MAbs have emerged as the fastest growing class of pharmaceutical products in recent years and in 2007 achieved worldwide sales of approximately $25 billion.

Despite exquisite specificity to their targets, many antibodies, particularly when used in a monotherapy setting, exhibit excellent therapeutic response in some patients but perform poorly in many others. This variability is due to the fact that patients are genetically and immunologically different. As a result, there is a significant market pressure to develop an effective method to stratify patients into potential responders versus non-responders.

By incorporating the 3×3 Matrix™ into the antibody engineering platform, PIKAMAB develops customized MAb therapies for each patient group in the matrix such that ADCC and ADCC-related mechanisms of action are fully enabled by effectively engaging the necessary immune cells such as the NK-cells, macrophages, and neutrophils. This patient and group-specific Fc engineering allows for effective engagement of IgGs to FcGR-3A and FcGR-2A in immune cells (NK-cells, macrophages, and neutrophils). For example, the Group-9 IgG construct in the matrix has the optimized Fc for FcGR-3A (F¹⁵⁸) and FcGR-2A (R¹³¹) receptors. Thus, effective ADCC and phagocytosis-mediated clearance of immune complexes can be achieved in each patient group.    

PIKAMAB captures these inventions in its theragnostic and MAb drug development programs. The theragnostic products, ADCC Therasight™ and Lupus Therasight™, stratify patients into nine distinct groups through a proprietary 3×3 Matrix™ and provide critical treatment guidance to biopharma companies, regulatory agencies, payers, and physicians.

• PIKAMAB’s theragnostic products that include ADCC Therasight™ and Lupus Therasight™ provide therapeutic guidance to marketed MAb therapies.
• ADCC is the major mechanism of action of several marketed MAbs (FcGR-3A and FcGR-2A receptors are directly involved in mediating ADCC).
• In several autoimmune diseases and inflammatory disorders, immune complexes are cleared in kidney, liver, and spleen by macrophage-mediated phagocytosis: FcGR-3A and FcGR-2A receptors play a central role in phagocytosis.

Neutrophils, macrophages, and natural killer (NK) cells play several key roles in immune functions, which include:

• Antibody-dependent cellular cytotoxicity (ADCC),
• Antibody-dependent cellular phagocytosis (ADCP),
• NK-cell mediated apoptosis,
• Clearance of immune complexes by phagocytosis, and
• Antibody-dependent cellular viral inhibition (ADCVI)

Selective engagement of these immune cells with antibodies, immune complexes (ICs), and other cells is mediated by Fc gamma receptors (FcGRs: 3A, 2A, 2B, 3B) expressed on the surface of these immune cells. Remarkably, single nucleotide substitutions in critical regions of these FcGR genes lead to single amino acid substitutions in these receptors. The occurrence of these polymorphisms profoundly alters the receptor biology and, in turn, the way these immune cells function. As a consequence, these functional polymorphisms in FcGRs play transformative roles in processes mediated by immune cells and have profound implications in oncology, autoimmune diseases, and inflammatory disorders.

In oncology, antibody-dependent cellular cytotoxicity (ADCC) is a major mechanism by which several therapeutic antibodies exert their therapeutic effect. NK-cells, macrophages, and neutrophils play critical roles in cancer cell killing of both solid tumors (for example, breast cancer or colorectal cancer) and hematological malignancies (for example, B-cell non-Hodgkins lymphoma).

Once an ADCC-capable antibody is bound to its tumor target cell, its natural cell killing and clearance mechanism is triggered by the Fc region of the antibody (its ‘tail’) upon binding to one of the Fc gamma receptors (FcGRs) located on immune cells such as NK cells, macrophages and neutrophils. This results in activation of the ADCC mechanism leading to tumor cell killing. The FcGR-3A is found on NK-cells while FcGR-2A is present on neutrophils. Both receptors are present on macrophages.

Cell-specific glycosylation differences have been observed in FcGR-3A expressed on NK-cells and macrophages, and such subtle variations are known to dramatically influence the ADCC capacities. Even though both FcGR-3A and FcGR-2A receptors are present, FcGR-2A receptor seems to play a crucial role in macrophage-mediated processes. Therefore both receptors, FcGR-3A and FcGR-2A, are immunologically relevant to achieve meaningful therapeutic outcomes for an ADCC-capable antibody.

In 1997, Rituximab (Rituxan®) was approved for the treatment of refractory or relapsed low-grade B-cell non-Hodgkin’s lymphoma (B-NHL) and has becoming a mainstay of treatment for low-grade B-NHL. To date, more than 1 million patients have been treated worldwide and the net sales of Rituxan reached $5.2 billion in 2007. Despite this remarkable commercial success, the therapeutic response to rituximab varies significantly in the patient population. Overall response rates (complete and partial responders combined) of ~40-70% have been reported under various clinical trial settings that included chemotherapy and/or radiation therapy with rituximab.

Based on 10 years of rituximab monotherapy study in B-NHL patients, Professor Ron Levy at Stanford University School of Medicine discovered that the therapeutic response rate of rituximab in patients can be unequivocally correlated to two specific polymorphisms in the receptors FcGR-3A (VF¹⁵⁸), and FcGR-2A (HR¹³¹). Systematic, retrospective analyses led to this seminal finding that both receptors independently and together dictate the therapeutic response rate and PFS, and that ADCC is the major mechanism of action by which rituximab elicits clinical response in B-NHL patients. In addition, this was the first study to discover the role for the FcGR-2A receptor and its HR¹³¹>polymorphism in dictating the therapeutic response rate and PFS of an antibody in an oncology indication (B-NHL). This study, however, only classified patients into three groups: excellent (VV¹⁵⁸and HH¹³¹), moderate (either VV¹⁵⁸or HH¹³¹), and poor (others).

