Periodic Reporting for period 2 - MUTAVAC (Targeting cancer with mutanome based stem cell vaccine)
Berichtszeitraum: 2023-10-01 bis 2025-09-30
The limited efficacy of current immunotherapies is partly due to their inability to target cancer stem cells (CSCs), which drive tumor heterogeneity, immune escape, recurrence, and metastatic spread. CSCs are intrinsically resistant to conventional therapies (chemo and radiotherapy), targeted therapies, and immunotherapies such as immune checkpoint inhibitors and CAR T cells. Moreover, existing immunotherapies cannot address the extensive antigenic diversity present in tumors and CSCs. Consequently, CSCs remain a major unmet challenge in oncology, with no approved therapies specifically targeting this cell population.
IPSirius is pioneering a new generation of iPSC based cancer vaccines (iPVAC) capable of targeting and eliminating CSCs by re engaging the immune system against multiple neoantigens. iPVAC induces strong and durable innate and adaptive immune responses, preventing tumor relapse and metastatic dissemination. Its immunogenicity has been demonstrated in vivo across several aggressive and metastatic cancer models.
The MUTAVAC project builds on this platform by developing iPVAC Mut, a next generation vaccine incorporating cancer associated mutanomes. Using proprietary technology, IPSirius generates ex vivo high mutational loads representative of cancer genomic instability, particularly relevant for cancers such as BRCA mutated tumors and Lynch syndrome. iPVAC Mut also serves as a powerful discovery tool for novel cancer targets.
MUTAVAC further advances the iPVAC value chain through the development of a predictive in vitro immunogenicity bioassay, enabling patient stratification, response prediction, and immune monitoring during clinical development.
By strengthening R&D and expanding into new therapeutic frontiers, the project is expected to generate new products, services, and revenue streams, including:
-An iPSC based discovery platform for first in class cancer targets and immunotherapy pipelines.
-Advanced preclinical development of iPVAC products with optimized antitumor immune responses.
-A high value potency assay integrating T cell immune monitoring and biomarker profiling for precision medicine and clinical translation.
IPSirius is pioneering a new generation of iPSC based cancer vaccines (iPVAC) capable of targeting and eliminating CSCs by re engaging the immune system against multiple neoantigens. iPVAC induces strong and durable innate and adaptive immune responses, preventing tumor relapse and metastatic dissemination. Its immunogenicity has been demonstrated in vivo across several aggressive and metastatic cancer models.
The MUTAVAC project builds on this platform by developing iPVAC Mut, a next generation vaccine incorporating cancer associated mutanomes. Using proprietary technology, IPSirius generates ex vivo high mutational loads representative of cancer genomic instability, particularly relevant for cancers such as BRCA mutated tumors and Lynch syndrome. iPVAC Mut also serves as a powerful discovery tool for novel cancer targets.
MUTAVAC further advances the iPVAC value chain through the development of a predictive in vitro immunogenicity bioassay, enabling patient stratification, response prediction, and immune monitoring during clinical development.
By strengthening R&D and expanding into new therapeutic frontiers, the project is expected to generate new products, services, and revenue streams, including:
-An iPSC based discovery platform for first in class cancer targets and immunotherapy pipelines.
-Advanced preclinical development of iPVAC products with optimized antitumor immune responses.
-A high value potency assay integrating T cell immune monitoring and biomarker profiling for precision medicine and clinical translation.
DEVELOPMENT OF IPVAC MUT
The primary objective of the MUTAVAC project is to develop mutated versions of IPVAC (iPVAC Mut) to increase antigen diversity and enhance immune responses against multiple cancer associated targets. IPVAC is the first vaccine candidate developed by IPSirius and is based on engineered induced pluripotent stem cell (iPSC) lines shown to effectively target cancer stem cells (CSCs), resulting in a significant reduction of tumor burden and metastatic spread in vivo.
