ITEM 1A. Risk Factors 31
ITEM 1B. Unresolved Staff Comments 86
ITEM 1C. Cybersecurity 86
ITEM 2. Properties 87
ITEM 3. Legal Proceedings 87
ITEM 4. Mine Safety Disclosures 87
PART II
ITEM 6. [Reserved] 89
ITEM 7A. Quantitative and Qualitative Disclosures About Market Risk 94
ITEM 8. Financial Statements and Supplementary Data 94
ITEM 9A. Controls and Procedures 94
ITEM 9B. Other Information 95
ITEM 9C. Disclosure Regarding Foreign Jurisdictions that Prevent Inspections 95
PART III
ITEM 10. Directors, Executive Officers and Corporate Governance 96
ITEM 11. Executive Compensation 105
ITEM 14. Principal Accountant Fees and Services 113
PART IV
ITEM 15. Exhibits and Financial Statement Schedules 114
i
CAUTIONARY NOTE REGARDING FORWARD-LOOKING STATEMENTS
This Annual Report on
Form 10-K (this “Annual Report”) contains forward-looking statements within
the meaning of Section 27A of the Securities Act of 1933, as amended (the “Securities Act”), and Section 21E of the
Securities Exchange Act of 1934, as amended (the “Exchange Act”). All statements other than statements of
historical facts contained in this Annual Report, including statements regarding our future results of operations and financial
position, business strategy, drug candidates, planned preclinical studies and clinical trials, results of preclinical studies,
clinical trials, research and development (“R&D”) costs, regulatory approvals, timing and likelihood of success, as
well as plans and objectives of management for future operations, are forward-looking statements. These statements involve known and
unknown risks, uncertainties and other important factors that are in some cases beyond our control and may cause our actual results,
performance or achievements to be materially different from any future results, performance or achievements expressed or implied by
the forward-looking statements.
In some cases, you can identify
forward-looking statements by terms such as “may,” “will,” “should,” “would,” “expect,”
“plan,” “anticipate,” “could,” “intend,” “target,” “project,”
“believe,” “estimate,” “predict,” “potential,” or “continue” or the negative
of these terms or other similar expressions. Forward-looking statements contained in this Annual Report include, but are not limited to,
statements about:
● our financial performance;
● the rate and degree of market acceptance of our product candidates;
ii
● our ability to realize the anticipated benefits of any strategic transactions;
● our ability to maintain proper and effective internal controls;
We have based these forward-looking
statements largely on our current expectations and projections about our business, the industry in which we operate and financial trends
that we believe may affect our business, financial condition, results of operations and prospects, and these forward-looking statements
are not guarantees of future performance or development. These forward-looking statements speak only as of the date of this Annual Report
and are subject to a number of risks, uncertainties and assumptions described in “Risk Factors” and elsewhere in this
Annual Report. Because forward-looking statements are inherently subject to risks and uncertainties, some of which cannot be predicted
or quantified, you should not rely on these forward-looking statements as predictions of future events. The events and circumstances reflected
in our forward-looking statements may not be achieved or occur and actual results could differ materially from those projected in the
forward-looking statements. Except as required by applicable law, we do not plan to publicly update or revise any forward-looking statements
contained herein until after we distribute this Annual Report, whether as a result of any new information, future events or otherwise.
In addition, statements that
“we believe” and similar statements reflect our beliefs and opinions on the relevant subject. These statements are based upon
information available to us as of the date of this Annual Report, and while we believe such information forms a reasonable basis for such
statements, such information may be limited or incomplete, and our statements should not be read to indicate that we have conducted an
exhaustive inquiry into, or review of, all potentially available relevant information. These statements are inherently uncertain, and
you are cautioned not to unduly rely upon these statements.
This Annual Report includes
trademarks, tradenames and service marks that are the property of other organizations. Solely for convenience, trademarks and tradenames
referred to in this Annual Report appear without the ® and TM symbols, but those references are not intended to indicate, in
any way, that we will not assert, to the fullest extent under applicable law, our rights, or that the applicable owner will not assert
its rights, to these trademarks and tradenames.
Unless the context otherwise
requires, all references herein to “we,” “us,” or “our” refer to the business and operations of PBAX
and its subsidiaries prior to consummation of the Business Combination and to CERo Therapeutics Holdings, Inc. (“CERo”) and
its subsidiaries following the consummation of the Business Combination.
iii
RISK FACTORS SUMMARY
Our
business is subject to numerous risks and uncertainties that you should consider before investing in our securities. Some of the principal
risk factors are summarized below:
iv
● Our product candidates rely on the availability of specialty raw materials.
v
PART
I
Item 1. Business.
Overview
We
are an innovative immunotherapy company advancing the development of next-generation engineered T cell therapeutics for the treatment
of cancer. Our proprietary approach to T cell engineering, which enables us to integrate certain desirable characteristics of both innate
and adaptive immunity into a single therapeutic construct, is designed to engage the body’s full immune repertoire to achieve optimized
cancer therapy. Our novel cellular immunotherapy platform is designed to redirect patient-derived T cells to eliminate tumors by building
in engulfment pathways that employ phagocytic mechanisms to destroy cancer cells, creating what we refer to as CER-T cells. We believe
the differentiated activity of CER-T cells will afford them greater therapeutic application than currently approved chimeric antigen
receptor T (“CAR-T”) cell therapies, for use spanning both hematological malignancies and solid tumors. We are nearing completion
of extensive preclinical testing and studies which are needed to obtain regulatory clearance to initiate human clinical trials with CER-1236,
and have engaged in a pre-IND meeting with the Federal Drug Administration (the “FDA”). We anticipate filing an investigational new drug (“IND”) application
and, if allowed to proceed, initiating clinical trials for our lead drug candidate, CER-1236, in 2024. However, manufacturing delays
or other delays with IND-enabling studies, among other factors, may impact the timing and approval of such trials.
The
ability to enhance the activity of T cells against human cancers through genetic engineering has been among the most significant advances
in cancer therapy in the last decade. One of the more promising therapeutic uses of T cells to emerge has been CAR-T cell technology.
