Periodic Reporting for period 4 - SCUBA CANCERS (Finding the genetic causes of contagious metastases under the sea)
Reporting period: 2021-01-01 to 2022-06-30
Clonally transmissible cancers are somatic cell lineages that are transmitted between individuals via the transfer of living cancer cells. There are only three known types of naturally occurring clonally transmissible cancers, one of which is a leukemia-like cancer found in marine bivalves, called hemic neoplasia (HN). HN in cockles Cerastoderma edule offers an unique opportunity, over the other naturally occurring transmissible cancers, for the discovery of the genetic drivers of cancer transmissibility. Using HN in cockles as a model for clonally transmissible cancers, this project intended to identify the genomic alterations and mutational processes that drive transmissible cancers to depart from their hosts and evolve as parasitic clonal lineages in the marine environment, for illuminating universal processes that make a cancer contagious, and to identify new/unexpected biological insights into the general mechanisms of cancer metastasis.
To investigate the origin and evolution of contagious cancers in common cockles, we collected 6,854 C. edule specimens and diagnosed 390 cases of BTN. We then generated a reference genome for the species and assessed genomic variation in the genomes of 61 BTN tumours. Analysis of tumour-specific variants confirmed the existence of two cockle BTN lineages with independent clonal origins, and gene expression patterns supported their status as haemocyte-derived marine leukaemias. Examination of mitochondrial DNA sequences revealed several mitochondrial capture events in BTN, as well as co-infection of cockles by different tumour lineages. Mutational analyses identified two lineage-specific mutational signatures, one of which resembles a signature associated with DNA alkylation. Karyotypic and copy number analyses uncovered genomes marked by pervasive instability and polyploidy. Whole-genome duplication, amplification of oncogenes CCND3 and MDM2, and deletion of the DNA alkylation repair gene MGMT, are likely drivers of BTN evolution. Characterization of satellite DNA identified elements with vast expansions in the cockle germ line, yet absent from BTN tumours, suggesting ancient clonal origins. Our study illuminates the evolution of contagious cancers under the sea, and reveals long-term tolerance of extreme instability in neoplastic genomes.
Finally, to investigate the limits of marine contagious cancers, we collected 345 warty venus clams for which we described a contagious cancer present in two distant locations that originated in a different species, the striped venus clam. We performed whole-genome sequencing in eight warty venus clams that were diagnosed with HN, from two sampling points located more than 1000 nautical miles away in the Atlantic Ocean and the Mediterranean Sea Coasts of Spain. Mitochondrial genome sequencing analysis from neoplastic animals revealed the coexistence of haplotypes from two different clam species. Phylog- enies estimated from mitochondrial and nuclear markers confirmed this leukaemia originated in striped venus clams and later transmitted to clams of the species warty venus, in which it survives as a contagious cancer. The analysis of mitochondrial and nuclear gene sequences supports all studied tumours belong to a single neoplastic lineage that spreads in the Seas of Southern Europe.
To investigate the origin and evolution of contagious cancers in common cockles, we collected 6,854 C. edule specimens and diagnosed 390 cases of BTN. We then generated a reference genome for the species and assessed genomic variation in the genomes of 61 BTN tumours. Analysis of tumour-specific variants confirmed the existence of two cockle BTN lineages with independent clonal origins, and gene expression patterns supported their status as haemocyte-derived marine leukaemias. Examination of mitochondrial DNA sequences revealed several mitochondrial capture events in BTN, as well as co-infection of cockles by different tumour lineages. Mutational analyses identified two lineage-specific mutational signatures, one of which resembles a signature associated with DNA alkylation. Karyotypic and copy number analyses uncovered genomes marked by pervasive instability and polyploidy. Whole-genome duplication, amplification of oncogenes CCND3 and MDM2, and deletion of the DNA alkylation repair gene MGMT, are likely drivers of BTN evolution. Characterization of satellite DNA identified elements with vast expansions in the cockle germ line, yet absent from BTN tumours, suggesting ancient clonal origins. Our study illuminates the evolution of contagious cancers under the sea, and reveals long-term tolerance of extreme instability in neoplastic genomes.
Finally, to investigate the limits of marine contagious cancers, we collected 345 warty venus clams for which we described a contagious cancer present in two distant locations that originated in a different species, the striped venus clam. We performed whole-genome sequencing in eight warty venus clams that were diagnosed with HN, from two sampling points located more than 1000 nautical miles away in the Atlantic Ocean and the Mediterranean Sea Coasts of Spain. Mitochondrial genome sequencing analysis from neoplastic animals revealed the coexistence of haplotypes from two different clam species. Phylog- enies estimated from mitochondrial and nuclear markers confirmed this leukaemia originated in striped venus clams and later transmitted to clams of the species warty venus, in which it survives as a contagious cancer. The analysis of mitochondrial and nuclear gene sequences supports all studied tumours belong to a single neoplastic lineage that spreads in the Seas of Southern Europe.