PIKAMAB’s proprietary breakthrough, the 3×3 Matrix™, stratifies patients into nine distinct groups based on the FcGR-3A (VF¹⁵⁸) and FcGR-2A (HR¹³¹) polymorphisms. Each patient group is immunologically distinct from any other group in the matrix. Patients with VV158 and HH131 FcGR polymorphisms (Group-1; ~5% of the patient population) are expected to have excellent response rates to treatment and longer PFS. Patients with other polymorphic patterns (F and R carriers) in their FcGRs are expected to have moderate or poorer response rate and shorter PFS. Accordingly, patients in Group-9 in the 3×3 Matrix™, defined as patients having FF¹⁵⁸in FcGR-3A and RR¹³¹in FcGR-2A, are expected to have the worst outcome.

Furthermore, PIKAMAB’s stratification goes beyond DNA/RNA-based genotyping of patients by generating relevant, patient-specific functional information. In conjunction with our proprietary 3×3 Matrix™, these data provide critical theragnostic information relevant to patient outcomes.

In recent years, the role for FcGR-3A and FcGR-2A polymorphisms in treatment outcomes and PFS has been established in patients treated with MAb therapies such as trastuzumab (Herceptin^®), and cetuximab (Erbitux^®).

Representative Publications:

Rituximab:
Two immunoglobulin G fragment C receptor polymorphisms independently predict response to rituximab in patients with follicular lymphoma. (N=87 patients)
Weng and Levy,J. Clin. Oncology21: 3940-3947. (2003)                

Cetuximab:
Impact of FcGRIIa-FcGRIIIa polymorphisms and KRAS mutations on the clinical outcome of patients with metastatic colorectal cancer treated with cetuximab plus irinotecan. (N=69 patients)
Bibeau et al.,J. Clin. Oncology27: 1122-1129. (2009)

rastuzumab:
Immunoglobulin G fragment C receptor polymorphisms and clinical efficacy of trastuzumab-basedtherapy in patients with HER-2/neu–positive metastatic breast cancer. (N=54 patients)
Musolino et al.,J. Clin. Oncology26:1789-1796. (2008)

Alemtuzumab:
CD52 is expressed on human mast cells and is a potential therapeutic target in Waldenström’s macroglobulinemia and mast cell disorders.
Santos et al.,Clinical Lymphoma & Myeloma6:478-483. (2006)

Autoimmune Diseases

In a variety of autoimmune diseases (such as systemic lupus erythematosus (SLE), lupus nephritis, rheumatoid arthritis, systemic vasculitis, Sjögren’s syndrome, Wegener’s granulomatosis etc.), the clearance of immune complexes (ICs; antigen-antibody complexes) is mediated by the mononuclear phagocyte system of the liver and spleen macrophages as well as the mesangial cells in kidneys. The receptors FcGR-2A and FcGR-3A are directly involved in the clearance of ICs and, as such, this clearance is directly correlated to the allelic polymorphisms in these receptors. Patients with RR131 polymorphism in FcGR-2A and FF¹⁵⁸polymorphism in FcGR-3A tend to have poor clearance, which leads to deposition and accumulation of ICs in tissues and organs. The susceptibility, progression, and severity of pathophysiological symptoms in SLE and lupus nephritis patients may be directly correlated with FcGR polymorphisms.

Lupus Therasight™ can provide critical treatment guidance to drug developers, regulatory agencies, payers, and physicians. For instance, in SLE and lupus nephritis, patients with increased risk of renal disease can be identified and then treated prophylactically and aggressively before substantial kidney damage has occurred. In addition, such systematic patient stratification will also lead us to the development of customized therapies.

Representative Publications:

FcGR-2A alleles are heritable risk factors for lupus nephritis in African Americans. (N=257 SLE patients)
Salmon et al.,J. Clin. Investigation97:1348-1354. (1996)

FcG receptor polymorphisms in systemic lupus erythematosus. (N=230 SLE patients)
Dijstelbloem et al.,Arthritis & Rheumatism43:2793-2800. (2000).

Genetic linkage and association of FcGR-3A (CD16A) on Chromosome 1q23 with human systemic lupus erythematosus. (N=438 SLE patients)

Edberg et al.,Arthritis & Rheumatism46:2132-2140. (2002)

Identification of IgG subclasses and C-Reactive Protein in lupus nephritis (N=80 patients)

Zuniga et al.,Arthritis & Rheumatism48:460-470. (2003)

Decreased transcription of the human FcGR-2B gene mediated by the -343 G/C propmoter polymorphism and association with systemic lupus erythematosus. (N=391 ethnicity agnostic; N=190 European Americans)

Blank et al.,Human Genetics117:220-227. (2005)

Emerging Market

The blockbuster antibody market, currently led by MAbs such as Rituxan, Remicade, and Herceptin, is projected to exceed $20 billion in sales in 2009. Currently, there are over 200 antibody product candidates in clinical development.

PIKAMAB is planning to initiate an antibody engineering R&D efforts in India to operationalize the generation of these stratified MAb products. PIKAMAB is also in the process of establishing a CLIA-certified testing facility in the U.S. to commercialize theragnostic products. In the second phase, we plan to expand theragnostic operations into Europe and Asia.