Yet as remarkable a development as CAR-T cell therapy has been, its use has been largely limited to the treatment of certain hematological
cancers due to CAR-T cells’ limited ability to proliferate, traffic, and circulate in solid tumors. Curative cell therapies for
solid tumors currently do not exist, and the significance of this limitation is underscored by the prevalence of solid tumor malignancies.
The American Cancer Society estimates that solid tumor cancers accounted for more than 1.7 million of the 1.9 million people
newly diagnosed with cancer in 2022. Even in hematological malignancies with approved CAR-T cell therapies, cure rates do not exceed
60%. Nevertheless, despite such limitations, sales of CAR-T cell therapies are anticipated to grow rapidly over the next several years
and are expected to exceed $10 billion globally by 2030.
We
believe that the preferential attributes engineered into our CER-T cell therapy enables us to overcome many of the limitations which
hinder the wider application of CAR-T technology. Our CER-T cells employ a novel targeting mechanism that enables the use of phagocytic
pathways. Specifically, they target phosphatidylserine (“PS”), a critical component of the cell’s plasma membrane that
has a key role in cell cycle regulation. Exposure of PS on the outer surface of the plasma membrane acts as an “eat-me” signal
and marks abnormal, stressed and dying or dead cells for phagocytosis. The pro-phagocytic activities of CER-T cells are designed to integrate
innate immune effector functions into cytotoxic killer T cells, creating within a single T cell the ability to directly mediate cytotoxic
effects and indirectly prime other immune cells. As externally oriented PS is ubiquitously expressed by numerous cancer cell types, we
believe a single CER-T construct may have broad clinical utility in treating an array of cancers. Moreover, in preclinical studies, we
have observed CER-T cells to exhibit superior cross-presentation abilities compared to conventional T cells, potentially triggering a
broad complement of immune effector cells against tumors. In consequence, we envision CER-T therapeutics as potentially having differentiated
therapeutic utility with application across a wide array of cancer types.
We
have patterned the design of our CER-T constructs based upon many of the components found in existing conventional CAR-T cell therapies,
which we believe could shorten development timelines and enhance commercial application. The processes and protocols used to genetically
modify a patient’s T cells to produce CAR-T cells are already well recognized, as is the use of lentivirus in the manufacture of
these therapies. Accordingly, we have developed CER-T cell manufacturing processes that closely resemble those used to produce existing
engineered CAR-T cells. We also expect to benefit from the well-defined and recognized regulatory guidelines established by both U.S. and
European regulatory authorities related to CAR-T therapy and its use. In contrast to these attributes, we believe that other emerging
CAR-based drug candidates which involve immune effector cells other than T cells, such as CAR-NK and CAR-M therapies, that are in the
earlier stages of clinical development are unlikely to enjoy similar benefits.
In
preclinical studies, we have observed CER-1236 to display attractive functional attributes, among which are:
● phagocytosis of tumor cells;
● enhanced antigen acquisition, processing and presentation;
1
● no evidence of T cell exhaustion despite repeated challenges;
● no observed off-target or off-tumor toxicities;
● well defined and scalable manufacturing protocols.
Based
on the preclinical data regarding the use of CER-1236 T cells to combat hematologic malignancies, we currently intend to file an IND
application to begin clinical trials in 2024. We anticipate that our initial targets will be relapsed, remitting acute myeloid leukemia
(“AML”) patients as well as aggressive, difficult-to-treat B cell malignancies, including aggressive mantle cell lymphoma
(“MCL”) and refractory chronic lymphocytic leukemia (“CLL”). AML is a heterogenous and aggressive hematopoietic
malignancy characterized by the rapid buildup of immature myeloid cells in the bone marrow and blood. This process results in the inhibition
of normal hematopoiesis, manifesting as neutropenia, anemia, thrombocytopenia, and the clinical features of bone marrow failure. AML
accounts for 90% of all acute leukemias in adults, with an estimated 20,240 new cases and 11,400 deaths expected in the United States
in 2023. The disease often presents with signs and symptoms related to infiltration of leukemic blasts into the bone marrow resulting
in infections and disruption of normal hematopiesis and is associated with a variety of laboratory derangements in addition to abnormal
blood counts. The current treatment has remained largely unchanged over several decades with combination chemotherapy with cytarabine
for 7 days and an anthracycline for 3 days (“7+3”). Newer, targeted approaches that include multi-kinase domain inhibitors
and antibody-drug conjugates are now available during induction chemotherapy for certain patients. For patients that are sufficiently
healthy and at unfavorable risk, allogeneic Hematopoietic Stem Cell Transplants (“HSCTs”) are commonly performed. Despite
these interventions, there is significant unmet medical need for novel therapies, including cell therapeutic approaches. In the difficult-to-treat
B-cell malignancies, durable responses with CAR-T cell therapy are often evasive and the high frequency of acute multi-organ complications
often limits its use, particularly among chronically ill or elderly patients. Existing FDA-approved CD19-targeted CAR-T cell therapies produce an overall response rate of between 50% and 80%. Our Phase 1 clinical trial
of around 25 patients, is intended to evaluate the safety, potential therapeutic utility and applicable dose of CER-1236. Following a
trial in these hematological malignancies, we intend to expand the clinical development of CER-1236 to include solid tumors such as non-small
cell lung cancer (“NSCLC”) and ovarian cancer. We believe that CER-1236 has the potential to be a therapy for the unmet needs
of targeted indications, if approved, and differentiated by its safety, tolerability, efficacy and clinical benefit over current therapeutic
alternatives, which have been observed in preclinical studies. None of the abovementioned statements regarding any of our products in
development are intended to be a prediction or conclusion of efficacy. No clinical trials on our product candidates have commenced so
no conclusions relating to such attributes can be made.
Our Strategy
Our
intent is to become a leading biopharmaceutical company focused on the capital-efficient advancement of innovative anti-cancer product
candidates targeting the unmet medical need associated with aggressive and difficult-to-treat hematological malignancies and solid tumors.
To accomplish this objective, the key elements of our strategy include:
2
The Immune
System and its Function
The
immune system is a host defense system comprising multiple structures and processes within an organism that protects against disease.
