Trial Information

The Development of Human Immunologic Assays Specific to Folate Receptor Antigen

Incidence of Ovarian Cancer

Ovarian cancer is the first mortality rate of gynecologic malignancies with an overall
5-year survival rate of only 20-30%. It became a more and more important disease in recent
years and the incidence of ovarian cancer also increased in recent years in Taiwan. The lack
of symptoms, difficulties in early diagnosis, insufficient accurate tumor markers, and lack
of information about ovarian tumor biology contribute to the poor prognosis in ovarian
cancer patients. The prognostic parameters for ovarian carcinomas are tumor stage,
histologic subtype, degree of malignancy, and residual tumor after surgical treatment.
However, these factors present an incomplete picture of the tumor biology of ovarian cancer
and are frequently interrelated. Thus, the identification of new biologic factors predictive
of individual disease course and prognosis would be extremely useful. From the
above-mentioned data, ovarian cancer is indeed a disease that should be respected, however,
there was very little research work focusing on it in Taiwan.

Treatment of ovarian cancer

Epithelial ovarian cancer (EOC) and extraovarian Müllerian carcinoma are similar pathologic
entities that share a preference for peritoneal cavity involvement. The spread pattern of
these tumors presents a challenge and unique opportunities for immunotherapy. Patients with
EOC who have stage I disease (localized to ovaries) after optimal surgical staging, have a
5-year survival rate of 90%, with no significant change at 10 years. In contrast, patients
with spreading beyond the ovaries have median survival rates that decrease to < 10% for
patients with bulky residual disease after surgery and treatment with platinum-based
combination chemotherapy. A randomized trial of first-line chemotherapy in patients with EOC
with residual masses larger than 1 cm after initial surgery, showed a median survival period
of 38 months for cisplatin/paclitaxel, significantly greater than 24 months for the
cisplatin/cytoxan treatment arm. In an interim analysis of an equivalency trial, survival
after carboplatin/paclitaxel was not worse than cisplatin/paclitaxel. Even though early
diagnosis is an important goal of ongoing clinical research efforts, it is unclear whether
advanced EOC starts as a multicentric process involving the ovaries and the peritoneal
surface. It is now established that hereditary factors contribute to the development of EOC.
Germline BRCA1 and BRCA2 mutations account for approximately 10% of all EOC. In a woman with
a BRCA 1 or 2 mutation, lifetime risk for ovarian cancer ranges from 16%-44%. With the
commercial availability of genetic testing for BRCA1 and BRCA2, more women are being
identified as being at high risk for ovarian cancer. There are no clear guidelines on cancer
prevention for these individuals. Although prophylactic oophorectomy is a reasonable option
for women who have completed childbearing, these women are still at risk for developing
peritoneal cancer. Clearly, other options for prevention are needed.

Immunotherapy for ovarian cancer

In developing effective immune-based strategies for EOC, there should be a distinction
between preventative and therapeutic approaches. It is anticipated that immunoprevention
(immunoprophylaxis) will be developed for women who are at an increased risk for the
development of EOC. In contrast, immunotherapy would be used as an adjuvant to surgery or in
combination with chemotherapy or other biologics as chemoimmunotherapy or
biochemoimmunotherapy. Patients with undetectable disease after being restaged after
chemotherapy could be considered for immunotherapy with the presumption that a majority does
in fact have micrometastases. Development of effective immune-based concepts for prevention
or treatment of EOC will require an understanding of tumor-immunology principles, mechanisms
of action of the expanding array of immune modulating molecules, identification and
characterization of tumor antigens, and determination of the microenvironment factors that
could impact on the different immune-effector mechanisms. The clinical researcher has been
provided with many immune directed agents, but progress on their integration into standard
therapies has been somewhat slow.

Folate receptor

The folate receptor (FR), a 38 kDa membrane glycoprotein, is represented by a homologous
family of glycoproteins, two of which (FR-a and FR-b) are attached to the cell surface by a
glycosyl-phosphatidylinositol anchor; the third isoform (FR-) and its truncated version
(FR-9) are constitutively secreted because of a lack of an efficient signal for
glycosyl-phosphatidylinositol modification. FR-a is expressed in some normal epithelial
cells and is elevated in certain carcinomas, whereas FR-b is a myeloid differentiation
marker and is elevated in some nonepithelial malignancies. FR-9 is expressed in
hematopoietic tissues. At present, FR is a major focus as a tumor target for multiple
experimental approaches in cancer therapy. One novel approach uses bifunctional antibodies
to target T cells to the FR on the surface of ovarian carcinoma cells. Selective growth
inhibition of the tumor cells was obtained by this approach. The chimeric antibodies, which
bind to both FR and either CD3 or CD28, produced impressive results in a xenogeneic model
and in patients with advanced ovarian cancer. Similarly, a chimeric molecule consisting of a
single-chain, Fv of anti-FR antibody and interleukin 2 was effective in inhibiting tumor
growth in vivo. Alternatively, folic acid conjugates of single chain anti-T-cell receptor
antibody could mobilize T-cell response against FR-rich tumors. Taking advantage of the
nondestructive nature of FR-mediated internalization of folate-coupled macromolecules,
cytotoxins such as momordin, Pseudomonas exotoxin, and maytansinoids were shown to produce
selective killing of FR-rich cells. Furthermore, the toxicity of such conjugates was
dependent upon receptor density on the cell surface. Folate-conjugated radiopharmaceuticals
also appear to offer a means of tumor imaging/radiation therapy. Folate-coated liposomes
were shown to selectively target FR-rich tumor cells, and selective killing of the malignant
cells was obtained by encapsulating doxorubicin in the liposomes. By a similar strategy, it
was possible to deliver antisense oligonucleotides against the epidermal growth factor
receptor to FR-rich tumor cells. Furthermore, selective targeting of an adenoviral vector to
FR-rich tumor cells has been achieved in the presence of an antibody to ablate the
endogenous viral tropism. Finally, several studies have shown that FR, when expressed at
high levels, could offer the preferred uptake route of novel classes of antifolate drugs
that target glycineamide ribonucleotide formyltransferase and thymidylate synthase. A
soluble form of FR has been detected in the serum and ascites of patients with ovarian
cancer. The FR has been reported to be selectively overexpressed in 90% of non-mucinous
ovarian carcinomas and in some other malignant tissues. The receptor has only been detected
in a few other normal cell types, but not in normal ovarian surface epithelium. It seems
that FR could be a potential target antigen for the immunotherapy of ovarian cancer.

Epitopes of folate receptor for human haplotype

The incidence of HLA-A2 haplotype is over 50% in the Western countries. The incidence of
HLA-A2 haplotype is around 30% in Taiwan. The specific epitopes of the folate receptor in
the HLA-A2 haplotype have been identified. They are E39 (FR, 191-199) EIWTHSTKV and E75 (FR,
245-253) LLSLALMLL, respectively. It seems that folate receptors could be a target antigen
for the immunotherapy of ovarian cancer.

Our research team has focused on the development of a cancer vaccine and immunotherapy for
several years. Our laboratory facilities have also been set up to evaluate the human
immunologic assays for human papilloma virus type 16 E7 antigen by the grant supported from
National Taiwan University Hospital. It is very important to set up various folate
receptor-specific immunologic assays of human beings to evaluate the effect of cancer
vaccine or immunotherapy for ovarian cancer in future clinical trials. So we would like to
provide this proposal to address the development of folate receptor-specific immunologic
assays in human beings. There are several aims in this project:

1. to develop and utilize assays to measure CTLs to folate receptors and,

2. to evaluate the folate receptor-specific immunologic responses between normal controls
and ovarian cancer patients.