HPV infection leads to cancer, especially in African populations which suffer high incidence and mortality rates, have yet to be clearly delineated.
A recent study in Nature Genetics describes a large-scale genomic, epigenomic, and transcriptomic study of 118 cervical tumors from Ugandan patients, including 72 from patients who had tested positive for human immunodeficiency virus (HIV+). An additional 89 cases, of which 16 were HIV+, received targeted sequencing to help validate findings. The work was performed by NCI’s HIV+ Tumor Molecular Characterization Project (HTMCP), a collaboration between the Office of Cancer Genomics (OCG) and the Office of HIV and AIDS Malignancies aiming to understand the genetic events driving HIV-associated cancers.
Characterizing HIV+ and HIV- Cervical Cancer in an Understudied Population
People infected with HIV have a substantially higher risk of cervical and other types of cancer. By comparing HIV+ and HIV- tumors, researchers hoped to identify important molecular characteristics that could inform diagnostics and how the virus may influence HPV infection or cancer prognosis.
Somewhat surprisingly, the tumors in the study did not have significant differences in mutation burden or signatures (characteristic combinations of mutation types) between the HIV+ and HIV- cases. PIK3CA, a recurrent significantly mutated gene in many cancers, was altered in a higher proportion of HIV- tumors.
Comparing copy number alterations, while many were shared between the two groups, HIV+ tumors exhibited more unique focal amplifications and deletions than the HIV- tumors.
Comparing the genomic data from this cohort to those from The Cancer Genome Atlas (TCGA) cervical study, which included patients of mostly American European ancestry, showed some differences, again mainly in copy number variations. The TCGA samples exhibited a larger number of deleted regions affecting 11 chromosomes. These data suggest that a paradigm of how genetic background, in addition to infection with HIV and HPV, may affect copy number changes is worth exploring.
High-Risk HPV Types Dominate the Ugandan Cohort
The researchers also examined the distribution of HPV types in the cohort. There are over 170 types of HPV that have previously been identified, which can be classified into genetic groupings called clades.
17 HPV types were detected in the cohort, with types belonging to known high-risk clade A7 (HPV types 18 and 45) and clade A9 (HPV type 16) among the most abundant. Clade A7 cervical cancers are more clinically aggressive than clade A9 cancers.
Clade A7 was found to be more prevalent in this Ugandan cohort, particularly among squamous cell carcinomas (SCCs), compared to TCGA’s study. Somewhat unexpectedly, no difference in HPV types were found between HIV+ and HIV- tumors.
Gene expression and methylation analyses suggested tumors infected with different clades were undergoing different cellular activities. Clade A7-infected samples—again known to be more clinically aggressive—exhibited increased activity of extracellular matrix organization, cell adhesion and migration pathways. Clade A9 samples showed epithelial differentiation, production of virus during HPV infection and uncontrolled cell growth.
HPV Integration Sites May Epigenetically Alter Gene Expression and Contribute to Tumorigenesis
Clear differences were found when comparing epigenomes of tumors of different HPV clades. The researchers started by examining the locations and genomic impacts of HPV integration sites for many of the tumors. After grouping these sites by proximity to each other, the researchers identified 257 so-called “integration events”.
Many genes near the integration events showed higher expression, especially if events had a higher number of integration sites. Clade-A7 samples, which had significantly more HPV integrations sites per event than clade-A9 samples, showed a more pronounced effect on expression than clade-A9 samples.
Changes in histone modifications near integration sites suggest epigenomic mechanisms may be at play in altering gene expression of the tumors. The researchers checked for changes in histone mark enrichment near integration events for about a quarter of the samples.
Genes near the integration events with increased expression also showed increased enrichment of active histone marks—epigenetic modifications associated with active gene transcription. The number of integration sites per event was also associated with increased histone mark enrichment and increased gene expression.
Finally, for integration events that were in non-coding regions, the researchers found endogenous retroviral (ERV) sequences could be at play. The ERVs nearby integration events showed similar increased expression and enrichment of active histone marks.
Beyond Uganda: Still Much to Understand about Viruses and Cancer
This study produced 118 cancer whole genomes, transcriptomes, and epigenomes, one of the largest number of whole genome sequences of invasive cervical cancer available and one of the few specific to African populations.
The study started to uncover some of the HPV clade-specific differences in cervical tumors, including epigenetic changes, which may help explain the difference in prognosis for the groups. From the expression and epigenetic analyses, a model is emerging where A7-infected tumors may be less differentiated, more likely to have viral transcripts integrated into the genome, and more aggressive with a poor prognosis. In contrast, A9-infected tumors may be more epithelial differentiated and have more non-integrated viral transcripts.
A more complete picture of how specific HPV strains affect disease development may help researchers understand the poor prognosis linked to clade A7-infected tumors, which may in turn shed light on the increased incidence and mortality of cervical cancer in African countries. Fortunately, the most recently approved HPV vaccines protect against the most abundant HPV strains found in this cohort, including clade A7, underscoring the protective value of the vaccine even in this under-studied population.
Still, much remains to be uncovered about the epigenomic effects of HPV, and the data produced in the study provides a launching pad for researchers. “This resource provides a wonderful opportunity to examine the non-coding regions of cervical cancer genomes,” says Dr. Janet Rader, who helped supervise the work. “We can start to examine how HPV alters the human genome to cause cancer progression and metastasis.”
Beyond this study in Uganda, HTMCP is looking to molecularly characterize other common tumor types that arise in HIV+ patients, such as diffuse large B cell lymphomas (DLBCLs) and non-small cell lung carcinomas (NSCLCs). The project continues to accrue these types of tumors in hopes of identifying key differences between HIV+ and HIV- patients. Tissue sources with these tumor types are encouraged to contact OCG for potential participation in HTMCP.
“The HTMCP studies of DLBCL and NSCLC will clarify if HIV-associated cancers have different rates of mutations in known cancer-causing genes and/or copy number changes in tumors from patients that have been diagnosed with HIV infection,” says Dr. Daniela S. Gerhard, director of OCG and a senior author of the study. “The effects of HIV in cancer etiology are still unclear, so HTMCP is still seeking new tissue source sites that may have tissues from HIV+ patients.”
The raw data from this study are available at NCI’s Genomic Data Commons and harmonized data will also be available soon for the research community.