As with other mammalian species, the human immune system is segregated into two separate yet interconnected components, the innate immune
system and the adaptive immune system. The innate immune system is responsible for an immediate, non-specific response to infected or
diseased cells. Triggering its activation are pathogen-associated and damage-associated molecular patterns recognized by preconfigured
pattern recognition receptors which reside on the surface of various types of leukocytes, or white blood cells, that make up the innate
immune system, including macrophages, dendritic cells, eosinophils and natural killer (“NK”) cells. In addition to its direct
participation in eliminating damaged or diseased cells, certain components of the innate immune system function significantly as antigen-presenting
cells (“APCs”) promoting the activity of the adaptive immune system.
The
adaptive immune system is composed of special types of leukoctyes known as T and B lymphocytes, also known as T and B cells, respectively.
T cells participate primarily in the cell-mediated immune response while B cells are involved in the humoral immune response. T
cells are an essential component of the adaptive immune system, targeting specific antigens and either destroying targeted cells directly
or participating in their destruction by activating other immune cells. T cells use T cell specific receptors to recognize antigens presented
via major histocompatibility complex (“MHC”) molecules on APCs. Through this mechanism, T cells have the ability to target
tumor-transformed or virus infected cells, as well as help coordinate the activity of other immune cells.
T
cells are differentiated by the expression of protein markers on their surface. The two most prominent types of T cells are those
that express CD8 molecules and are known as CD8 T cells, and those that express CD4 molecules and are known as CD4 T cells. CD8 T cells,
also referred to as cytotoxic lymphocytes (“CTLs”), eliminate cells which they encounter that are recognized as being infected
with viruses or other pathogens or are otherwise damaged or dysfunctional through a process referred to as cell lysis, which involves
the release by these killer T cells of perforins and granzymes to compromise the integrity of the target cell’s membrane. Endogenous
pathogens are broken down by mechanisms present in virtually all cells into smaller fragments and presented to CD8 T cells in combination
with an MHC class I molecule. CD4 T cells, also referred to as T helper cells, have limited cytotoxic activity and typically do
not kill infected or dysfunctional cells or eliminate pathogens directly. Instead, they participate in the immune response by providing
signals which activate and orchestrate other types of immune cells to perform these tasks. Professional APCs, such as dendritic cells
and macrophages, process exogenous pathogens and then present small fragments of the degraded pathogen to CD4 T cells in combination
with an MHC class II molecule, through a phenomenon known as cross-presentation, antigens of exogenous origin are coupled with an
MHC Class I molecule to amplify CD8 T cell activity. Antigen cross presentation is of particular importance in the immune system’s
response to cancer.
Genetically
Engineered T Cells
The
ability to enhance the activity of T cells against human cancers through genetic engineering has been among the most significant advances
in cancer therapy in the last decade. Advances in understanding T cells and their role in immunology, and an appreciation of their potential
use to treat cancer, has increased interest in the clinical application of T cells in recent years, with the field of adoptive immunotherapy
attaining increased prominence as a means of enhancing immune control over tumors. Modern molecular biological techniques allow scientists
to introduce genes into human T cells that enhance T cell activity, expand their numbers and infuse them back into the patient from whom
they were originally harvested. We have developed a novel approach to T cell engineering which has enabled us to integrate certain desirable
characteristics of both the innate immune system and the adaptive immune system into a single therapeutic construct intended to optimize
cancer therapy. This novel cellular immunotherapy platform is designed to redirect T cells to eliminate tumors by building in engulfment
pathways that employ phagocytic programs, creating our CER-T cell therapy.
3
Phagocytosis
is a vital cellular process by which a phagocytic cell engulfs and internalizes a target for elimination and is a major mechanism for
the removal of pathogens and unwanted cells to maintain tissue homeostasis. The human body removes billions of cells daily through phagocytic
processes. Phagocytic removal employs specific cell clearance programs and machinery to eliminate target cells. The process is a crucial
part of the innate immune system and is distinct from the adaptive immune response which involves the generation of cytotoxic T cells
to elicit antigen-specific, cytolytic target elimination. To optimize anti-tumor function, we developed CER-T cell therapy to collaboratively
mediate both cytotoxic and phagocytic mechanisms. By leveraging the strength of both immune responses, engulfment has the potential for
more silent and nontoxic cell removal compared to current CAR-T cell therapies. By leveraging both immune responses, we believe CER-T
cell therapy has the potential to eliminate cancer cells more effectively and with fewer side effects than traditional CAR-T cell therapies.
The
recognition of phagocytosis as a therapeutic modality to directly clear cancer cells and initiate anti-tumor T cell immune responses
has fueled interest in effectively engaging phagocytes for use in cancer therapy. Macrophage cell engineering and macrophage-targeting
approaches that enhance cytotoxic, phagocytic and cytokine-mediated anti-tumor function are in development. Early clinical trial data
from therapeutic candidates targeting myeloid inhibitor function has demonstrated the potential to elicit clinical responses. However,
the diverse pro-tumor functions of myelo-monocytic cells may offset these efforts by supporting cancer cell survival, proliferation and
the release of factors that may impede anti-tumor immune responses. Limited in vivo proliferation and manufacturing challenges have also
been hurdles in the development of mononuclear phagocyte-based cellular therapy.
Experimental
evidence demonstrates the ability of CER-T cells to engulf targeted cells, employ cytolytic and non-cytolytic killing mechanisms, and
exhibit pro-inflammatory and antigen processing capabilities that augment the current capabilities of T cell immunotherapy. To that end,
we believe CER-T cell therapy, if approved, may become a component of standard of care treatment regimens, used in combination with both
small molecule therapeutics and biologics including monoclonal antibodies, and CAR-T and high affinity T cell receptor (“TCR”)
T cell therapies to direct robust tumor elimination.
The Increasing
Prominence of CAR-T Technology
Immunotherapy
is a treatment that harnesses the components and mechanics of the immune system to address diseases and disorders. Cellular immunotherapy
is a form of immunotherapy that focuses on modulating or enhancing the activity of different immune cells. One of the more prominent
and promising therapeutic uses of T-cells to emerge has been CAR-T cell technology.
