Fungi have been detected in 35 types of cancer, often intracellularly

Fungi have been detected in 35 types of cancer, often intracellularly

In a recent article published in the journal cell Researchers found fungal deoxyribonucleic acid (DNA) and cells at low levels in many human cancers, with cancer type-dependent variations in fungal community composition and fungal-bacteriome interactions.

Learn: Pan-cancer analyzes reveal cancer type-specific fungal ecologies and bacteriome interactions. Photo credit: Kateryna Kon / Shutterstock

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Studies have shown that tumors exhibit spatially heterogeneous, intracellular and polymicrobial communities. Sepich-Poore et al. showed that nutrient limitation in the tumor microenvironment (TME) and antibiotics induce selection pressures that influence the composition of fungal-bacterial-cancer immune cells.

Fungi appear to be important opportunistic pathogens that shape host immunity and infect cancer patients; However, they are understaffed. It also remains unknown whether they could be part of polymorphic microbiomes that represent cancer. This provided sufficient motivation to study clonal cancer evolution as a multispecies process and to characterize the pan-cancer mycobiome. Furthermore, since bacteria and fungi share symbiotic and antagonistic relationships in nature, studying their interactions in tumors could potentially provide synergistic diagnostic performance for certain cancers.

About studying

In the present study, researchers profiled fungal DNA in two large cohorts of cancer samples, the Weizmann (WIS) and Cancer Genome Atlas (TCGA) cohorts. They studied patients with 35 different types of cancer, obtained tissue, blood and plasma samples from 17,401 patients and proceeded to characterize their cancer mycobiome.

The WIS cohort included 1,183 formalin-fixed, paraffin-embedded (FFPE), or frozen tumor samples and normal adjacent tissue [(NAT); often paired)] from eight types of tissues obtained from lung, melanoma, ovary, breast, colon, brain, bone and pancreas, as well as normal breast tissue that is not cancer. The second cohort included whole genome sequencing (WGS) and ribonucleic acid sequencing (RNA-seq) data.

The team examined all cancer samples for the presence of fungi and characterized them using internal transcribed spacer 2 (ITS2) amplicon sequencing. In addition, they quantified fungal DNA using quantitative polymerase chain reaction (qPCR) of the 5.8S fungal ribosomal gene in a random subset of the WIS cohort, which included 261 tumor and 137 negative control samples. In addition, the team compared the data on the presence (or absence) of fungi at different taxonomic levels to estimate the normalized mutual intradomain information for the WIS cohort.

Studies have shown that bacteriomes, immunomes, and mycobiomes exhibit cancer type specificity. It is therefore likely that multidomain fungal clusters vary by cancer type. The team compared WIS overlapping fungal and bacterial genera in TCGA using a neural network method previously developed to estimate microbiome and metabolite co-occurrence.

The team also tested whether mycotypes were associated with immune responses, C1 to C6, previously identified in TCGA patients and patient survival. Additionally, they determined whether machine learning (ML) discriminated mycobiomes between and within cancer types. Finally, researchers applied differential abundance testing (DA) and ML between stage I and IV tumor mycobiomes.

Study results

All tumors tested had higher fungal loads than negative controls, but fungal loads differed between tumor types, with the highest fungal DNA loads occurring in breast and bone cancers. ITS2 amplicon and sequencing also found more fungal reads in all tumor types than in negative controls. Notably, colon and lung tumors had significantly higher fungal loads than NAT. The researchers found a similar trend in breast tumors compared to NAT and normal tissue.

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Compared to matched bacteriomes, tumor-specific fungi exhibited lower diversity and abundance. Interestingly, although fungi were present in all cancers examined, not all tumors showed a positive fungal signal. However, imaging showed that most fungi were intracellular, like intratumoral bacteria. Furthermore, mycobiome richness was lower for the WIS (amplicon) cohort than for the TCGA (shotgun metagenomic) cohort. Interestingly, four of seven cancers that shared WIS and TCGA showed significant positive correlations between intratumoral fungal and bacterial richness.

In contrast to bacteria, there is a lack of published fungal genomes, limiting inference of gene content from amplicon data. In addition, low fungal abundances in tumors make their functional characterization difficult. However, the study results pointed to Malassezia globosa, a species of fungus that promotes pancreatic oncogenesis. The researchers also found significant correlations between some fungal species and other parameters such as age, tumor subtypes and response to immunotherapy. However, the researchers were unable to determine the exact nature of these associations.

The researchers observed positive correlations between microbiomes and mycobiomes in several cancer types. However, their diversity, frequency and co-occurrence varied depending on the cancer type. This raises the possibility that, unlike the gut, TMEs are a non-competitive space for microbial colonization, which the researchers termed a “permissive” phenotype. They referred to these distinct fungal-bacterial-immune clusters driven by fungal co-occurrence as mycotypes. For example, breast cancer had the most significant co-occurrences of fungi and bacteria (96.5%), with Aspergillus and Malassezia as hubs.

Unsupervised analyzes revealed three mycotypes, namely F1 (Malassezia-Ramularia-Trichosporon), F2 (Aspergillus-Candida), and F3 (multigenera, including Yarrowia). Interestingly, the mycotype log ratios varied between the TCGA and WIS cancer types. Six of nine TCGA log ratios between domains were significantly correlated (e.g., fungal F1/F2 vs. bacterial F1/F2), suggesting similar shifts within multidomain ecologies across different human cancers and validating inferred co-occurrences. Furthermore, the log ratios of immune cells co-occurring with F1, F2, or F3 cluster fungi distinguished immune response subtypes.

Fungal-induced pancancer mycotypes exhibited distinct immune responses that stratified patient survival. Although sparse, these fungi were immunologically potent, analogous to programmed death cells (PD)1+ in immunotherapy. The associations of fungi with clinical parameters could enable the detection of early-stage cancers and support their clinical utility as potential biomarkers and therapeutic targets. Finally, DA assays revealed cancer stage-specific fungi for gastric, rectal, and kidney cancers among RNA-seq samples, while ML data supported the differentiation of gastric and kidney cancer stages.

Conclusions

The study provided the first analysis of plasma mycobiomes in early-stage cancer. The researchers discovered fungi in 35 types of cancer, and most fungi were located intracellularly in cancer and immune cells, analogous to intratumoral bacteria. Although they were unable to pinpoint the sources of fungi from cell-free plasma, these species could help diagnose cancer in its early stages. In addition, they discovered multiple fungal-bacterial immunoecologies across tumors. Interestingly, intratumoral fungi stratified clinical outcomes, including response to immunotherapy.

Reference:

• Pan-Krebs-Analysen zeigen krebstypspezifische Pilzökologien und Bakteriom-Interaktionen, Lian Narunsky-Haziza, Gregory D. Sepich-Poore, Ilana Livyatan, Omer Asraf, Cameron Martino, Deborah Nejman, Nancy Gavert, Jason E. Stajich, Guy Amit , Antonio González, Stephen Wandro, Gili Perry, Ruthie Ariel, Arnon Meltser, Justin P. Shaffer, Qiyun Zhu, Nora Balint-Lahat, Iris Barshack, Maya Dadiani, Einav N. Gal-Yam, Sandip Pravin Patel, Amir Bashan, Austin D. Swafford, Yitzhak Pilpel, Rob Knight, Ravid Straussman, Cell 2022, DOI: https://doi.org/10.1016/j.cell.2022.09.005, https://www.cell.com/cell/fulltext/S0092-8674(22)01127-8

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