IPSirius has generated multiple iPSC lines from healthy donors as well as patients with hematological malignancies, oncogenic somatic mutations, or hereditary cancer predispositions using the Yamanaka reprogramming approach. Within MUTAVAC, highly mutated iPSC lines (iPVAC Mut) were generated from multiple donors through controlled exposure to the mutagen N ethyl N nitrosourea (ENU), following a proprietary IPSirius protocol. All iPSC lines were produced under research grade conditions, with mutagenesis optimized to induce multiple mutations while preserving cell viability.
KEY OUTCOMES – MUTAGENESIS AND GENOMIC CHARACTERIZATION
Three main outcomes were achieved. First, an ENU based in vitro mutagenesis process compatible with future clinical development was standardized. Six distinct iPSC lines were exposed to ENU at varying doses for 60 days and expanded for genomic analysis.
Second, cancer like mutational signatures were characterized by whole exome sequencing of parental iPSC lines (IPVAC) and ENU treated lines (iPVAC Mut). Between 150 and 300 mutations per line were identified, including pathogenic somatic variants and a tumor mutational burden of approximately 11.9 mutations/Mb, comparable to primary human cancers.
These data demonstrate that iPVAC Mut lines:
-Were successfully generated across all donor derived iPSCs
-Recapitulate genomic and transcriptomic features of primary cancers
-Harbor hundreds of novel cancer associated mutations
-Maintain mutational stability over multiple culture passages
DEVELOPMENT AND VALIDATION OF A HUMAN IMMUNE POTENCY BIOASSAY
A human immune potency bioassay was developed to assess T cell reactivity against the mutational diversity of iPVAC Mut candidates. A standardized Mixed Lymphocyte Reaction (MLR) assay was established using PBMCs from HLA A2 healthy donors provided by the French Blood Establishment.
The assay relies on co culture of antigen loaded dendritic cells derived from CD14⁺ monocytes with naïve CD8⁺ T cells, with immune activation quantified by IFN γ ELISPOT. Robust isolation, dendritic cell differentiation, and expansion of functional CD8⁺ T cells were demonstrated.
The MLR platform was validated using the highly immunogenic MART 1 peptide in PBMCs from ten HLA A2 donors, resulting in strong IFN γ secretion, priming of up to 22% MART 1 specific CD8⁺ T cells, and increased CD137 expression, confirming effective T cell activation.
IMMUNOGENICITY AND MECHANISM OF ACTION
The immunogenicity of IPVAC and iPVAC Mut was assessed using dendritic cells loaded with iPSC derived lysates. CD8⁺ T cells primed with these antigens exhibited cytotoxic activity against iPSCs and multiple cancer cell lines (SK MEL 5, MDA MB 231, SW 620). Notably, iPVAC Mut induced stronger cytotoxic responses than IPVAC, accompanied by increased expression of activation and effector markers (CD107a, CD137, IFN γ, TNF α).
Mechanistic insights were obtained through single cell RNA sequencing of CD8⁺ T cells, revealing over 70,000 distinct TCR α/β clonotypes and a 22% increase in clonotype diversity in iPVAC Mut relative to IPVAC. This was associated with enhanced neoantigen diversity, increased NKG7 expression, and elevated granzyme A production.
PORTFOLIO ACTIVITY PLAN
To address genetic instability risks inherent to iPSC based therapies, IPSirius implemented Optical Genome Mapping (OGM). Genome integrity analysis of 50 cGMP iPSC samples across multiple stages revealed structural variants undetected by conventional karyotyping.
These results confirm OGM as a robust tool for baseline genomic profiling and quality control of iPSC master cell lines. While regulatory acceptance criteria for ATMPs are still evolving, relevant parameters include the absence of large (>1 Mb) oncogenic rearrangements, tumor associated translocations, and unexpected aneuploidies, consistent with EMA and FDA expectations.
The primary objective of the MUTAVAC project is to develop mutated versions of IPVAC (iPVAC Mut) to increase antigen diversity and enhance immune responses against multiple cancer associated targets. IPVAC is the first vaccine candidate developed by IPSirius and is based on engineered induced pluripotent stem cell (iPSC) lines shown to effectively target cancer stem cells (CSCs), resulting in a significant reduction of tumor burden and metastatic spread in vivo.