CAR-T
therapy recognizes specific antigens that are present on the surface of tumor cells and destroys them. The concept of CAR-T builds upon
the normal biology of CTLs, whereby naturally occurring receptors serve to activate these cells when a foreign pathogen or cancerous
cell is detected. Conventional CAR-T cell therapy involves the genetic manipulation of a patient’s T cells to enable the expression
by those modified cells of a receptor designed to bind to a specific surface antigen. After the removal of the T cells from the patient’s
blood, a viral vector containing the genetic instructions for the CAR is employed to insert those genes into the genome of the T cell
through a process known as transduction. Aggregated in a single viral vector are the genes encoding for each component of the CAR. Typical
of the prevailing generation of CAR architecture is the inclusion of these components:
The
assembly of these core CAR components is depicted in the schematic presented below to which certain non-coding regulatory sequences may
be used to augment viral gene expression.
4
Delivery
of conventional CAR-T cell therapies involves a single viral vector.
Conventional
CAR-T cell therapies often utilize a lentiviral vector for the delivery of CAR specific genes. Lentiviral particles offer a well-characterized
transduction mechanism and are recognized as efficient and convenient vehicles for gene transfer as they demonstrate broad tropism, or
activity, in a wide array of cell types, and can be used to target quiescent, or non-dividing, cells. In addition, they do not integrate
close to the promoter regions of genes with the frequency of other gene delivery alternatives and lack the immunogenicity of DNA-based
vectors, characteristics which provide for enhanced safety. The use of a lentiviral vector to facilitate ex vivo clinical gene transfer
has been demonstrated to be safe in humans for two decades with no genotoxicity observed in hundreds of patients following gene transfer
into T cells or hematopoietic progenitor cells.
Currently,
six CAR-T cell therapies have been approved by the FDA for the treatment of certain types of hematological cancers. The first two, approved
in 2017, are axicabtagene ciloleucel, sold by Gilead Sciences under the brand name Yescarta, and tisagenlecleucel, sold by Novartis under
the brand name Kymriah. A third CAR-T cell therapy, brexucabtagene autoleucel, which is comparable to Yescarta and sold by Gilead under
the tradename Tecartus, was approved in 2020. Lisocabtagene matraleucel, sold by Bristol Myers Squibb under the brand name Breyanzi,
received FDA approval in February 2021 with Bristol Myers Squibb also receiving approval for idecabtagene vicleucel, sold under
the tradename Abecma, in March of that year. Most recently, Janssen Biotech received FDA approval for ciltacabtagene autoleucel, brand
name Carvykti, to treat adult patients with relapsed or refractory multiple myeloma and which targets the BMCA protein expressed on cancer
cells rather than CD19, the target of the other approved CAR-T cell therapies. Each of these therapies is an autologous therapy and is
made from T cells first collected from the patient, which are then genetically modified and administered back to the same patient. Sales
of CAR-T cell therapies are anticipated to grow rapidly over the next several years and are expected to exceed $10 billion
by 2030. CAR-constructs incorporating alternate immune effector cell types, including NK cells and macrophages, are in earlier stages
of clinical development and have only recently entered clinical trials. To date, no CAR-based therapies that employ NK cells or macrophages
have received FDA approval. There are at present no FDA approved CAR T cell products for AML.
The Limitations
of Current CAR-T Technology
Much
of the excitement of cellular therapy surrounds the curative potential of adoptive transfer of genetically engineered T cells. Adoptively
transferred T cells proliferate upon their engagement with target antigens and represent a form of therapy that can be appropriately
characterized as living and expanding. Efficient targeted killing and tumor elimination may be achieved in a short period of time. However,
multiple barriers limit the efficacy of conventional CAR-T cell therapy. A high rate of side effects often accompany treatment with currently
approved products, especially in those patients with high tumor burdens. In addition, partial responses occur, often associated with
immune escape of the tumor from the TCR or the display by the T cells of an exhaustion phenotype. Moreover, while engineered CAR-T cells
have shown remarkable potential in the treatment of hematological cancers, they have not demonstrated equivalent efficacy in the treatment
of solid tumors. Curative cell therapies for solid tumors currently do not exist and the importance of this limitation is underscored
by the prevalence of solid tumor malignances. The American Cancer Society estimates that solid tumor cancers accounted for more than
1.7 million of the 1.9 million people newly diagnosed with cancer in 2021. Even in hematological malignancies with approved
CAR-T cell therapies, less toxic orthogonal treatment approaches are needed as cure rates for CD19-targeted CAR-T cell therapies do not
exceed 60%.
Challenges
to the use of cellular therapy to address solid tumors often relate to difficulty in developing receptors directed towards targets expressed
in high frequency on cancer cells as well as overcoming the immunosuppressive microenvironments that contribute to ineffective immune
responses. The tumor stroma, made up of a dense fibrotic matrix, often surrounds solid tumors and acts as a physical barrier, which restricts
CAR-T cell access to the tumor. CAR-T cell activity may be further hindered by the tumor microenvironment (“TME”). In the
TME, multiple cell types which drive immunosuppression infiltrate solid tumors, including myeloid-derived suppressor cells, tumor-associated
macrophages, and regulatory T cells. The interaction of these cells and the tumor cells increases the expression of signaling molecules
that enable tumor cell proliferation while dampening the generation of co-stimulatory signals necessary for T cell expansion and persistence.
In addition, TME-associated immune dysfunction may result in a down regulation of MHC class I molecules, limiting proper antigen
presentation and T cell proliferation. Collectively, these attributes of solid tumors enable them to avoid normal immune surveillance.
Increased engagement of the endogenous host response is also an important, if not critical, component of CAR-T cell therapy clinical
success as the recruitment into the tumor of bystander lymphocytes has been observed in tumor biopsies from patients with curative CAR-T
cell therapy. Enhancing the host’s own response to tumor cells offers an important opportunity to improve current CAR T cell responses.
5
CAR-T
recipients may also incur serious adverse events (“SAEs”), perhaps the most prominent of which is cytokine release syndrome
(“CRS”). Believed to be related to the rapid proliferation and activation of T cells upon detection of a target antigen,
severe or life-threatening CRS was noted in a significant number of patients who participated in the registrational trials of FDA-approved
CAR-T therapies. These SAEs can result in patients who receive conventional CAR-T therapy requiring longer hospitalizations and more
intensive medical care. The frequency and severity of observed SAEs was one of the primary reasons that administration of currently approved
CAR-T therapy is restricted to a select number of treatment centers. Moreover, aside from the low-level expression of certain cancer
specific neoantigens, most tumor associated antigens are also found on normal cells which may lead to serious, if not life threatening,
“on-target, off-tumor” toxicities.