During the 5 years of SCUBA CANCERS project we have met the planned goals and unraveled unexpected findings. A list of the major results follow:
Prevalence of disseminated neoplasia in common cockles
Reference genome and transcriptome of the common cockle
Two transmissible cancer lineages propagate through cockle populations
Haemocytic origin of cockle transmissible neoplasia
Mitochondrial transfer delineates the clonal structure of CedBTN
Lineage-specific mutational processes operate in CedBTN
Pervasive genomic instability drives the evolution of CedBTN
Satellite DNA expansions illuminate the emergence of CedBTN
Leukaemia-like cancer in warty venus clams
Mitochondrial sequencing reveals cancer contagion in warty venus clams
Nuclear evidence of cancer contagion in warty venus clams
Cancer inspection in the origin species (C. gallina) of warty venus clam cancer
Two publications report all these findings:
-Bruzos et al. (2022)
-García Souto et al. (2022)
Prevalence of disseminated neoplasia in common cockles
Reference genome and transcriptome of the common cockle
Two transmissible cancer lineages propagate through cockle populations
Haemocytic origin of cockle transmissible neoplasia
Mitochondrial transfer delineates the clonal structure of CedBTN
Lineage-specific mutational processes operate in CedBTN
Pervasive genomic instability drives the evolution of CedBTN
Satellite DNA expansions illuminate the emergence of CedBTN
Leukaemia-like cancer in warty venus clams
Mitochondrial sequencing reveals cancer contagion in warty venus clams
Nuclear evidence of cancer contagion in warty venus clams
Cancer inspection in the origin species (C. gallina) of warty venus clam cancer
Two publications report all these findings:
-Bruzos et al. (2022)
-García Souto et al. (2022)
Despite several BTN clones having been newly described in recent years, no analyses of whole BTN genomes have yet been reported. Combining a range of approaches, our study provides a first outlook into the genomes of these singular marine leukaemias in European common cockles, complementing the work of Hart et al. on American soft-shell clams. Both studies reveal neoplastic genomes marked by structural instability. In the case of cockle BTN, we find evidence for ongoing extreme genomic instability, most likely activated by early whole-genome duplication and fuelled by recurrent chromosome mis-segregation during mitosis. This is in stark contrast with the three transmissible cancers described in terrestrial mammals (dogs and Tasmanian devils), which present remarkable karyotypic stability, and thus challenges the notion that development of a durable genome architecture is required for long-term survival of cancer lineages. Although our data do not allow estimation of precise ages for cockle BTN, multiple lines of evidence suggest that these clones may have emerged centuries or millennia ago. These include the broad geographic distribution of tumours, the marked genetic divergence between tumours and modern cockles, the recurrent capture of host mitochondria by tumours, and the absence in tumours of satellite DNA elements that are vastly expanded in the cockle germ line. Furthermore, Hart et al. estimate an age of ~500 years for the BTN clone affecting soft-shell clams, supporting that long-term survival of marine transmissible cancers is possible. Taken together, our findings suggest that CedBTN lineages have undergone a relatively long history of pervasive genomic instability. Studying the mechanisms that enable BTN cells to overcome the effects of such instability will broaden our understanding of the conditions required for cancers to survive and adapt over the long term.
In adiition, the results of warty venus clams reveal the existence of a transmissible leukemia originated in a striped venus clam, most likely C. gallina, which was transmitted to a second species, the warty venus clam (V. verrucosa), and among whose specimens it currently propagates. We identified this parasitic cancer in warty venus clams from two sampling points that are more than 1,000 nautical miles away in the coasts of Spain, bathed by two different seas, the Atlantic Ocean and the Mediterranean Sea. The analysis of mitochondrial and nuclear gene sequences revealed no nucleotide diversity within the seven tumours sequenced, which supports that all belong to the same neoplastic lineage that spreads between Veneridae clams in the Seas of Southern Europe. Although we ignore the age of this cancer clone, we can confirm it arose before 2011, when the neoplastic warty venus specimen EVVV11-02 was collected. The apparent lack of genetic variation between all tumours, even from distant sampling points, suggests either that this cancer is very recent, or that it may have been unintentionally scattered by the action of man, a way of transmission that has been proposed for other bivalve transmissible cancers.
In adiition, the results of warty venus clams reveal the existence of a transmissible leukemia originated in a striped venus clam, most likely C. gallina, which was transmitted to a second species, the warty venus clam (V. verrucosa), and among whose specimens it currently propagates. We identified this parasitic cancer in warty venus clams from two sampling points that are more than 1,000 nautical miles away in the coasts of Spain, bathed by two different seas, the Atlantic Ocean and the Mediterranean Sea. The analysis of mitochondrial and nuclear gene sequences revealed no nucleotide diversity within the seven tumours sequenced, which supports that all belong to the same neoplastic lineage that spreads between Veneridae clams in the Seas of Southern Europe. Although we ignore the age of this cancer clone, we can confirm it arose before 2011, when the neoplastic warty venus specimen EVVV11-02 was collected. The apparent lack of genetic variation between all tumours, even from distant sampling points, suggests either that this cancer is very recent, or that it may have been unintentionally scattered by the action of man, a way of transmission that has been proposed for other bivalve transmissible cancers.