IPSirius has generated multiple iPSC lines from healthy donors as well as patients with hematological malignancies, oncogenic somatic mutations, or hereditary cancer predispositions using the Yamanaka reprogramming approach. Within MUTAVAC, highly mutated iPSC lines (iPVAC Mut) were generated from multiple donors through controlled exposure to the mutagen N ethyl N nitrosourea (ENU), following a proprietary IPSirius protocol. All iPSC lines were produced under research grade conditions, with mutagenesis optimized to induce multiple mutations while preserving cell viability.
KEY OUTCOMES – MUTAGENESIS AND GENOMIC CHARACTERIZATION
Three main outcomes were achieved. First, an ENU based in vitro mutagenesis process compatible with future clinical development was standardized. Six distinct iPSC lines were exposed to ENU at varying doses for 60 days and expanded for genomic analysis.
Second, cancer like mutational signatures were characterized by whole exome sequencing of parental iPSC lines (IPVAC) and ENU treated lines (iPVAC Mut). Between 150 and 300 mutations per line were identified, including pathogenic somatic variants and a tumor mutational burden of approximately 11.9 mutations/Mb, comparable to primary human cancers.
These data demonstrate that iPVAC Mut lines:
-Were successfully generated across all donor derived iPSCs
-Recapitulate genomic and transcriptomic features of primary cancers
-Harbor hundreds of novel cancer associated mutations
-Maintain mutational stability over multiple culture passages
DEVELOPMENT AND VALIDATION OF A HUMAN IMMUNE POTENCY BIOASSAY
A human immune potency bioassay was developed to assess T cell reactivity against the mutational diversity of iPVAC Mut candidates. A standardized Mixed Lymphocyte Reaction (MLR) assay was established using PBMCs from HLA A2 healthy donors provided by the French Blood Establishment.
The assay relies on co culture of antigen loaded dendritic cells derived from CD14⁺ monocytes with naïve CD8⁺ T cells, with immune activation quantified by IFN γ ELISPOT. Robust isolation, dendritic cell differentiation, and expansion of functional CD8⁺ T cells were demonstrated.
The MLR platform was validated using the highly immunogenic MART 1 peptide in PBMCs from ten HLA A2 donors, resulting in strong IFN γ secretion, priming of up to 22% MART 1 specific CD8⁺ T cells, and increased CD137 expression, confirming effective T cell activation.
IMMUNOGENICITY AND MECHANISM OF ACTION
The immunogenicity of IPVAC and iPVAC Mut was assessed using dendritic cells loaded with iPSC derived lysates. CD8⁺ T cells primed with these antigens exhibited cytotoxic activity against iPSCs and multiple cancer cell lines (SK MEL 5, MDA MB 231, SW 620). Notably, iPVAC Mut induced stronger cytotoxic responses than IPVAC, accompanied by increased expression of activation and effector markers (CD107a, CD137, IFN γ, TNF α).
Mechanistic insights were obtained through single cell RNA sequencing of CD8⁺ T cells, revealing over 70,000 distinct TCR α/β clonotypes and a 22% increase in clonotype diversity in iPVAC Mut relative to IPVAC. This was associated with enhanced neoantigen diversity, increased NKG7 expression, and elevated granzyme A production.
PORTFOLIO ACTIVITY PLAN
To address genetic instability risks inherent to iPSC based therapies, IPSirius implemented Optical Genome Mapping (OGM). Genome integrity analysis of 50 cGMP iPSC samples across multiple stages revealed structural variants undetected by conventional karyotyping.
These results confirm OGM as a robust tool for baseline genomic profiling and quality control of iPSC master cell lines. While regulatory acceptance criteria for ATMPs are still evolving, relevant parameters include the absence of large (>1 Mb) oncogenic rearrangements, tumor associated translocations, and unexpected aneuploidies, consistent with EMA and FDA expectations.