We
believe that the preferential attributes engineered into our CER-T cell therapies have the potential to represent a next-generation adoptive
cellular immunotherapy approach and enable us to overcome many of the limitations which hinder the wider application of current CAR-T
technology. The prophagocytic and immunomodulatory properties of CER-T cells are designed to overcome some of the immunosuppressive elements
in many solid tumors. In addition, their anticipated superior antigen presentation properties may enhance a patient’s ongoing immune
response against tumor antigens. In consequence, we envision CER-T therapeutics as having a differentiated mechanism for tumor clearance
that enables the potential for enhanced activity across a broad array of hematological malignancies and solid tumors.
CER-T
Cell Therapy Technology
Distinguishing
our CER-T cell therapy candidate is the integration into a single therapeutic construct of many of the anti-tumor capabilities resident
in both the innate and the adaptive immune systems. We believe the coupling of these functions better emulates normal immune system activity
which may promote enhanced T cell activation, proliferation and durability for more robust elimination of cancerous cells and reduction
in tumor burden.
We
have designed our CER-T constructs to embrace many of the components found in conventional CAR-T cell therapies. The processes and protocols
used to genetically modify a patient’s T cells to produce CAR-T cells are well recognized, as is the use of lentivirus in the manufacture
of these therapies. Accordingly, we have constructed CER-T cell manufacturing processes to be similar to those of CAR-T cells. We expect
to benefit from the well-defined regulatory guidelines established by both U.S. and European regulatory authorities related to CAR-T
cell therapy and its use.
The biological
foundations for CER-T cell therapy
PS
is a component of a cell’s plasma membrane and has a key role in cell removal. Under normal physiological conditions, PS is restricted
to the inner leaflet of the phospholipid bilayer which makes up the plasma membrane of a cell. However, cellular stresses cause the externalization
of PS to the cell surface. Exposure of PS on the outer surface acts as an “eat-me” signal and marks abnormal, stressed and
dying or dead cells for phagocytic clearance. A variety of tumors have been shown to have increased surface PS as a result of altered
plasma membrane regulation. Among hematologic tumors, loss-of-function mutations in the flippase chaperone transmembrane protein 30A
(“TMEM30A”), have been identified in approximately 5% to 11% of patients with diffuse large B cell lymphoma (“DLBCL”)
and among a cohort of newly diagnosed patients, this mutation was correlated with improved response to the standard therapeutic regimen
suggesting the host’s immune elimination of PS positive tumor cells enhances tumor clearance. We are seeking to exploit the presence
of PS expressed on the outer cell surface of both hematological malignancies and solid tumors to create our next generation anti-cancer
agents.
CER-1236:
Our Lead Development Candidate
As
externally oriented PS is present on many cancerous cells regardless of tumor type, we believe a single CER construct may demonstrate
clinical utility in treating an array of cancers. To that end, we have focused our development activities on optimizing the cancer killing
capabilities of a specific CER-T therapeutic design. These efforts have resulted in our lead clinical candidate, CER-1236. In preclinical
studies, we have observed CER-1236 to display attractive functional capabilities and product characteristics, among which are:
● tumor cell phagocytosis;
● enhanced antigen acquisition, processing and presentation;
6
● no evidence of T cell exhaustion despite repeated challenges;
● no observed off-target or off-tumor toxicities;
● well defined and scalable manufacturing protocols.
We
have designed CER-1236 to align with components included in the current generation of conventional CAR-T configurations by fusing the
external domain of TIM-4, a phagocytic receptor, with intracellular signaling domains from T cells and innate immune cells. TIM-4 harbors
endogenous phagocytic capacity through its binding to the pro-phagocytic “eat-me” signal PS. CER-1236’s intracellular
signaling domains, including TLR2/TIR, CD28 and CD3ξ motifs, are designed to augment both TIM-4 mediated phagocytosis and cytotoxic
T cell function. Another similarity between conventional CAR-T therapeutic formats and our CER-T design is the delivery vehicle used
in transduction. As is found in many approved CAR-T therapies, our CER-T technology also employs a lentiviral vector to facilitate gene
delivery to patient-derived T cells. A schematic of the structural elements of CER-1236 is presented below.
Schematic
of CER-1236
Abbreviations:
TIM-4 = ectodomain of the T cell immunoglobulin mucin domain protein 4; TLR2 = toll-like receptor 2; TIR = toll/interleukin-1 receptor.
CER-1236
employs an innovative mechanism of action
CER-1236
is an autologous T cell therapy candidate designed to target PS through the external domain of the prophagocytic receptor TIM-4 protein.
This therapeutic construct was developed to combine adaptive T cell killing activity with phagocytic clearance and antigen presentation
activity to create T cells with enhanced cancer immunotherapy capabilities. The approach builds on the early success of adoptive T cell
transfer, which has demonstrated the ability of T cells to proliferate, traffic, and circulate within both primary and metastatic tumors.
By
enhancing phagocytic clearance and antigen presentation activity and integrating them into T cells, we believe CER-T cells offer the
potential for more effective elimination of cancer cells. The industry’s decades-long experience with engineered T cell use provides
a solid foundation for the development of CER-1236.
As
the target ligand of our initial CER-T cell is not an antigen restricted to only certain tumors, CER-1236 T cells may provide clinical
benefit across multiple tumor types. The functional interaction of CER-1236 T cells is depicted in the illustration presented below.
CER-1236
T cells are designed to harness the power of both the innate and adaptive immune systems
7
CER-1236
expresses the external domain of the prophagocytic receptor TIM-4 which is linked to T cell and innate immune cell intracellular signaling
domains. TIM-4 is normally expressed on subsets of macrophages and dendritic cells and harbors endogenous phagocytic capacity through
its binding to and recognition of PS. The intracellular signaling domains in CER-1236 are designed to trigger T cell cytotoxic function
and enhance TIM-4 mediated phagocytosis. CD3ξ is the signaling component of the TCR and CD28 is a co-stimulatory domain needed for
optimal activation. The TLR2/TIR domain is involved in both innate and adaptive immune responses and activation of TIR further enhances
signaling through both NFξB and the mitogen-activated protein (“MAP”) kinase family, promoting T cell activity and phagocytic
uptake. Both CD28 and CD3ξ signaling domains are incorporated into approved CAR-T cell products. A third generation anti-CD19 CAR-T
cell that incorporates a TLR2/TIR is currently in clinical development.
By
virtue of the TIM-4 engulfment receptor and the intracellular signaling domains, CER-1236 combines attributes of both T cells and phagocytic
cells. In phagocytic cells, such as macrophages and dendritic cells, recognition of the TIM-4 ligand, PS, on the surface of apoptotic
cells by native TIM-4 leads internalization by utilizing integrin coreceptors to activate phagocytic signaling. TIM-4-mediated phagocytosis
depends on activation of the RAC1 GTPase which is similarly targeted by TLR signaling, especially TLR9 and TLR2. However, it has been
shown that deletion of the intracellular portion of TIM-4 is not required for phagocytosis, and therefore the extracellular domain (“ECD”)
of TIM-4 appears to function as a tether during phagocytosis to allow intracellular signaling by other transmembrane phagocytic molecules
with which it associates, such as the integrins which are expressed ubiquitously on T cells. Since CER-1236 contains only the ECD of
TIM-4, binding to PS on tumor cells recruits the cell-surface phagocytosis machinery, and simultaneously directly activates CER-1236
T cells through the intracellular CD3ξ and CD28 costimulatory domains. Phagocytosis and cytokine secretion are further enhanced by
the TLR2/TIR intracellular signaling domain.
In preclinical
studies, CER-1236 empowers T cells with phagocytic and cytotoxic potency
In
an in vitro evaluation of the phagocytic potential of CER-1236, CER-transduced T cells demonstrated robust phagocytosis of PS. CER-1236
T cells were produced by transducing donor T cells using a lentiviral vector encoding for the chimeric receptor CER-1236, yielding a
high percentage of T cells expressing the TIM-4 receptor, in similar CD4:CD8 ratios to untransduced cells. CER-1251 T cells, which express
matching intracellular signaling domains but are unable to bind to PS due to a mutation in the gene encoding for the TIM-4 binding site,
were also produced as a negative control.
PS-coated
agarose beads were prelabeled with pHrodo red, a pH-sensitive dye which displays limited fluorescence at neutral pH but generates significant
fluorescence in acidic pH. The post-phagocytic fusion of phagosomes and lysosomes leads to a drop in pH which can be detected by
pH-sensitive dyes. As is illustrated in the graphic below, CER-1236 T cells co-cultured with PS-coated beads displayed significant phagocytic
activity with up to 60% of CER-T cells acquiring a pHrodo red signal, indicative of bead capture and internalization. By contrast, untransduced
T cells and CER-1251 T cells, with a mutation in the TIM-4 binding site, demonstrated minimal pHrodo red binding.
CER-1236
displays robust, target-specific phagocytic activity
Gene
expression patterns demonstrate the combined cytotoxic and phagocytic functions which reside in the CER-1236 T cell. RNA-sequencing enabled
the interrogation of the transcriptional profile of CER-1236 T cells after stimulation, with defined separation between the CER-1236
activated cells and the untransduced and CER-1251 control T cells. As is presented in the gene expression profile below, over 1,700 genes
were noted to be differentially expressed in CER-1236 stimulated T cells in comparison to CER-1251 stimulated T cells. Among these genes
were those related to pathways with well-known involvement in regulating phagocytosis, genes involved in nucleation of the ARP-WASP complex,
Rho family GTPases, RAC signaling and phagosome formation. Of note, the RhoG subfamily of GTPase has been previously implicated in TCR-driven
phagocytic processes. This aggregate of transcriptional signatures is indicative of the multi-modal immune response elicited by CER-1236
T cells.
8
Phagocytic
and cytotoxic transcriptional signatures demonstrate the plasticity of CER-1236 T cells
CER-1236
T cells were also observed to generate potent anti-cancer responses in cell lines derived from specific hematological malignancies and
solid tumors. Using an MCL cell line that has been modified to constitutively express externalized cell surface PS, MCL cells were co-cultured
with either CER-1236 T cells or untransduced T cells. Notably, CER-1236 T cells eliminated 87% of the MCL cells while the untransduced
cells demonstrated minimal cytotoxic ability. In addition, CER-1236 T cells secreted multiple cytokines, including IFNξ, granzyme
B and TNFξ, all indicative of robust and sustained T cell cytotoxicity. Cytokine secretion was determined to be dependent of binding
to PS, as CER-1251 T cells did not secrete cytokines despite exposure to cell surface PS. Further visual evidence of the cancer-killing
capacity of CER-1236 T cells is illustrated in the staining assays depicted in the graphs presented below. In the assays with no CER-1236
T cells, a significant proliferation of cancer cells was observed, as evidenced by the increase in red staining, while the growth of
cancer cells when exposed to CER-1236 T cells was limited. These results are presented in the graph to the left below.
9
CER-1236
T cells demonstrates potent cytotoxic responses to cancer cells in vitro
Significant
cytotoxic activity of CER-1236 was also noted in an advanced NSCLC cell line which had a mutation in its epidermal growth factor receptor
(“EGFR”) gene, a cancer type accounting for between 10% and 15% of all lung adenocarcinoma cases in persons of European descent
and higher among the Asian population. As is depicted in the above, right graph, while the addition of CER-1236 alone to a NSCLC cell
line which harbors L858R double mutations, demonstrated moderate cancer cell killing activity, the addition of osimertinib, the preferred
tyrosine kinase inhibitor option for first-line treatment of EGFR-mutation positive advanced NSCLC, substantially enhanced CER-1236 T
cell killing in a tyrosine kinase inhibitor-concentration dependent manner. In contrast, HCC827 cells co-cultured with untransduced T
cells displayed minimal changes in cell number as compared to cells incubated in the absence of T cells, at all drug concentrations tested.
Conditional cytokine proliferation was also observed with CER-1236 T cell treatment, with IFNξ levels over 400-fold higher in cancer
cell cultures which used CER-1236 T cells, in contrast to co-cultures which used untransduced T cells. The addition of osimertinib to
co-cultures further increased IFNξ levels by more than two-fold, compared with CER-1236 treatment alone. Similar trends were observed
with TNFξ and Granzyme B levels and increases in osimertinib concentrations led to dose-dependent CER-1236 T cell proliferation. These
results demonstrated that CER-1236 T cell activity could be significantly enhanced by upregulating target expression through concomitant
dosing of standard of care medication.