SCIENTIFIC IMPACT
Our results demonstrate that cancer associated mutational profiles can be reproducibly generated in vitro by exposing induced pluripotent stem cells (iPSCs) to the mutagen ENU. To our knowledge, this represents the first pluripotent stem cell based platform capable of reproducing mutational patterns commonly observed in human tumors.
This disruptive technology enables the discovery of novel immune and therapeutic targets derived from tumor specific mutational signatures and opens new avenues for first in class immunotherapies, including cancer vaccines, TCR based adoptive cell therapies, and CAR based approaches, particularly for aggressive and metastatic cancers.
IPSirius has established a strong intellectual property position, with a portfolio of 31 patents across 12 countries covering pluripotent stem cell based universal cancer vaccines. Outcomes from MUTAVAC are expected to further strengthen this portfolio through additional patent filings.
INDUSTRIALIZATION AND COMMERCIALIZATION
For clinical translation, IPSirius has identified a CDMO for manufacturing clinical grade IPVAC products and a CRO to support development in compliance with EMA requirements. The clinical strategy and commercial potential of the lead IPVAC product have been assessed in key markets (US, EU, UK, Canada, Australia), with market analyses estimating maximum annual revenue potential based on pricing assumptions.
ECONOMIC IMPACT
IPSirius is a pioneer in the use of allogeneic iPSC technologies to develop a new generation of off the shelf cancer vaccines. The antigen agnostic and scalable nature of this approach offers significant economic advantages, enabling reductions in production and treatment costs.
Our results demonstrate a standardized, scalable, and industrially compatible manufacturing process for IPVAC and iPVAC Mut products. The industrialization of off the shelf cancer vaccines represents a major breakthrough with high potential impact, addressing critical unmet needs in the treatment of aggressive and metastatic cancers.
MARKET ACCESS AND SOCIETAL IMPACT
MUTAVAC aligns closely with European policy priorities, including Horizon Europe and the Health 2030 framework, by fostering innovation, improving patient outcomes, promoting sustainability through streamlined manufacturing, and contributing to economic growth and biomedical sovereignty.
Our results demonstrate that cancer associated mutational profiles can be reproducibly generated in vitro by exposing induced pluripotent stem cells (iPSCs) to the mutagen ENU. To our knowledge, this represents the first pluripotent stem cell based platform capable of reproducing mutational patterns commonly observed in human tumors.
This disruptive technology enables the discovery of novel immune and therapeutic targets derived from tumor specific mutational signatures and opens new avenues for first in class immunotherapies, including cancer vaccines, TCR based adoptive cell therapies, and CAR based approaches, particularly for aggressive and metastatic cancers.
IPSirius has established a strong intellectual property position, with a portfolio of 31 patents across 12 countries covering pluripotent stem cell based universal cancer vaccines. Outcomes from MUTAVAC are expected to further strengthen this portfolio through additional patent filings.
INDUSTRIALIZATION AND COMMERCIALIZATION
For clinical translation, IPSirius has identified a CDMO for manufacturing clinical grade IPVAC products and a CRO to support development in compliance with EMA requirements. The clinical strategy and commercial potential of the lead IPVAC product have been assessed in key markets (US, EU, UK, Canada, Australia), with market analyses estimating maximum annual revenue potential based on pricing assumptions.
ECONOMIC IMPACT
IPSirius is a pioneer in the use of allogeneic iPSC technologies to develop a new generation of off the shelf cancer vaccines. The antigen agnostic and scalable nature of this approach offers significant economic advantages, enabling reductions in production and treatment costs.
Our results demonstrate a standardized, scalable, and industrially compatible manufacturing process for IPVAC and iPVAC Mut products. The industrialization of off the shelf cancer vaccines represents a major breakthrough with high potential impact, addressing critical unmet needs in the treatment of aggressive and metastatic cancers.
MARKET ACCESS AND SOCIETAL IMPACT
MUTAVAC aligns closely with European policy priorities, including Horizon Europe and the Health 2030 framework, by fostering innovation, improving patient outcomes, promoting sustainability through streamlined manufacturing, and contributing to economic growth and biomedical sovereignty.