PS,
a lipid moiety recognized by phagocytic cells as an “eat me” signal, has previously been shown to be aberrantly upregulated
on acute promyelocytic (“APL”) blasts, a subset of AML. To further interrogate phosphatidylserine across other AML subtypes,
we evaluated a panel of primary bone marrow samples and peripheral blood from AML patients. We screened a preliminary panel of primary,
treatment-naïve or on-therapy AML bone marrow and PBMC samples by flow cytometry: (n=5 adverse, n=5 intermediate, n=1 APL, n=1 familial,
n=5 N/A) (Table 1). We observed both high percent (35.5 % ± 21.6) and gMFI of cell surface PS on a range of AML bone marrow samples.
The median MFI of tertiles 1-3 was: T1 n=7, gMFI = 5033; T2 n=8, gMFI = 1873; T3 n=8, gMFI = 611. Of note, the two on-therapy samples
showed high percent and gMFI of cell surface PS, with a patient receiving 5-azacytidine showing 1.8 fold PS gMFI over median. The second
patient receiving TKI therapy showed 3.3 fold PS gMFI over median. Healthy donor samples had much lower cell surface PS, with a mean
gMFI of 582. Circulating AML leukemic blasts were also evaluated for cell surface PS and showed high concordance with BM blasts, with
high levels of cell surface PS compared to healthy donor peripheral blood mononuclear cells (“PBMCs”).
10
Table
1. AML patient characteristics
130781611 Newly Diagnosed none 62 Female White 81.67 Intermediate N/A Normal
11
AML
from bone marrow or PBMC have elevated cell surface “Eat Me” signal
CER-1236
T cells were also observed to generate potent anti-cancer responses against myeloid malignancies. AML is a heterogenous, and aggressive
hematopoietic malignancy characterized by the rapid buildup of immature myeloid cells in the bone marrow and blood. We used AML cell
lines depicted in the graph below, Kasumi-1 and MV4-11 to demonstrate cytotoxic anti-AML responses in co-culture studies with CER-1236.
Similar to in vitro cytotoxicity results observed with B cell malignancy and NSCLC cell lines we show the addition of CER-1236 alone
to AML cell lines demonstrates potent cell killing activity. Kasumi-1 harbors a p53 mutation, marking a subset of unfavorable disease
risk AML patients, while MV4-11 cells carry a FLT-3 mutation, a proliferative AML leukemia subset. Both cell lines co-cultured with untransduced
T cells displayed minimal changes in cell number as compared to cells incubated in the absence of T cells. CER-1236 T cells secreted
multiple cytokines in co-cultures with AML cell lines, including IFNξ, granzyme B and TNFξ, all indicative of robust and sustained
T cell cytotoxicity.
CER-1236
T cells demonstrate robust in vivo elimination of MCL xenografts
The
cancer killing capacity of CER-1236 that was demonstrated in studies involving MCL cell lines was also noted in a mouse xenograft model.
Immune deficient NOD scid gamma (“NSG”) mice were xenografted with the human REC-1 cell line at Day -2 and then treated with
8 mg/kg ibrutinib or vehicle and administered CER-1236 T cells daily from Day -1 to study completion. Administration of 7.5e6 CER-1236
T cells in the presence of ibrutinib resulted in the elimination of REC-1 tumor burden in all 11 of the mice in this treatment cohort.
The administration of CER-1236 T cells in the absence of ibrutinib eliminated the tumors in all nine animals treated with CER-1236 T
cells alone. No tumor growth inhibition was observed in either the vehicle-treated or ibrutinib-treated control groups. Median survival
for mice receiving CER-1236 T cells with or without co-administration of ibrutinib was not reached during the study period. The results
of this study are presented in the charts below.
A
single infusion of CER-1236 T cells eliminates tumors and improves survival
12
The
level of CER-1236 T cells in peripheral blood displayed robust expansion at Day 7, with or without the concomitant administration
of ibrutinib. Animals that received CER-1236 T cells demonstrated an expansion of over 400-fold as compared to Day 2 levels both in the
absence and presence of ibrutinib. High levels of CER-1236 T cells did not persist in the periphery and animals that received CER-1236
T cells showed a greater than 95% contraction in cell count from peak numbers by Day 14 with subsequent CER-T cell expansion likely prompted
by residual tumor cell encounters. CER-1236 T cells also maintained robust proliferative capacity despite repeated antigen challenges
with no evidence of T cell exhaustion noted. These findings are illustrated in the following charts.
A
single infusion of CER-1236 T cells generated rapid cell expansion across repeated challenges.
CER-1236
demonstrates in vivo tumor clearance in NSCLC adenocarcinoma xenograft.
We
envisioned that the simultaneous exposure to both osimertinib and CER-1236 would lead to synergistic in vivo anti-tumor responses. HCC827
NSCLC cells were inoculated into the flanks of NSG mice. Once established, the mice were dosed with a short course of the EGFR inhibitor
osimeretinib to prime PS antigen on tumors and administered 2.5e6 CER-1236 T cells. Treatment groups that received the EGFR inhibitor
alone, after initial tumor regression, developed progressive disease. In contrast, animals infused with CER-1236 T cells demonstrated
potent anti-tumor responses in the presence of osimertinib. CER-1236 T cells expanded rapidly in the blood, with the highest expansion
observed in the osimertinib-treated cohorts. Importantly, no evidence of organ toxicity or weight loss was observed with increases in
body weight recorded in all groups over the course of the study. Analysis of the tumors post-infusion indicated extensive infiltration
of T cells compared to untransduced controls.
CER-1236
T cells infused to Osimertinib dosed animals showed higher levels of T cell expansion
We
believe that the preclinical models of AML, MCL, ovarian cancer and EGFR-mutation positive NSCLC demonstrate the ability of CER-1236
T cells to induce collaborative innate-adaptive anti-tumor immune responses in both in vitro and in vivo studies. Moreover, concurrent
treatment with standard-of-care therapeutics for each of these indications increases target ligand, conditionally bolstering CER-1236
T cell function to augment anti-tumor activity. Additionally, in antigen presentation assays, activated CER-1236 T cells exhibited superior
cross-presentation ability relative to conventional T cells, triggering specific TCR-T cell responses in an MHC class I and TLR-2
dependent manner, overcoming the limited antigen presentation capabilities of conventional T cells. These results indicate that CER-1236
T cells have the potential to achieve optimal tumor control by eliciting both cytotoxic effects and mediated cross-priming.
13
CER-1236
T cells did not elicit safety signals in preclinical studies
Importantly,
no evidence of toxicity was observed after the single administration of 7.5e6 CER-1236 T cells. No incidence of anemia, thrombocytopenia,
neutropenia or coagulation abnormalities were recorded. Complete blood count, prothrombin time, and partial thromboplastin time were
measured in blood taken at the peak of T cell expansion and after T cell contraction. Hematologic indices, including hemoglobin/hematocrit,
platelets and neutrophils remained stable throughout the study. Prothrombin and partial thromboplastin time were unaffected at the peak
of T cell expansion and after contraction. None of the animals experienced weight loss, morbidity or unexpected mortality. CER-1236 T
cells showed a restricted pattern of tissue distribution similar to other T cell products and did not result in tissue damage with no
histological abnormalities observed in any organ evaluated, including heart, lung, liver, kidney and brain.
CER-1236
Clinical Development Strategy
Based
on the extensive preclinical data that we have assembled regarding the use of CER-1236 T cells to combat cancer, we currently anticipate
filing an initial IND in 2024 to commence clinical trials with our initial treatment target being patients suffering from relapsed, or
refractory AML as well as certain B cell lymphomas with unmet medical need. We subsequently intend to expand the clinical development
of CER-1236 to include solid tumors such as NSCLC and ovarian cancer. We expect these clinical trials to evaluate the safety, the potential
therapeutic utility and applicable dose of CER-1236. In addition, we anticipate that these clinical trials may provide insight into the
possible use of CER-1236 to treat an array of hematologic and solid tumors. We believe this drug candidate has the potential to be a
therapy for the unmet needs of targeted indications, if approved, and by leveraging the innate immune system’s phagocytic capabilities,
could be differentiated by its safety, tolerability, efficacy and clinical benefit over current therapeutic approaches, which have been
observed in preclinical studies. None of the abovementioned statements regarding any of our products in development are intended to be
a prediction or conclusion of efficacy. No clinical trials on our product candidates have commenced so no conclusions relating to such
attributes can be made.
Disease
background
Acute
Myeloid Leukemia
Approximately, 20,000
patients in the U.S. are diagnosed with AML annually. In adults, AML, characterized by the rapid buildup of abnormal myeloid cells in
the bone marrow and blood, is the most common acute leukemia worldwide. In children and young adults under 20 years old AML
comprises 74% of acute leukemia cases. Despite the many available treatments for AML, prognosis for patients remains poor. There are
several molecular alterations of AML that make it difficult to treat with individual therapies. The process of categorizing the
disease state as well a patient’s degree of “fitness” for toxic chemotherapeutic regimens is the subject of
ongoing debate and discussion. Moreover, the development of engineered T cell therapies for AML has proven difficult in part due to
the identification of suitable target antigens.
B cell
lymphoma
B
cell lymphomas include patients diagnosed with Hodgkin’s disease and those diagnosed with non-Hodgkin’s lymphoma (“NHL”).
In the U.S., the American Cancer Society estimates that in 2023, approximately 8,800 cases of Hodgkin’s lymphoma and 81,000 cases
of NHL will be diagnosed, with the number of new diagnoses expected to increase annually. Aggressive B cell lymphomas are a heterogenous
group of cancers that arise from B lymphocytes in various stages of development that make up part of the immune system. Included in this
group is DLBCL and its variants, MCL, Burkitt lymphoma and B cell lymphoblastic lymphoma. There are an estimated 15,000 patients receiving
second or third-line therapies for refractory or recurrent NHL.
We
intend to focus the initial development of CER-1236 on the lymphoid malignancies MCL and CLL, a malignancy of mature B cells. MCL is
a rare and difficult-to-treat B cell lymphoma with a relapsing and remitting clinical course. Most cases of MCL are diagnosed with advanced-stage
disease. Extranodal involvement is common, including involvement of the bone marrow, the gastrointestinal tract and peripheral blood
which can result in significant morbidity. MCL accounts for approximately 6% of all cases of NHL diagnosed annually. CLL tends to progress
slowly, with a benign phase followed by a terminal phase marked by progressive disease resistant to treatment. The course of CLL is complicated
by immune dysfunction, which leads to an increased risk of infection, autoimmune complications and other cancers. An estimated 19,000
new cases of CLL are diagnosed annually in the U.S. and CLL is the cause of approximately 4,300 deaths per year.
Current
therapies and their limitations
Until
recent years, AML has been treated with decades-old combination chemotherapy regimens, including cytarabine and anthracycline. This
regimen has about a 70-80% complete response (“CR”) rates of adults younger than 60 years and 40-60% of fit
adults older than 60 years old. For those eligible for the chemotherapy regimen and experiencing a CR, many patients with adverse
features (70%) undergo allogeneic HSCT which, in some patients are “curative.” Unfortunately, a significant proportion (up
to 50%) of AML patients are over the age of 65 and are “unfit” for intensive chemotherapy, requiring different treatment
approaches for medically unfit patients. The treatment landscape for older unfit adults with AML fundamentally changed with the recent
availability of new drugs, in particular the oral B-cell lymphoma 2 inhibitor venetoclax. Venetoclax is used in conjunction with azacytidine
to treat these patients, with a complete response rate ~65%. However the majority of adult patients with AML experience relapse despite
initially attaining CR; a venetoclax-based doublet therapy for medically less-fit adults carries a median survival of ~14.7 months
and 4-year survival of less than 20%. The prognosis for patients who are refractory to or relapse after frontline azacitidine venetoclax
is dismal with median overall survival of 2.4 months, making this an area of high unmet need. Such patients who do not respond to
frontline therapy with azacitidine or venetoclax, and the subset who do not respond to targeted therapies, e.g., IDH1/2 inhibitors, are